CXCR4 and CXCL12 are inversely expressed in colorectal cancer cells and modulate cancer cell migration, invasion and MMP-9 activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brand, Stephan; Dambacher, Julia; Beigel, Florian

2005-10-15

Colorectal cancer (CRC) is characterized by a distinct metastatic pattern resembling chemokine-induced leukocyte trafficking. This prompted us to investigate expression, signal transduction and specific functions of the chemokine receptor CXCR4 in CRC cells and metastases. Using RT-PCR analysis and Western blotting, we demonstrated CXCR4 and CXCL12 expression in CRC and CRC metastases. Cell differentiation increases CXCL12 mRNA levels. Moreover, CXCR4 and its ligand are inversely expressed in CRC cell lines with high CXCR4 and low or not detectable CXCL12 expression. CXCL12 activates ERK-1/2, SAPK/JNK kinases, Akt and matrix metalloproteinase-9. These CXCL12-induced signals mediate reorganization of the actin cytoskeleton resulting inmore » increased cancer cell migration and invasion. Moreover, CXCL12 increases vascular endothelial growth factor (VEGF) expression and cell proliferation but has no effect on CRC apoptosis. Therefore, the CXCL12/CXCR4 system is an important mediator of invasion and metastasis of CXCR4 expressing CRC cells.« less

Estrone Sulfate Transport and Steroid Sulfatase Activity in Colorectal Cancer: Implications for Hormone Replacement Therapy.

PubMed

Gilligan, Lorna C; Gondal, Ali; Tang, Vivien; Hussain, Maryam T; Arvaniti, Anastasia; Hewitt, Anne-Marie; Foster, Paul A

2017-01-01

Hormone replacement therapy (HRT) affects the incidence and potential progression of colorectal cancer (CRC). As HRT primarily consists of estrone sulfate (E 1 S), understanding whether this conjugated estrogen is transported and metabolized in CRC will define its potential effect in this malignancy. Here, we show that a panel of CRC cell lines (Colo205, Caco2, HCT116, HT-29) have steroid sulfatase (STS) activity, and thus can hydrolyze E 1 S. STS activity is significantly higher in CRC cell lysate, suggesting the importance of E 1 S transport in intracellular STS substrate availability. As E 1 S transport is regulated by the expression pattern of certain solute carrier organic anion transporter polypeptides, we show that in CRC OATP4A1 is the most abundantly expressed transporter. All four CRC cell lines rapidly transported E 1 S into cells, with this effect significantly inhibited by the competitive OATP inhibitor BSP. Transient knockdown of OATP4A1 significantly disrupted E 1 S uptake. Examination of estrogen receptor status showed ERα was present in Colo205 and Caco2 cells. None of the cells expressed ERβ. Intriguingly, HCT116 and HT29 cells strongly expressed the G protein coupled estrogen receptor (GPER), and that stimulation of this receptor with estradiol (E 2 ) and G1, a GPER agonist, significantly ( p < 0.01) increased STS activity. Furthermore, tamoxifen and fulvestrant, known GPER agonist, also increased CRC STS activity, with this effect inhibited by the GPER antagonist G15. These results suggest that CRC can take up and hydrolyze E 1 S, and that subsequent GPER stimulation increases STS activity in a potentially novel positive feedback loop. As elevated STS expression is associated with poor prognosis in CRC, these results suggest HRT, tamoxifen and fulvestrant may negatively impact CRC patient outcomes.

Coding variants in NOD-like receptors: An association study on risk and survival of colorectal cancer.

PubMed

Huhn, Stefanie; da Silva Filho, Miguel I; Sanmuganantham, Tharmila; Pichulik, Tica; Catalano, Calogerina; Pardini, Barbara; Naccarati, Alessio; Polakova-Vymetálkova, Veronika; Jiraskova, Katerina; Vodickova, Ludmila; Vodicka, Pavel; Löffler, Markus W; Courth, Lioba; Wehkamp, Jan; Din, Farhat V N; Timofeeva, Maria; Farrington, Susan M; Jansen, Lina; Hemminki, Kari; Chang-Claude, Jenny; Brenner, Hermann; Hoffmeister, Michael; Dunlop, Malcolm G; Weber, Alexander N R; Försti, Asta

2018-01-01

Nod-like receptors (NLRs) are important innate pattern recognition receptors and regulators of inflammation or play a role during development. We systematically analysed 41 non-synonymous single nucleotide polymorphisms (SNPs) in 21 NLR genes in a Czech discovery cohort of sporadic colorectal cancer (CRC) (1237 cases, 787 controls) for their association with CRC risk and survival. Five SNPs were found to be associated with CRC risk and eight with survival at 5% significance level. In a replication analysis using data of two large genome-wide association studies (GWASs) from Germany (DACHS: 1798 cases and 1810 controls) and Scotland (2210 cases and 9350 controls) the associations found in the Czech discovery set were not confirmed. However, expression analysis in human gut-related tissues and immune cells revealed that the NLRs associated with CRC risk or survival in the discovery set were expressed in primary human colon or rectum cells, CRC tissue and/or cell lines, providing preliminary evidence for a potential involvement of NLRs in general in CRC development and/or progression. Most interesting was the finding that the enigmatic development-related NLRP5 (also known as MATER) was not expressed in normal colon tissue but in colon cancer tissue and cell lines. Future studies may show whether regulatory variants instead of coding variants might affect the expression of NLRs and contribute to CRC risk and survival.

miR-34a mediates oxaliplatin resistance of colorectal cancer cells by inhibiting macroautophagy via transforming growth factor-β/Smad4 pathway.

PubMed

Sun, Chen; Wang, Fu-Jing; Zhang, Hao-Gang; Xu, Xun-Zheng; Jia, Rui-Chun; Yao, Lei; Qiao, Peng-Fei

2017-03-14

To investigate whether microRNA (miR)-34a mediates oxaliplatin (OXA) resistance of colorectal cancer (CRC) cells by inhibiting macroautophagy via the transforming growth factor (TGF)-β/Smad4 pathway. miR-34a expression levels were detected in CRC tissues and CRC cell lines by quantitative real-time polymerase chain reaction. Computational search, functional luciferase assay and western blotting were used to demonstrate the downstream target of miR-34a in CRC cells. Cell viability was measured with Cell Counting Kit-8. Apoptosis and macroautophagy of CRC cells were analyzed by flow cytometry and transmission electron microscopy, and expression of beclin I and LC3-II was detected by western blotting. Expression of miR-34a was significantly reduced while expression of TGF-β and Smad4 was increased in CRC patients treated with OXA-based chemotherapy. OXA treatment also resulted in decreased miR-34a levels and increased TGF-β and Smad4 levels in both parental cells and the OXA-resistant CRC cells. Activation of macroautophagy contributed to OXA resistance in CRC cells. Expression levels of Smad4 and miR-34a in CRC patients had a significant inverse correlation and overexpressing miR-34a inhibited macroautophagy activation by directly targeting Smad4 through the TGF-β/Smad4 pathway. OXA-induced downregulation of miR-34a and increased drug resistance by activating macroautophagy in CRC cells. miR-34a mediates OXA resistance of CRC by inhibiting macroautophagy via the TGF-β/Smad4 pathway.

Bone morphogenetic protein and Notch signalling crosstalk in poor-prognosis, mesenchymal-subtype colorectal cancer.

PubMed

Irshad, Shazia; Bansal, Mukesh; Guarnieri, Paolo; Davis, Hayley; Al Haj Zen, Ayman; Baran, Brygida; Pinna, Claudia Maria Assunta; Rahman, Haseeb; Biswas, Sujata; Bardella, Chiara; Jeffery, Rosemary; Wang, Lai Mun; East, James Edward; Tomlinson, Ian; Lewis, Annabelle; Leedham, Simon John

2017-06-01

The functional role of bone morphogenetic protein (BMP) signalling in colorectal cancer (CRC) is poorly defined, with contradictory results in cancer cell line models reflecting the inherent difficulties of assessing a signalling pathway that is context-dependent and subject to genetic constraints. By assessing the transcriptional response of a diploid human colonic epithelial cell line to BMP ligand stimulation, we generated a prognostic BMP signalling signature, which was applied to multiple CRC datasets to investigate BMP heterogeneity across CRC molecular subtypes. We linked BMP and Notch signalling pathway activity and function in human colonic epithelial cells, and normal and neoplastic tissue. BMP induced Notch through a γ-secretase-independent interaction, regulated by the SMAD proteins. In homeostasis, BMP/Notch co-localization was restricted to cells at the top of the intestinal crypt, with more widespread interaction in some human CRC samples. BMP signalling was downregulated in the majority of CRCs, but was conserved specifically in mesenchymal-subtype tumours, where it interacts with Notch to induce an epithelial-mesenchymal transition (EMT) phenotype. In intestinal homeostasis, BMP-Notch pathway crosstalk is restricted to differentiating cells through stringent pathway segregation. Conserved BMP activity and loss of signalling stringency in mesenchymal-subtype tumours promotes a synergistic BMP-Notch interaction, and this correlates with poor patient prognosis. BMP signalling heterogeneity across CRC subtypes and cell lines can account for previous experimental contradictions. Crosstalk between the BMP and Notch pathways will render mesenchymal-subtype CRC insensitive to γ-secretase inhibition unless BMP activation is concomitantly addressed. © 2017 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Knockdown of stromal interaction molecule 1 inhibits proliferation of colorectal cancer cells by inducing apoptosis.

PubMed

Yang, Dong; Dai, Xiaoyu; Li, Keqiang; Xie, Yangyang; Zhao, Jianpei; Dong, Mingjun; Yu, Hua; Kong, Zhenfang

2018-06-01

Stromal interaction molecule 1 (STIM1) is an endoplasmic reticulum Ca 2+ sensor which has been reported to be overexpressed in numerous types of cancer, and is involved in the cell proliferation, invasion, migration and metastasis frequently observed in cancer. However, the role of STIM1 in colorectal cancer (CRC) remains unknown. The purpose of the present study was to investigate the effect of STIM1 in human CRC. The expression of STIM1 was specifically knocked down using lentivirus-mediated small hairpin RNA (shRNA) interference techniques in the CRC cell lines HCT116 and SW1116. Subsequently, the efficiency of infection was confirmed using green fluorescent protein (GFP)-positive signals. The knockdown efficiency was further determined using the reverse transcription-quantitative polymerase chain reaction and western blotting analysis. As a result, CRC cell lines with STIM1 silenced were successfully constructed and subsequently employed in a series of cell function assays. Knockdown of STIM1 significantly suppressed cell proliferation and colony formation, as revealed by an MTT and colony formation assay. Furthermore, it was identified that STIM1 silencing may promote cell apoptosis through the induction of mitochondria-associated apoptosis, as was identified by increased expression levels of B-cell lymphoma 2 (Bcl-2)-associated death promoter, Bcl-2-associated X protein and poly(ADP-ribose) polymerase cleavage. Therefore, STIM1 may serve a critical role in the progression of CRC by regulating cell proliferation and apoptosis, which may provide a potential therapeutic target for the treatment of CRC.

miR-34a mediates oxaliplatin resistance of colorectal cancer cells by inhibiting macroautophagy via transforming growth factor-β/Smad4 pathway

PubMed Central

Sun, Chen; Wang, Fu-Jing; Zhang, Hao-Gang; Xu, Xun-Zheng; Jia, Rui-Chun; Yao, Lei; Qiao, Peng-Fei

2017-01-01

AIM To investigate whether microRNA (miR)-34a mediates oxaliplatin (OXA) resistance of colorectal cancer (CRC) cells by inhibiting macroautophagy via the transforming growth factor (TGF)-β/Smad4 pathway. METHODS miR-34a expression levels were detected in CRC tissues and CRC cell lines by quantitative real-time polymerase chain reaction. Computational search, functional luciferase assay and western blotting were used to demonstrate the downstream target of miR-34a in CRC cells. Cell viability was measured with Cell Counting Kit-8. Apoptosis and macroautophagy of CRC cells were analyzed by flow cytometry and transmission electron microscopy, and expression of beclin I and LC3-II was detected by western blotting. RESULTS Expression of miR-34a was significantly reduced while expression of TGF-β and Smad4 was increased in CRC patients treated with OXA-based chemotherapy. OXA treatment also resulted in decreased miR-34a levels and increased TGF-β and Smad4 levels in both parental cells and the OXA-resistant CRC cells. Activation of macroautophagy contributed to OXA resistance in CRC cells. Expression levels of Smad4 and miR-34a in CRC patients had a significant inverse correlation and overexpressing miR-34a inhibited macroautophagy activation by directly targeting Smad4 through the TGF-β/Smad4 pathway. OXA-induced downregulation of miR-34a and increased drug resistance by activating macroautophagy in CRC cells. CONCLUSION miR-34a mediates OXA resistance of CRC by inhibiting macroautophagy via the TGF-β/Smad4 pathway. PMID:28348487

Facile construction of fused benzimidazole-isoquinolinones that induce cell-cycle arrest and apoptosis in colorectal cancer cells.

PubMed

He, Liu-Jun; Yang, Dong-Lin; Li, Shi-Qiang; Zhang, Ya-Jun; Tang, Yan; Lei, Jie; Frett, Brendan; Lin, Hui-Kuan; Li, Hong-Yu; Chen, Zhong-Zhu; Xu, Zhi-Gang

2018-06-12

Colorectal cancer (CRC) is one of the most frequent, malignant gastrointestinal tumors, and strategies and effectiveness of current therapy are limited. A series of benzimidazole-isoquinolinone derivatives (BIDs) was synthesized and screened to identify novel scaffolds for CRC. Of the compounds evaluated, 7g exhibited the most promising anti-cancer properties. Employing two CRC cell lines, SW620 and HT29, 7g was found to suppress growth and proliferation of the cell lines at a concentration of ∼20 µM. Treatment followed an increase in G 2 /M cell cycle arrest, which was attributed to cyclin B1 and cyclin-dependent kinase 1 (CDK1) signaling deficiencies with simultaneous enhancement in p21 and p53 activity. In addition, mitochondrial-mediated apoptosis was induced in CRC cells. Interestingly, 7g decreased phosphorylated AKT, mTOR and 4E-BP1 levels, while promoting the expression/stability of PTEN. Since PTEN controls input into the PI3K/AKT/mTOR pathway, antiproliferative effects can be attributed to PTEN-mediated tumor suppression. Collectively, these results suggest that BIDs exert antitumor activity in CRC by impairing PI3K/AKT/mTOR signaling. Against a small kinase panel, 7g exhibited low affinity at 5 µM suggesting anticancer properties likely stem through a non-kinase mechanism. Because of the novelty of BIDs, the structure can serve as a lead scaffold to design new CRC therapies. Copyright © 2018. Published by Elsevier Ltd.

FOXQ1, a Novel Target of the Wnt Pathway and a New Marker for Activation of Wnt Signaling in Solid Tumors

PubMed Central

Christensen, Jon; Bentz, Susanne; Sengstag, Thierry; Shastri, V. Prasad; Anderle, Pascale

2013-01-01

Background The forkhead box transcription factor FOXQ1 has been shown to be upregulated in colorectal cancer (CRC) and metastatic breast cancer and involved in tumor development, epithelial-mesenchymal transition and chemoresistance. Yet, its transcriptional regulation is still unknown. Methods FOXQ1 mRNA and protein expression were analysed in a panel of CRC cell lines, and laser micro-dissected human biopsy samples by qRT-PCR, microarray GeneChip® U133 Plus 2.0 and western blots. FOXQ1 regulation was assayed by chromatin immunoprecipitation and luciferase reporter assays. Results FOXQ1 was robustly induced in CRC compared to other tumors, but had no predictive value with regards to grade, metastasis and survival in CRC. Prototype-based gene coexpression and gene set enrichment analysis showed a significant association between FOXQ1 and the Wnt pathway in tumors and cancer cell lines from different tissues. In vitro experiments confirmed, on a molecular level, FOXQ1 as a direct Wnt target. Analysis of known Wnt targets identified FOXQ1 as the most suitable marker for canonical Wnt activation across a wide panel of cell lines derived from different tissues. Conclusions Our data show that FOXQ1 is one of the most over-expressed genes in CRC and a direct target of the canonical Wnt pathway. It is a potential new marker for detection of early CRC and Wnt activation in tumors of different origins. PMID:23555880

Expression of Abelson Interactor 1 (Abi1) Correlates with Inflammation, KRAS Mutation and Adenomatous Change during Colonic Carcinogenesis

PubMed Central

Steinestel, Konrad; Brüderlein, Silke; Steinestel, Julie; Märkl, Bruno; Schwerer, Michael J.; Arndt, Annette; Kraft, Klaus; Pröpper, Christian; Möller, Peter

2012-01-01

Background Abelson interactor 1 (Abi1) is an important regulator of actin dynamics during cytoskeletal reorganization. In this study, our aim was to investigate the expression of Abi1 in colonic mucosa with and without inflammation, colonic polyps, colorectal carcinomas (CRC) and metastases as well as in CRC cell lines with respect to BRAF/KRAS mutation status and to find out whether introduction of KRAS mutation or stimulation with TNFalpha enhances Abi1 protein expression in CRC cells. Methodology/Principal Findings We immunohistochemically analyzed Abi1 protein expression in 126 tissue specimens from 95 patients and in 5 colorectal carcinoma cell lines with different mutation status by western immunoblotting. We found that Abi1 expression correlated positively with KRAS, but not BRAF mutation status in the examined tissue samples. Furthermore, Abi1 is overexpressed in inflammatory mucosa, sessile serrated polyps and adenomas, tubular adenomas, invasive CRC and CRC metastasis when compared to healthy mucosa and BRAF-mutated as well as KRAS wild-type hyperplastic polyps. Abi1 expression in carcinoma was independent of microsatellite stability of the tumor. Abi1 protein expression correlated with KRAS mutation in the analyzed CRC cell lines, and upregulation of Abi1 could be induced by TNFalpha treatment as well as transfection of wild-type CRC cells with mutant KRAS. The overexpression of Abi1 could be abolished by treatment with the PI3K-inhibitor Wortmannin after KRAS transfection. Conclusions/Significance Our results support a role for Abi1 as a downstream target of inflammatory response and adenomatous change as well as oncogenic KRAS mutation via PI3K, but not BRAF activation. Furthermore, they highlight a possible role for Abi1 as a marker for early KRAS mutation in hyperplastic polyps. Since the protein is a key player in actin dynamics, our data encourages further studies concerning the exact role of Abi1 in actin reorganization upon enhanced KRAS/PI3K signalling during colonic tumorigenesis. PMID:22808230

Hsa-miR-875-5p exerts tumor suppressor function through down-regulation of EGFR in colorectal carcinoma (CRC)

PubMed Central

Li, Qi; Xue, Peng; Chen, Zhixiao; Dong, Xiao; Xue, Ying

2016-01-01

Hsa-miRNA-875-5p (miR-875-5p) has recently been discovered to have anticancer efficacy in different organs. However, the role of miR-875-5p on colorectal carcinoma (CRC) is still ambiguous. In this study, we investigated the role of miR-875-5p on the development of CRC. The results indicated that miR-875-5p was significantly down-regulated in primary tumor tissues and very low levels were found in CRC cell lines. Ectopic expression of miR-875-5p in CRC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4 and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-875-5p induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-875-5p inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene EGFR was revealed to be a putative target of miR-875-5p, which was inversely correlated with miR-875-5p expression in CRC. Taken together, our results demonstrated that miR-875-5p played a pivotal role on CRC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic EGFR. PMID:27302926

Hsa-miR-875-5p exerts tumor suppressor function through down-regulation of EGFR in colorectal carcinoma (CRC).

PubMed

Zhang, Tiening; Cai, Xun; Li, Qi; Xue, Peng; Chen, Zhixiao; Dong, Xiao; Xue, Ying

2016-07-05

Hsa-miRNA-875-5p (miR-875-5p) has recently been discovered to have anticancer efficacy in different organs. However, the role of miR-875-5p on colorectal carcinoma (CRC) is still ambiguous. In this study, we investigated the role of miR-875-5p on the development of CRC. The results indicated that miR-875-5p was significantly down-regulated in primary tumor tissues and very low levels were found in CRC cell lines. Ectopic expression of miR-875-5p in CRC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4 and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-875-5p induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-875-5p inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene EGFR was revealed to be a putative target of miR-875-5p, which was inversely correlated with miR-875-5p expression in CRC. Taken together, our results demonstrated that miR-875-5p played a pivotal role on CRC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic EGFR.

Myotubularin-related protein 7 inhibits insulin signaling in colorectal cancer

PubMed Central

Gutting, Tobias; Friedrich, Teresa; Gaiser, Timo; Magdeburg, Julia; Kienle, Peter; Ruh, Hermelindis; Hopf, Carsten; Behrens, Hans-Michael; Röcken, Christoph; Hanoch, Tamar; Seger, Rony; Ebert, Matthias P.A.; Burgermeister, Elke

2016-01-01

Phosphoinositide (PIP) phosphatases such as myotubularins (MTMs) inhibit growth factor receptor signaling. However, the function of myotubularin-related protein 7 (MTMR7) in cancer is unknown. We show that MTMR7 protein was down-regulated with increasing tumor grade (G), size (T) and stage (UICC) in patients with colorectal cancer (CRC) (n=1786). The presence of MTMR7 in the stroma correlated with poor prognosis, whereas MTMR7 expression in the tumor was not predictive for patients' survival. Insulin reduced MTMR7 protein levels in human CRC cell lines, and CRC patients with type 2 diabetes mellitus (T2DM) or loss of imprinting (LOI) of insulin-like growth factor 2 (IGF2) had an increased risk for MTMR7 loss. Mechanistically, MTMR7 lowered PIPs and inhibited insulin-mediated AKT-ERK1/2 signaling and proliferation in human CRC cell lines. MTMR7 provides a novel link between growth factor signaling and cancer, and may thus constitute a potential marker or drug target for human CRC. PMID:27409167

miR-106a suppresses tumor cells death in colorectal cancer through targeting ATG7.

PubMed

Hao, Haibin; Xia, Guangfeng; Wang, Chao; Zhong, Fuping; Liu, Laipeng; Zhang, Dong

2017-06-01

Autophagy-related gene 7 (ATG7) and miR-106a play an important role in cancer cell autophagy and apoptosis, but the outcome of ATG7 and miR-106a in colorectal cancer (CRC) still remains not clear. In this study, we found that ATG7 and miR-106a expression were mutually related with cell death and prognosis in CRC patients. In addition, we also showed that ATG7 and miR-106a expression were changeable in colorectal cancer cell lines when compared with normal cell lines, but ATG7 and miR-106a mRNA level was negatively correlated. Furthermore, ATG7 protein and mRNA levels decreased after over-expression of miR-106a, whereas the suppression of ATG7 had the opposite effect. We confirmed that miR-106a down-regulated ATG7 mRNA level by binding the specific sequence of ATG7 mRNA 3'UTR region. Moreover, the over-expression of ATG7 induced CRC cells death both in vitro and in vivo. Taken together, our study data demonstrated that ATG7 aggravated the cell death of CRC, which was inhibited by miR-106a.

Knockdown of long non‑coding RNA PVT1 reverses multidrug resistance in colorectal cancer cells.

PubMed

Fan, Heng; Zhu, Jian-Hua; Yao, Xue-Qing

2018-06-01

Multidrug resistance (MDR) is one of the primary causes of chemotherapy failure in colorectal cancer (CRC), and extensive biological studies into MDR are required. The non‑coding RNA plasmacytoma variant translocation 1 (PVT1) has been demonstrated to be associated with low survival rates in patients with CRC. However, whether PVT1 serves a critical function in the MDR of CRC remains to be determined. To determine the association between PVT1 expression and 5‑fluorouracil (5‑FU) resistance in CRC, the expression levels of PVT1 mRNA in 5‑FU‑resistant CRC tissues and cell lines (HCT‑8/5‑FU and HCT‑116/5‑FU) were assessed by a reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). Cytotoxicity was evaluated using a Cell Counting Kit‑8 assay and apoptosis rates were assessed via flow cytometry. In the present study, PVT1 mRNA was highly expressed in 5‑FU‑resistant CRC tissues and cell lines. HCT‑8/5‑FU and HCT‑116/5‑FU cells transfected with small interfering RNA PVT1 and treated with 5‑FU exhibited higher apoptotic rates and lower survival rates. By contrast, overexpression of PVT1 in HCT‑8 and HCT‑116 cells transfected with lentiviral vector‑PVT1‑green fluorescent protein and treated with 5‑FU exhibited lower apoptosis rates and higher survival rates. RT‑qPCR and western blotting demonstrated that the overexpression of PVT1 increased the mRNA and protein expression levels of multidrug resistance‑associated protein 1, P‑glycoprotein, serine/threonine‑protein kinase mTOR and apoptosis regulator Bcl2. The present study indicates that PVT1 overexpression may promote MDR in CRC cells, and suggested that inhibition of PVT1 expression may be an effective therapeutic strategy for reversing MDR in CRC.

Decreasing CNPY2 Expression Diminishes Colorectal Tumor Growth and Development through Activation of p53 Pathway.

PubMed

Yan, Ping; Gong, Hui; Zhai, Xiaoyan; Feng, Yi; Wu, Jun; He, Sheng; Guo, Jian; Wang, Xiaoxia; Guo, Rui; Xie, Jun; Li, Ren-Ke

2016-04-01

Neovascularization drives tumor development, and angiogenic factors are important neovascularization initiators. We recently identified the secreted angiogenic factor CNPY2, but its involvement in cancer has not been explored. Herein, we investigate CNPY2's role in human colorectal cancer (CRC) development. Tumor samples were obtained from CRC patients undergoing surgery. Canopy 2 (CNPY2) expression was analyzed in tumor and adjacent normal tissue. Stable lines of human HCT116 cells expressing CNPY2 shRNA or control shRNA were established. To determine CNPY2's effects on tumor xenografts in vivo, human CNPY2 shRNA HCT116 cells and controls were injected into nude mice, separately. Cellular apoptosis, growth, and angiogenesis in the xenografts were evaluated. CNPY2 expression was significantly higher in CRC tissues. CNPY2 knockdown in HCT116 cells inhibited growth and migration and promoted apoptosis. In xenografts, CNPY2 knockdown prevented tumor growth and angiogenesis and promoted apoptosis. Knockdown of CNPY2 in the HCT116 CRC cell line reversibly increased p53 activity. The p53 activation increased cyclin-dependent kinase inhibitor p21 and decreased cyclin-dependent kinase 2, thereby inhibiting tumor cell growth, inducing cell apoptosis, and reducing angiogenesis both in vitro and in vivo. CNPY2 may play a critical role in CRC development by enhancing cell proliferation, migration, and angiogenesis and by inhibiting apoptosis through negative regulation of the p53 pathway. Therefore, CNPY2 may represent a novel CRC therapeutic target and prognostic indicator. Copyright © 2016 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

NFATC3-PLA2G15 Fusion Transcript Identified by RNA Sequencing Promotes Tumor Invasion and Proliferation in Colorectal Cancer Cell Lines.

PubMed

Jang, Jee-Eun; Kim, Hwang-Phill; Han, Sae-Won; Jang, Hoon; Lee, Si-Hyun; Song, Sang-Hyun; Bang, Duhee; Kim, Tae-You

2018-06-14

This study was designed to identify novel fusion transcripts (FTs) and their functional significance in colorectal cancer lines. We performed paired-end RNA sequencing of 28 colorectal cancer (CRC) cell lines. FT candidates were identified using TopHat-fusion, ChimeraScan, and FusionMap tools and further experimental validation was conducted through reverse transcription-polymerase chain reaction and Sanger sequencing. FT was depleted in human CRC line and the effects on cell proliferation, cell migration, and cell invasion were analyzed. 1,380 FT candidates were detected through bioinformatics filtering. We selected 6 candidate FTs, including 4 inter-chromosomal and 2 intra-chromosomal FTs and each FT was found in at least 1 of the 28 cell lines. Moreover, when we tested 19 pairs of CRC tumor and adjacent normal tissue samples, NFATC3-PLA2G15 FT was found in 2. Knockdown of NFATC3-PLA2G15 using siRNA reduced mRNA expression of epithelial-mesenchymal transition (EMT) markers such as vimentin, twist, and fibronectin and increased mesenchymal-epithelial transition markers of E-cadherin, claudin-1, and FOXC2 in colo-320 cell line harboring NFATC3-PLA2G15 FT. The NFATC3-PLA2G15 knockdown also inhibited invasion, colony formation capacity, and cell proliferation. These results suggest that that NFATC3-PLA2G15 FTs may contribute to tumor progression by enhancing invasion by EMT and proliferation.

Protein and siRNA delivery by transportan and transportan 10 into colorectal cancer cell lines.

PubMed

Wierzbicki, Piotr M; Kogut-Wierzbicka, Marzena; Ruczynski, Jaroslaw; Siedlecka-Kroplewska, Kamila; Kaszubowska, Lucyna; Rybarczyk, Agnieszka; Alenowicz, Magdalena; Rekowski, Piotr; Kmiec, Zbigniew

2014-01-01

Cell penetrating peptides (CPPs) have the ability to translocate through cell membranes with high efficiency and therefore can introduce biological agents with pharmaceutical properties into the cell. Transportan (TP) and its shorter analog transportan 10 (TP10) are among the best studied CPPs, however, their effects on viability of and cargo introduction into colorectal cancer (CRC) cells have yet not been investigated. The aim of our study was to evaluate the cytotoxic effects of TP and TP10 on representative CRC lines and the efficiency of protein (streptavidin) and siRNA cargo delivery by TP-biotinylated derivatives (TP-biot). HT29 (early stage CRC model) and HCT116 (metastatic CRC model) cell lines were incubated with TP, TP10, TP-biot1, TP-biot13 and TP10-biot1. The effects of studied CPPs on cell viability and cell cycle were assessed by MTT and annexin V assays. The uptake of streptavidin-FITC complex into cells was determined by flow cytometry and fluorescence microscopy, with the inhibition of cellular vesicle trafficking by brefeldin A. The efficiency of siRNA for SASH1 gene delivery was measured by quantitative PCR (qPCR). Since up to 10 µM concentrations of each CPP showed no significant cytotoxic effect, the concentrations of 0.5-5 µM were used for further analyses. Within this concentration range none of the studied CPPs affected cell viability and cell cycle. The efficient and endocytosis-independent introduction of streptavidin-FITC complex into cells was observed for TP10-biot1 and TP-biot1 with the cytoplasmic location of the fluorescent cargo; decreased SASH1 mRNA level was noticed with the use of siRNA and analyzed CPPs. We conclude that TP, TP10 and their biotinylated derivatives can be used as efficient delivery vehicles of small and large cargoes into CRC cells.

Systemic analysis of different colorectal cancer cell lines and TCGA datasets identified IGF-1R/EGFR-PPAR-CASPASE axis as important indicator for radiotherapy sensitivity.

PubMed

Chen, Lin; Zhu, Zhe; Gao, Wei; Jiang, Qixin; Yu, Jiangming; Fu, Chuangang

2017-09-05

Insulin-like growth factor 1 receptor (IGF-1R) is proved to contribute the development of many types of cancers. But, little is known about its roles in radio-resistance of colorectal cancer (CRC). Here, we demonstrated that low IGF-1R expression value was associated with the better radiotherapy sensitivity of CRC. Besides, through Quantitative Real-time PCR (qRT-PCR), the elevated expression value of epidermal growth factor receptor (EGFR) was observed in CRC cell lines (HT29, RKO) with high radio-sensitivity compared with those with low sensitivity (SW480, LOVO). The irradiation induced apoptosis rates of wild type and EGFR agonist (EGF) or IGF-1R inhibitor (NVP-ADW742) treated HT29 and SW480 cells were quantified by flow cytometry. As a result, the apoptosis rate of EGF and NVP-ADW742 treated HT29 cells was significantly higher than that of those wild type ones, which indicated that high EGFR and low IGF-1R expression level in CRC was associated with the high sensitivity to radiotherapy. We next conducted systemic bioinformatics analysis of genome-wide expression profiles of CRC samples from the Cancer Genome Atlas (TCGA). Differential expression analysis between IGF-1R and EGFR abnormal CRC samples, i.e. CRC samples with higher IGF-1R and lower EGFR expression levels based on their median expression values, and the rest of CRC samples identified potential genes contribute to radiotherapy sensitivity. Functional enrichment of analysis of those differential expression genes (DEGs) in the Database for Annotation, Visualization and Integrated Discovery (DAVID) indicated PPAR signaling pathway as an important pathway for the radio-resistance of CRC. Our study identified the potential biomarkers for the rational selection of radiotherapy for CRC patients. Copyright © 2017 Elsevier B.V. All rights reserved.

Gab3 is required for human colorectal cancer cell proliferation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xiang, Shihao; Wang, Na; Hui, Pingping

Here, we focused on the potential function of Gab3, an uncommon Gab family protein, in human colorectal cancer (CRC) cells. We found that Gab3 was only expressed in human colon cancer tissues as well as in established (HCT-116 and HT-29 lines) and primary human CRC cells. It was however absent in normal human colon cancer tissues and in FHC colon epithelial cells. Knockdown of Gab3 by targeted-shRNAs inhibited proliferation of the CRC cells. Reversely, exogenous over-expression of Gab3 promoted CRC cell proliferation. At the signaling level, Gab3 co-precipitated with p85 and SHP2 in CRC cells, which was required for subsequentmore » Akt and Erk activation. Gab3 shRNA knockdown inhibited Akt and Erk activation, yet Gab3 over-expression augmented it. In vivo, HCT-116 xenograft tumor growth in severe combined immune deficient (SCID) mice was suppressed following expressing Gab3 shRNAs. Meanwhile, Akt and Erk activation in Gab3 shRNA-expressing tumors was also largely inhibited. Together, our results suggest that Gab3 expression in CRC cells is important for Akt-Erk activation and cell proliferation. - Highlights: • Gab3 is only expressed in colorectal cancer (CRC) cells, but not in colon epithelial cells. • Gab3 shRNA knockdown inhibits CRC cell proliferation. • Exogenous over-expression of Gab3 promotes HCT-116 cell proliferation. • Gab3 co-precipitates with p85 and SHP2 to mediate Akt and Erk activation in CRC cells. • HCT-116 tumor growth in SCID mice is suppressed with expression of Gab3 shRNAs.« less

Collaborative Model for Acceleration of Individualized Therapy of Colon Cancer

DTIC Science & Technology

2013-10-01

Methods: The anti-proliferative effects of TAK-960 as a single agent and in combination with irinotecan (SN38) or cetuximab were assessed using an assay...following treatment with TAK-960 alone or in combination with standard agents (irinotecan or cetuximab ). Results: CRC cell lines were quite...sensitive to TAK-960 with IC50 values ranging from 0.007 to 1 umol/L. While no synergy was observed in the KRAS WT CRC cell lines in the cetuximab

Hyperphosphorylation of PP2A in colorectal cancer and the potential therapeutic value showed by its forskolin-induced dephosphorylation and activation.

PubMed

Cristóbal, Ion; Rincón, Raúl; Manso, Rebeca; Madoz-Gúrpide, Juan; Caramés, Cristina; del Puerto-Nevado, Laura; Rojo, Federico; García-Foncillas, Jesús

2014-09-01

The tumor suppressor protein phosphatase 2A (PP2A) is frequently inactivated in human cancer and phosphorylation of its catalytic subunit (p-PP2A-C) at tyrosine-307 (Y307) has been described to inhibit this phosphatase. However, its molecular and clinical relevance in colorectal cancer (CRC) remains unclear. p-PP2A-C Y307 was determined by immunoblotting in 7 CRC cell lines and 35 CRC patients. CRC cells were treated with the PP2A activator forskolin alone or combined with the PP2A inhibitor okadaic acid, 5-fluorouracil and oxaliplatin. We examined cell growth, colonosphere formation, caspase activity and AKT and ERK activation. PP2A-C was found hyperphosphorylated in CRC cell lines. Forskolin dephosphorylated and activated PP2A, impairing proliferation and colonosphere formation, and inducing activation of caspase 3/7 and changes in AKT and ERK phosphorylation. Moreover, forskolin showed additive effects with 5-fluorouracil and oxaliplatin treatments. Analysis of p-PP2A-C Y307 in primary tumors confirmed the presence of this alteration in a subgroup of CRC patients. Our data show that PP2A-C hyperphosphorylation is a frequent event that contributes to PP2A inhibition in CRC. Antitumoral effects of forskolin-mediated PP2A activation suggest that the analysis of p-PP2A-C Y307 status could be used to identify a subgroup of patients who would benefit from treatments based on PP2A activators. Copyright © 2014 Elsevier B.V. All rights reserved.

Heat shock protein 70-2 (HSP70-2) is a novel therapeutic target for colorectal cancer and is associated with tumor growth.

PubMed

Jagadish, Nirmala; Parashar, Deepak; Gupta, Namita; Agarwal, Sumit; Suri, Vaishali; Kumar, Rajive; Suri, Vitusha; Sadasukhi, Trilok Chand; Gupta, Anju; Ansari, Abdul S; Lohiya, Nirmal Kumar; Suri, Anil

2016-07-29

Colorectal cancer (CRC) is the third leading cause of cancer related deaths worldwide both in men and women. Our recent studies have indicated an association of heat shock protein 70-2 (HSP70-2) with bladder urothelial carcinoma. In the present study, we investigated the association of HSP70-2 with various malignant properties of colorectal cancer cells and clinic-pathological features of CRC in clinical specimens. HSP70-2 mRNA and protein was investigated expression by RT-PCR, immunohistochemistry, immunofluorescence, flow cytometry and Western blotting in CRC clinical specimens and COLO205 and HCT116 cell lines. Plasmid-based gene silencing approach was employed to study the association of HSP70-2 with various malignant properties of COLO205 and HCT116 cells in in vitro and with tumor progression in in vivo COLO205 human xenograft mice model. HSP70-2 expression was detected in 78 % of CRC patients irrespective of various stages and grades by RT-PCR and IHC. Our analysis further revealed that HSP70-2 expression was detected in both COLO205 and HCT116 cell lines. Ablation of HSP70-2 expression resulted in reduced cellular growth, colony forming ability, migratory and invasive ability of CRC cells. In addition, ablation of HSP70-2 expression showed significant reduction in tumor growth in COLO205 human xenograft in in vivo mouse model. Collectively, our results indicate that HSP70-2 is associated with CRC clinical specimens. In addition, down regulation of HSP70-2 expression reduces cellular proliferation and tumor growth indicating that HSP70-2 may be a potential therapeutic target for CRC treatment.

Curcumin enhances the effects of irinotecan on colorectal cancer cells through the generation of reactive oxygen species and activation of the endoplasmic reticulum stress pathway.

PubMed

Huang, Yan-Feng; Zhu, Da-Jian; Chen, Xiao-Wu; Chen, Qi-Kang; Luo, Zhen-Tao; Liu, Chang-Chun; Wang, Guo-Xin; Zhang, Wei-Jie; Liao, Nv-Zhu

2017-06-20

Although initially effective against metastatic colorectal cancer (CRC), irinotecan-based chemotherapy leads to resistance and adverse toxicity. Curcumin is well known for its anti-cancer effects in many cancers, including CRC. Here, we describe reactive oxygen species (ROS) generation and endoplasmic reticulum (ER) stress as important mechanisms by which curcumin enhances irinotecan's effects on CRC cells. CRC cell lines were treated with curcumin and/or irinotecan for 24 h, and then evaluated using cell proliferation assays, cell apoptosis assays, cell cycle analysis, intracellular Ca2+ measurements, ROS measurements and immunoblotting for key ER stress-related proteins. We found that cell viability was inhibited and apoptosis was increased, accompanied by ROS generation and ER stress activation in CRC cells treated with curcumin alone or in combination with irinotecan. Blocking ROS production attenuated the expression of two markers of ER stress: binding of immunoglobulin protein (BIP) and CCAAT/enhancer-binding protein homologous protein (CHOP). Blocking CHOP expression using RNA interference also inhibited ROS generation. These results demonstrated that curcumin could enhance the effects of irinotecan on CRC cells by inhibiting cell viability and inducing cell cycle arrest and apoptosis, and that these effects may be mediated, in part, by ROS generation and activation of the ER stress pathway.

Frequent Truncating Mutation of TFAM Induces Mitochondrial DNA Depletion and Apoptotic Resistance in Microsatellite-Unstable Colorectal Cancer

PubMed Central

Guo, Jianhui; Zheng, Li; Liu, Wenyong; Wang, Xianshu; Wang, Zemin; Wang, Zehua; French, Amy J.; Kang, Dongchon; Chen, Lin; Thibodeau, Stephen N.; Liu, Wanguo

2013-01-01

The mitochondrial transcription factor A (TFAM) is required for mitochondrial DNA (mtDNA) replication and transcription. Disruption of TFAM results in heart failure and premature aging in mice. But very little is known about the role of TFAM in cancer development. Here, we report the identification of frequent frameshift mutations in the coding mononucleotide repeat of TFAM in sporadic colorectal cancer (CRC) cell lines and in primary tumors with microsatellite instability (MSI), but not in microsatellite stable (MSS) CRC cell lines and tumors. The presence of the TFAM truncating mutation, in CRC cells with MSI, reduced the TFAM protein level in vivo and in vitro and correlated with mtDNA depletion. Furthermore, forced overexpression of wild-type TFAM in RKO cells carrying a TFAM truncating mutation suppressed cell proliferation and inhibited RKO cell-induced xenograft tumor growth. Moreover, these cells showed more susceptibility to cisplatin-induced apoptosis due to an increase of cytochrome b (Cyt b) expression and its release from mitochondria. An interaction assay between TFAM and the heavy-strand promoter (HSP) of mitochondria revealed that mutant TFAM exhibited reduced binding to HSP, leading to reduction in Cyt b transcription. Collectively, these data provide evidence that a high incidence of TFAM truncating mutations leads to mitochondrial copy number reduction and mitochondrial instability, distinguishing most CRC with MSI from MSS CRC. These mutations may play an important role in tumorigenesis and cisplatin-induced apoptotic resistance of most microsatellite-unstable CRCs. PMID:21467167

The impact of polyunsaturated fatty acids on DNA methylation and expression of DNMTs in human colorectal cancer cells.

PubMed

Sarabi, Mostafa Moradi; Naghibalhossaini, Fakhraddin

2018-05-01

Growing evidence suggests a role of polyunsaturated fatty acids (PUFA) in the prevention of various types of malignancy, including colorectal cancer (CRC). No published studies have yet examined the direct effect of PUFA treatment on DNA methylation in CRC cells. In this study, 5 human CRC cells were treated with 100 μM DHA, EPA, and LA for 6 days and changes in their global- and gene-specific DNA methylation status as well as expression of DNA methyl transferases (DNMT) were investigated. Cell-type specific differences in DNA methylation and expression of DNMTs were observed in PUFA-treated cells. DHA and EPA treatment induced global hypermethylation in HT29/219 and HCT116 cells, but reduced methylation in Caco2 cells (p < 0.05). Among 10 tumor related genes tested in 5 CRC cell lines, DHA and EPA induced promoter demethylation of Cox2 in HT29/219, p14 and PPARγ in HCT116, and ECAD in SW742 cells. Cell-type specific differences in expression of DNMT1, DNMT3a, and 3b genes were also observed between PUFA-treated and control cells (p < 0.05). Overall, treatment of PUFAs coordinately induced the expression of DNMTs in HT29/219, but suppressed in other 4 cell lines investigated in this study. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Long Non-Coding RNA SH3PXD2A-AS1 Promotes Cell Progression Partly Through Epigenetic Silencing P57 and KLF2 in Colorectal Cancer.

PubMed

Ma, Zhonghua; Peng, Peng; Zhou, Jing; Hui, Bingqing; Ji, Hao; Wang, Juan; Wang, Keming

2018-05-03

Colorectal cancer (CRC) is one of the most commonly diagnosed malignancies worldwide. Current evidence has revealed the key roles of long non-coding RNAs (IncRNAs) in multiple cancers, including CRC. In this study we identified the lncRNA SH3PXD2A-AS1 as a novel molecule associated with CRC progression by analyzing the publicly available data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) datasets. Quantitative real-time polymerase chain reaction (qRT-PCR) assays were performed to examine the expression levels of SH3PXD2A-AS1 in CRC tissue samples and CRC cell lines. Cell viability examination, colony-formation experiments, ethynyl deoxyuridine (Edu) assays and flow cytometry were performed to investigate the roles of SH3PXD2A-AS1 in CRC proliferation, cell cycle regulation, and apoptosis. Transwell assays were used to explore the effects of SH3PXD2A-AS1 on CRC cells migration and invasion. A nude mice model was used to assess the effects of SH3PXD2A-AS1 on tumorigenesis in vivo. Subcellular fractionation, RNA immunoprecipitation (RIP), and chromatin immunoprecipitation (ChIP) assays were conducted to detect the molecular mechanisms of SH3PXD2A-ASl-mediated gene expression. Rescue assays were used to determine whether P57 and Kruppel-like factor 2 (KLF2) were involved in SH3PXD2A-ASl-dependent CRC proliferation. We firstly found that SH3PXD2A-AS1 was significantly upregulated in CRC tissues and cell lines, and overexpression of SH3PXD2A-AS1 was correlated with tumor size, TNM stage, and lymph node metastasis in patients with CRC. Furthermore, SH3PXD2A-AS1 knockdown inhibited CRC cells proliferation, migration and invasion in vitro, and suppressed tumorigenesis in vivo. Mechanistic studies indicated that SH3PXD2A-AS1 could epiqenetically repress P57 and KLF2 expression through interaction with EZH2. Rescue experiments suggested that SH3PXD2A-ASl-mediated oncogenesis was impaired by overexpression of P57 or KLF2. Interestingly, the expression of SH3PXD2A-AS1 was inversely correlated with the expression of P57 and KLF2 in CRC tissue samples. Our research presents the first evidence that SH3PXD2A-AS1 acts as an oncogene in CRC, and may be a promising diagnostic or therapeutic target in patients with CRC. © 2018 The Author(s). Published by S. Karger AG, Basel.

Connective tissue growth factor acts as a therapeutic agent and predictor for peritoneal carcinomatosis of colorectal cancer.

PubMed

Lin, Been-Ren; Chang, Cheng-Chi; Chen, Robert Jeen-Chen; Jeng, Yung-Ming; Liang, Jin-Tung; Lee, Po-Huang; Chang, King-Jen; Kuo, Min-Liang

2011-05-15

Here, we aimed to investigate the role of connective tissue growth factor (CTGF) in peritoneal carcinomatosis (PC) associated with colorectal cancer (CRC) and to characterize the underlying mechanism of CTGF mediating adhesion. A cohort of 136 CRC patient specimens was analyzed in this study. CRC cell lines were used for in vitro adhesion assay and in vivo peritoneal dissemination experiment. Recombinant CTGF protein treatment, transfection of CTGF expression plasmids, and knockdown of CTGF expression in CRC cells were utilized to evaluate the integrin α5, which served as a target of CTGF in inhibiting peritoneal seeding. The analysis of CRC tissues revealed an inverse correlation between CTGF expression and prevalence of PC. Lower CTGF level in CRC patients was associated with higher peritoneal recurrence rate after surgery. Inducing CTGF expression in cancer cells resulted in decreased incidence of PC and increased rate of mice survival. The mice received intraperitoneal injection of recombinant CTGF protein simultaneously with cancer cells or following tumor formation; in both cases, peritoneal tumor dissemination was found to be effectively inhibited in the mouse model. Functional assay revealed that CTGF significantly decreased the CRC cell adhesion ability, and integrin α5 was confirmed by reverse transcriptase PCR and functional blocking assay as a downstream effector in the CTGF-mediated inhibition of CRC cell adhesion. CTGF acts as a molecular predictor of PC and could be a potential therapeutic target for the chemoprevention and treatment of PC in CRC patients. ©2011 AACR.

Potential role of TRIM3 as a novel tumour suppressor in colorectal cancer (CRC) development.

PubMed

Piao, Mei-Yu; Cao, Hai-Long; He, Na-Na; Xu, Meng-Que; Dong, Wen-Xiao; Wang, Wei-Qiang; Wang, Bang-Mao; Zhou, Bing

2016-01-01

Colorectal cancer (CRC) is the third leading cause of cancer-related mortality in the United States. Recent cancer genome-sequencing efforts and complementary functional studies have led to the identification of a collection of candidate 'driver' genes involved in CRC tumorigenesis. Tripartite motif (TRIM3) is recently identified as a tumour suppressor in glioblastoma but this tumour-suppressive function has not been investigated in CRC. In this study, we investigated the potential role of TRIM3 as a tumour suppressor in CRC development by manipulating the expression of TRIM3 in two authentic CRC cell lines, HCT116 and DLD1, followed by various functional assays, including cell proliferation, colony formation, scratch wound healing, soft agar, and invasion assays. Xenograft experiment was performed to examine in vivo tumour-suppressive properties of TRIM3. Small-interfering RNA (siRNA) mediated knockdown of TRIM3 conferred growth advantage in CRC cells. In contrast, overexpression of TRIM3 affected cell survival, cell migration, anchorage independent growth and invasive potential in CRC cells. In addition, TRIM3 was found to be down-regulated in human colon cancer tissues compared with matched normal colon tissues. Overexpression of TRIM3 significantly inhibited tumour growth in vivo using xenograft mouse models. Mechanistic investigation revealed that TRIM3 can regulate p53 protein level through its stabilisation. TRIM3 functions as a tumour suppressor in CRC progression. This tumour-suppressive function is exerted partially through regulation of p53 protein. Therefore, this protein may represent a novel therapeutic target for prevention or intervention of CRC.

Isolated Lung Perfusion as an Adjuvant Treatment of Colorectal Cancer Lung Metastases: A Preclinical Study in a Pig Model

PubMed Central

Pagès, Pierre-Benoit; Facy, Olivier; Mordant, Pierre; Ladoire, Sylvain; Magnin, Guy; Lokiec, Francois; Ghiringhelli, Francois; Bernard, Alain

2013-01-01

Background The lung is a frequent site of colorectal cancer (CRC) metastases. After surgical resection, lung metastases recurrences have been related to the presence of micrometastases, potentially accessible to a high dose chemotherapy administered via adjuvant isolated lung perfusion (ILP). We sought to determine in vitro the most efficient drug when administered to CRC cell lines during a short exposure and in vivo its immediate and delayed tolerance when administered via ILP. Methods First, efficacy of various cytotoxic molecules against a panel of human CRC cell lines was tested in vitro using cytotoxic assay after a 30-minute exposure. Then, early (operative) and delayed (1 month) tolerance of two concentrations of the molecule administered via ILP was tested on 19 adult pigs using hemodynamic, biological and histological criteria. Results In vitro, gemcitabine (GEM) was the most efficient drug against selected CRC cell lines. In vivo, GEM was administered via ILP at regular (20 µg/ml) or high (100 µg/ml) concentrations. GEM administration was associated with transient and dose-dependant pulmonary vasoconstriction, leading to a voluntary decrease in pump inflow in order to maintain a stable pulmonary artery pressure. After this modulation, ILP using GEM was not associated with any systemic leak, systemic damage, and acute or delayed histological pulmonary toxicity. Pharmacokinetics studies revealed dose-dependant uptake associated with heterogenous distribution of the molecule into the lung parenchyma, and persistent cytotoxicity of venous effluent. Conclusions GEM is effective against CRC cells even after a short exposure. ILP with GEM is a safe and reproducible technique. PMID:23527205

LRIG1, a 3p tumor suppressor, represses EGFR signaling and is a novel epigenetic silenced gene in colorectal cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kou, Changhua, E-mail: chkoukou@hotmail.com; Zhou, Tian; Han, Xilin

2015-08-21

Downregulation of LRIG1 was found in many types of cancer. However, data concerning the possible mechanism of LRIG1 reduction in cancers were not reported yet. To analyze the regulation and function of LRIG1 in colorectal cancer (CRC), 6 cell lines, 46 paired tissues from primary CRC cases were employed in this study. In CRC cell lines, under-expression of LRIG1 was correlated with promoter region hypermethylation, and restoration of LRIG1 was induced by 5-Aza-2'-deoxyazacytidine treatment. Subsequently, we ectopically expressed LRIG1 in LRIG1 low-expressing HCT-116 cells and suppressed LRIG1 in LRIG1 high-expressing LoVo cells. We found that over-expression of LRIG1 inhibits cellmore » proliferation and colony formation and tumor growth, while knockdown of LRIG1 promotes cell proliferation and colony formation. Decreased and increased EGFR/AKT signaling pathway may partially explain the lower and higher rates of proliferation in CRC cells transfected with LRIG1 cDNA or shRNA. In clinical samples, we compared the methylation, mRNA and protein expression of LRIG1 in samples of CRC tissues. A significant increase in LRIG1 methylation was identified in CRC specimens compared to adjacent normal tissues and that it was negatively correlated with its mRNA and protein expression. In conclusion, LRIG1 is frequently methylated in human CRC and consequent mRNA and protein downregulation may contribute to tumor growth by activating EGFR/AKT signaling. - Highlights: • Promoter methylation of LRIG1 occurred in colorectal cancer cells and tumors. • Restoration of LRIG1 inhibits tumor growth in vitro and in vivo. • Overexpression or knockdown of LRIG1 regulates EGFR/AKT and downstream apoptosis. • Methylation of LRIG1 correlates with its mRNA and protein downregulation. • LRIG1 was firstly identified as an epigenetic target in cancer.« less

Acriflavine enhances the antitumor activity of the chemotherapeutic drug 5-fluorouracil in colorectal cancer cells.

PubMed

Zargar, Parisa; Ghani, Esmaeel; Mashayekhi, Farideh Jalali; Ramezani, Amin; Eftekhar, Ebrahim

2018-06-01

5-Fluorouracil (5-FU)-based chemotherapy improves the overall survival rates of patients with colorectal cancer (CRC). However, only a small proportion of patients respond to 5-FU when used as a single agent. The aim of the present study was to investigate whether the anticancer property of 5-FU is potentiated by combination treatment with acriflavine (ACF) in CRC cells. Additionally, the potential underlying molecular mechanisms of the cytotoxic effect of ACF were determined. The cytotoxic effects of ACF, 5-FU and irinotecan on different CRC cell lines with different p53 status were investigated using an MTT assay. SW480 cells that express a mutated form of p53 and two other CRC cell lines were used, HCT116 and LS174T, with wild-type p53. To determine the effect of ACF on the sensitivity of cells to 5-FU, cells were co-treated with the 30% maximal inhibitory concentration (IC 30 ) of ACF and various concentrations of 5-FU, or pretreated with the IC 30 of ACF and various concentrations of 5-FU. To assess the mechanism of action of ACF, cells were treated with IC 30 values of the compound and then the reverse transcription-quantitative polymerase chain reaction was used to evaluate mRNA levels of hypoxia-inducible factor-1α (HIF-1α) and topoisomerase 2. Results indicate that pretreatment with ACF markedly sensitized CRC cells to the cytotoxic effects of 5-FU, whereas simultaneous treatment with ACF and 5-FU were not able to alter the resistance of CRC cells to 5-FU. In comparison with irinotecan, ACF was a more potent agent for enhancing the antitumor activity of 5-FU. ACF did not alter the mRNA levels of either HIF-1α or topoisomerase 2. The results of the present study reveal for the first time that pretreatment of CRC cells with ACF markedly increases the cytotoxic effects of 5-FU, regardless of the p53 status of cells.

Integrative Genomics Reveals Mechanisms of Copy Number Alterations Responsible for Transcriptional Deregulation in Colorectal Cancer

PubMed Central

Camps, Jordi; Nguyen, Quang Tri; Padilla-Nash, Hesed M.; Knutsen, Turid; McNeil, Nicole E.; Wangsa, Danny; Hummon, Amanda B.; Grade, Marian; Ried, Thomas; Difilippantonio, Michael J.

2016-01-01

To evaluate the mechanisms and consequences of chromosomal aberrations in colorectal cancer (CRC), we used a combination of spectral karyotyping, array comparative genomic hybridization (aCGH), and array-based global gene expression profiling on 31 primary carcinomas and 15 established cell lines. Importantly, aCGH showed that the genomic profiles of primary tumors are recapitulated in the cell lines. We revealed a preponderance of chromosome breakpoints at sites of copy number variants (CNVs) in the CRC cell lines, a novel mechanism of DNA breakage in cancer. The integration of gene expression and aCGH led to the identification of 157 genes localized within high-level copy number changes whose transcriptional deregulation was significantly affected across all of the samples, thereby suggesting that these genes play a functional role in CRC. Genomic amplification at 8q24 was the most recurrent event and led to the overexpression of MYC and FAM84B. Copy number dependent gene expression resulted in deregulation of known cancer genes such as APC, FGFR2, and ERBB2. The identification of only 36 genes whose localization near a breakpoint could account for their observed deregulated expression demonstrates that the major mechanism for transcriptional deregulation in CRC is genomic copy number changes resulting from chromosomal aberrations. PMID:19691111

JC Virus Mediates Invasion and Migration in Colorectal Metastasis

PubMed Central

Link, Alexander; Shin, Sung Kwan; Nagasaka, Takeshi; Balaguer, Francesc; Koi, Minoru; Jung, Barbara; Boland, C. Richard; Goel, Ajay

2009-01-01

Introduction JC Virus (JCV), a human polyomavirus, is frequently present in colorectal cancers (CRCs). JCV large T-Ag (T-Ag) expressed in approximately half of all CRC's, however, its functional role in CRC is poorly understood. We hypothesized that JCV T-Ag may mediate metastasis in CRC cells through increased migration and invasion. Material and Methods CRC cell lines (HCT116 and SW837) were stably transfected with JCV early transcript sequences cloned into pCR3 or empty vectors. Migration and invasion assays were performed using Boyden chambers. Global gene expression analysis was performed to identify genetic targets and pathways altered by T-Ag expression. Microarray results were validated by qRT-PCR, protein expression analyses and immunohistochemistry. Matching primary CRCs and liver metastases from 33 patients were analyzed for T-Ag expression by immunohistochemistry. Results T-Ag expressing cell lines showed 2 to 3-fold increase in migration and invasion compared to controls. JCV T-Ag expression resulted in differential expression of several genetic targets, including genes that mediate cell migration and invasion. Pathway analysis suggested a significant involvement of these genes with AKT and MAPK signaling. Treatment with selective PI3K/AKT and MAPK pathway inhibitors resulted in reduced migration and invasion. In support of our in-vitro results, immunohistochemical staining of the advanced stage tumors revealed frequent JCV T-Ag expression in metastatic primary tumors (92%) as well as in their matching liver metastasis (73%). Conclusion These data suggest that JCV T-Ag expression in CRC associates with a metastatic phenotype, which may partly be mediated through the AKT/MAPK signaling pathway. Frequent expression of JCV T-Ag in CRC liver metastasis provides further clues supporting a mechanistic role for JCV as a possible mediator of cellular motility and invasion in CRC. PMID:19997600

Down-regulation of long non-coding RNA RP11-708H21.4 is associated with poor prognosis for colorectal cancer and promotes tumorigenesis through regulating AKT/mTOR pathway.

PubMed

Sun, Longci; Jiang, Chunhui; Xu, Chunjie; Xue, Hanbing; Zhou, Hong; Gu, Lei; Liu, Ye; Xu, Qing

2017-04-25

Long non-coding RNAs (lncRNAs) serve critical roles in cancer development and progression. Herein, through next generation RNA sequencing and experimental validations, we determined the expression status of RP11-708H21.4 in colorectal cancer (CRC) and explored its clinical significance and biological functions in CRC. Differentially expressed lncRNAs from CRC samples and corresponding normal mucosa tissues was screened through RNA sequencing, and RP11-708H21.4 was selected for further experimental validation. The expression levels of RP11-708H21.4 in CRC tissues and cell lines were determined using qRT-PCR. Also, the relationship between the clinicopathological features and RP11-708H21.4 expression was analyzed. Cell viability was examined by CCK-8 and colony assays; cell migration and invasion were detected by transwell assays; cell cycle and cell apoptosis were analyzed by flow cytometry. The chemosensitivity of CRC cells to 5-Fluorouracil (5-FU) was also determined using CCK-8 assay. CRC xenograft tumor models were established to determine the biological functions of RP11-708H21.4 in vivo. Levels of cell cycle-related proteins and AKT/mTOR pathway-related proteins were detected by western blot assay. RP11-708H21.4 expression was aberrantly decreased in CRC, and its expression was closely associated with aggressive clinicopathologic features and unfavorable prognosis of CRC patients. Overexpressed RP11-708H21.4 suppresses CRC cell proliferation through inducing G1 arrest. Moreover, up-regulation of RP11-708H21.4 inhibits cell migration and invasion, causes cell apoptosis, and enhances 5-FU sensitivity of CRC cells. Finally, increased RP11-708H21.4 expression blocked AKT/mTOR pathway, and repressed in vivo CRC xenograft tumor growth. The results indicated that RP11-708H21.4 might have potential roles as a biomarker and a therapeutic target for CRC.

Tumor-associated macrophages recruit CCR6+ regulatory T cells and promote the development of colorectal cancer via enhancing CCL20 production in mice.

PubMed

Liu, Jinlin; Zhang, Ning; Li, Qun; Zhang, Weiwei; Ke, Fang; Leng, Qibin; Wang, Hong; Chen, Jinfei; Wang, Honglin

2011-04-29

Tumor-associated macrophages (TAMs) remodel the colorectal cancer (CRC) microenvironment. Yet, findings on the role of TAMs in CRC seem to be contradictory compared with other cancers. FoxP3(+) regulatory T (Treg)-cells dominantly infiltrate CRC. However, the underlying molecular mechanism in which TAMs may contribute to the trafficking of Treg-cells to the tumor mass remains unknown. CRC was either induced by N-methyl-N-nitrosourea (MNU) and H. pylori or established by subcutaneous injection of mouse colorectal tumor cell line (CMT93) in mice. CMT93 cells were co-cultured with primary macrophages in a transwell apparatus. Recruitment of FoxP3 green fluorescence protein positive (FoxP3(GFP+)) Treg-cells was assessed using the IVIS Imaging System or immunofluorescence staining. A role for macrophages in trafficking of Treg-cells and in the development of CRC was investigated in CD11b diphtheria toxin receptor (CD11b-DTR) transgenic C57BL/6J mice in which macrophages can be selectively depleted. Treg-cells remarkably infiltrated solid tumor, and predominantly expressed the homing chemokine receptor (CCR) 6 in the induced CRC model. Both CMT93 cancer cells and macrophages produced a large amount of CCL20, the sole ligand of CCR6 in vitro and in vivo. Injection of recombinant mouse CCL20 into tumor sites promoted its development with a marked recruitment of Treg-cells in the graft CRC model. Conditional macrophage ablation decreased CCL20 levels, blocked Treg-cell recruitment and inhibited tumor growth in CD11b-DTR mice grafted with CMT93. TAMs recruit CCR6(+) Treg-cells to tumor mass and promote its development via enhancing the production of CCL20 in a CRC mouse model.

In Vitro Killing of Colorectal Carcinoma Cells by Autologous Activated NK Cells is Boosted by Anti-Epidermal Growth Factor Receptor-induced ADCC Regardless of RAS Mutation Status.

PubMed

Turin, Ilaria; Delfanti, Sara; Ferulli, Federica; Brugnatelli, Silvia; Tanzi, Matteo; Maestri, Marcello; Cobianchi, Lorenzo; Lisini, Daniela; Luinetti, Ombretta; Paulli, Marco; Perotti, Cesare; Todisco, Elisabetta; Pedrazzoli, Paolo; Montagna, Daniela

2018-05-01

Treatment of advanced metastatic colorectal cancer (mCRC) patients is associated with a poor prognosis and significant morbidity. Moreover, targeted therapies such as anti-epidermal growth factor receptor (EGFR) have no effect in metastatic patients with tumors harboring a mutation in the RAS gene. The failure of conventional treatment to improve outcomes in mCRC patients has prompted the development of adoptive immunotherapy approaches including natural killer (NK)-based therapies. In this study, after confirmation that patients' NK cells were not impaired in their cytotoxic activity, evaluated against long-term tumor cell lines, we evaluated their interactions with autologous mCRC cells. Molecular and phenotypical evaluation of mCRC cells, expanded in vitro from liver metastasis, showed that they expressed high levels of polio virus receptor and Nectin-2, whereas UL16-binding proteins were less expressed in all tumor samples evaluated. Two different patterns of MICA/B and HLA class I expression on the membrane of mCRC were documented; approximately half of mCRC patients expressed high levels of these molecules on the membrane surface, whereas, in the remaining, very low levels were documented. Resting NK cells were unable to display sizeable levels of cytotoxic activity against mCRC cells, whereas their cytotoxic activity was enhanced after overnight or 5-day incubation with IL-2 or IL-15. The susceptibility of NK-mediated mCRC lysis was further significantly enhanced after coating with cetuximab, irrespective of their RAS mutation and HLA class I expression. These data open perspectives for combined NK-based immunotherapy with anti-epidermal growth factor receptor antibodies in a cohort of mCRC patients with a poor prognosis refractory to conventional therapies.

H19 mediates methotrexate resistance in colorectal cancer through activating Wnt/β-catenin pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Ke-feng; Liang, Wei-Cheng; Feng, Lu

Colorectal cancer (CRC) is a common malignancy, most of which remain unresponsive to chemotherapy. As one of the earliest cytotoxic drugs, methotrexate (MTX) serves as an anti-metabolite and anti-folate chemotherapy for various cancers. Unfortunately, MTX resistance prevents its clinical application in cancer therapy. Thereby, overcoming the drug resistance is an alternative strategy to maximize the therapeutic efficacy of MTX in clinics. Long noncoding RNAs (lncRNAs) have gained widespread attention in recent years. More and more emerging evidences have demonstrated that they play important regulatory roles in various biological activities and disease progression including drug resistance. In the present study, amore » MTX-resistant colorectal cell line HT-29 (HT-29-R) was developed, which displayed the active proliferation and shortened cell cycle. LncRNA H19 was found to be significantly upregulated in this resistant cell line. Further investigation showed that H19 knockdown sensitized the MTX resistance in HT-29-R cells while its overexpression improved the MTX resistance in the parental cells, suggesting that H19 mediate MTX resistance. The Wnt/β-catenin signaling was activated in HT-29-R cells, and H19 knockdown suppressed this signaling in the parental cells. In conclusion, H19 mediated MTX resistance via activating Wnt/β-catenin signaling, which help to develop H19 as a promising therapeutic target for MTX resistant CRC. - Highlights: • A methotrexate (MTX) -resistant colorectal cancer cell line HT-29 (HT-29-R) has been developed. • H19 was upregulated in HT-29-R cells. • H19 mediated MTX resistance in colorectal cancer (CRC). • Wnt/β-catenin pathway was involved in the H19-mediated MTX resistance in CRC cells.« less

Pokemon enhances proliferation, cell cycle progression and anti-apoptosis activity of colorectal cancer independently of p14ARF-MDM2-p53 pathway.

PubMed

Zhao, Yi; Yao, Yun-hong; Li, Li; An, Wei-fang; Chen, Hong-zen; Sun, Li-ping; Kang, Hai-xian; Wang, Sen; Hu, Xin-rong

2014-12-01

Pokemon has been showed to directly suppress p14(ARF) expression and also to overexpress in multiple cancers. However, p14(ARF)-MDM2-p53 pathway is usually aberrant in colorectal cancer (CRC). The aim is to confirm whether Pokemon plays a role in CRC and explore whether Pokemon works through p14(ARF)-MDM2-p53 pathway in CRC. Immunohistochemistry for Pokemon, p14(ARF) and Mtp53 protein was applied to 45 colorectal epitheliums (CREs), 42 colorectal adenomas (CRAs) and 66 CRCs. Pokemon was knocked down with RNAi technique in CRC cell line Lovo to detect mRNA expression of p14(ARF) with qRT-PCR, cell proliferation with CCK8 assay, and cell cycle and apoptosis with flowcytometry analysis. The protein expression rates were significantly higher in CRC (75.8%) than in CRE (22.2 %) or CRA (38.1%) for Pokemon and higher in CRC (53.0%) than in CRE (0) or CRA (4.8%) for Mtp53, but not significantly different in CRC (86.4 %) versus CRE (93.3%) or CRA (90.5 %) for p14(ARF). Higher expression rate of Pokemon was associated with lymph node metastasis and higher Duke's stage. After knockdown of Pokemon in Lovo cells, the mRNA level of p14(ARF) was not significantly changed, the cell proliferation ability was decreased by 20.6%, cell cycle was arrested by 55.7% in G0/G1 phase, and apoptosis rate was increased by 19.0%. Pokemon enhanced the oncogenesis of CRC by promoting proliferation, cell cycle progression and anti-apoptosis activity of CRC cells independently of p14(ARF)-MDM2-p53 pathway. This finding provided a novel idea for understanding and further studying the molecular mechanism of Pokemon on carcinogenesis of CRC.

Carnosic acid inhibits the proliferation and migration capacity of human colorectal cancer cells

PubMed Central

BARNI, M.V.; CARLINI, M.J.; CAFFERATA, E.G.; PURICELLI, L.; MORENO, S.

2012-01-01

Colorectal cancer (CRC) is the third most common malignant neoplasm worldwide. The objective of this study was to examine whether carnosic acid (CA), the main antioxidant compound of Rosmarinus officinalis L., would inhibit the cell viability of three CRC cell lines: Caco-2, HT29 and LoVo in a dose-dependent manner, with IC50 values in the range of 24–96 μM. CA induced cell death by apoptosis in Caco-2 line after 24 h of treatment and inhibited cell adhesion and migration, possibly by reducing the activity of secreted proteases such as urokinase plasminogen activator (uPA) and metalloproteinases (MMPs). These effects may be associated through a mechanism involving the inhibition of the COX-2 pathway, because we have determined that CA downregulates the expression of COX-2 in Caco-2 cells at both the mRNA and protein levels. Therefore, CA modulates different targets involved in the development of CRC. These findings indicate that carnosic acid may have anticancer activity and may be useful as a novel chemotherapeutic agent. PMID:22246562

NSC30049 inhibits Chk1 pathway in 5-FU-resistant CRC bulk and stem cell populations.

PubMed

Narayan, Satya; Jaiswal, Aruna S; Sharma, Ritika; Nawab, Akbar; Duckworth, Lizette Vila; Law, Brian K; Zajac-Kaye, Maria; George, Thomas J; Sharma, Jay; Sharma, Arun K; Hromas, Robert A

2017-08-22

The 5-fluorouracil (5-FU) treatment induces DNA damage and stalling of DNA replication forks. These stalled replication forks then collapse to form one sided double-strand breaks, leading to apoptosis. However, colorectal cancer (CRC) stem cells rapidly repair the stalled/collapsed replication forks and overcome treatment effects. Recent evidence suggests a critical role of checkpoint kinase 1 (Chk1) in preventing the replicative stress. Therefore, Chk1 kinase has been a target for developing mono or combination therapeutic agents. In the present study, we have identified a novel orphan molecule NSC30049 (NSC49L) that is effective alone, and in combination potentiates 5-FU-mediated growth inhibition of CRC heterogeneous bulk and FOLFOX-resistant cell lines in culture with minimal effect on normal colonic epithelial cells. It also inhibits the sphere forming activity of CRC stem cells, and decreases the expression levels of mRNAs of CRC stem cell marker genes. Results showed that NSC49L induces 5-FU-mediated S-phase cell cycle arrest due to increased load of DNA damage and increased γ-H2AX staining as a mechanism of cytotoxicity. The pharmacokinetic analysis showed a higher bioavailability of this compound, however, with a short plasma half-life. The drug is highly tolerated by animals with no pathological aberrations. Furthermore, NSC49L showed very potent activity in a HDTX model of CRC stem cell tumors either alone or in combination with 5-FU. Thus, NSC49L as a single agent or combined with 5-FU can be developed as a therapeutic agent by targeting the Chk1 pathway in 5-FU-resistant CRC heterogeneous bulk and CRC stem cell populations.

Targeting autophagy enhances apatinib-induced apoptosis via endoplasmic reticulum stress for human colorectal cancer.

PubMed

Cheng, Xi; Feng, Haoran; Wu, Haoxuan; Jin, Zhijian; Shen, Xiaonan; Kuang, Jie; Huo, Zhen; Chen, Xianze; Gao, Haoji; Ye, Feng; Ji, Xiaopin; Jing, Xiaoqian; Zhang, Yaqi; Zhang, Tao; Qiu, Weihua; Zhao, Ren

2018-05-30

Apatinib, a novel tyrosine kinase inhibitor (TKI), has been confirmed for its efficacy and safety in the treatment of advanced gastric carcinoma and some other solid tumors. However, the direct functional mechanisms of tumor lethality mediated by apatinib have not yet been fully characterized, and the precise mechanisms of drug resistance are largely unknown. Here, in this study, we demonstrated that apatinib could induce both apoptosis and autophagy in human colorectal cancer (CRC) via a mechanism that involved endoplasmic reticulum (ER) stress. Moreover, activation of the IRE1α pathway from apatinib-induced ER stress is responsible for the induction of autophagy; however, blocking autophagy could enhance the apoptosis in apatinib-treated human CRC cell lines. Furthermore, the combination of apatinib with autophagy inhibitor chloroquine (CQ) tends to have the most significant anti-tumor effect of CRC both in vitro and in vivo. Overall, our data show that because apatinib treatment could induce ER stress-related apoptosis and protective autophagy in human CRC cell lines, targeting autophagy is a promising therapeutic strategy to relieve apatinib drug resistance in CRC. Copyright © 2018 The Author(s). Published by Elsevier B.V. All rights reserved.

Cell-surface markers for colon adenoma and adenocarcinoma

PubMed Central

Sewda, Kamini; Coppola, Domenico; Enkemann, Steven; Yue, Binglin; Kim, Jongphil; Lopez, Alexis S.; Wojtkowiak, Jonathan W.; Stark, Valerie E.; Morse, Brian; Shibata, David; Vignesh, Shivakumar; Morse, David L.

2016-01-01

Early detection of colorectal cancer (CRC) is crucial for effective treatment. Among CRC screening techniques, optical colonoscopy is widely considered the gold standard. However, it is a costly and invasive procedure with a low rate of compliance. Our long-term goal is to develop molecular imaging agents for the non-invasive detection of CRC by molecular imaging-based colonoscopy using CT, MRI or fluorescence. To achieve this, cell surface targets must be identified and validated. Here, we report the discovery of cell-surface markers that distinguish CRC from surrounding tissues that could be used as molecular imaging targets. Profiling of mRNA expression microarray data from patient tissues including adenoma, adenocarcinoma, and normal gastrointestinal tissues was used to identify potential CRC specific cell-surface markers. Of the identified markers, six were selected for further validation (CLDN1, GPR56, GRM8, LY6G6D/F, SLCO1B3 and TLR4). Protein expression was confirmed by immunohistochemistry of patient tissues. Except for SLCO1B3, diffuse and low expression was observed for each marker in normal colon tissues. The three markers with the greatest protein overexpression were CLDN1, LY6G6D/F and TLR4, where at least one of these markers was overexpressed in 97% of the CRC samples. GPR56, LY6G6D/F and SLCO1B3 protein expression was significantly correlated with the proximal tumor location and with expression of mismatch repair genes. Marker expression was further validated in CRC cell lines. Hence, three cell-surface markers were discovered that distinguish CRC from surrounding normal tissues. These markers can be used to develop imaging or therapeutic agents targeted to the luminal surface of CRC. PMID:26894861

Cell-surface markers for colon adenoma and adenocarcinoma.

PubMed

Sewda, Kamini; Coppola, Domenico; Enkemann, Steven; Yue, Binglin; Kim, Jongphil; Lopez, Alexis S; Wojtkowiak, Jonathan W; Stark, Valerie E; Morse, Brian; Shibata, David; Vignesh, Shivakumar; Morse, David L

2016-04-05

Early detection of colorectal cancer (CRC) is crucial for effective treatment. Among CRC screening techniques, optical colonoscopy is widely considered the gold standard. However, it is a costly and invasive procedure with a low rate of compliance. Our long-term goal is to develop molecular imaging agents for the non-invasive detection of CRC by molecular imaging-based colonoscopy using CT, MRI or fluorescence. To achieve this, cell surface targets must be identified and validated. Here, we report the discovery of cell-surface markers that distinguish CRC from surrounding tissues that could be used as molecular imaging targets. Profiling of mRNA expression microarray data from patient tissues including adenoma, adenocarcinoma, and normal gastrointestinal tissues was used to identify potential CRC specific cell-surface markers. Of the identified markers, six were selected for further validation (CLDN1, GPR56, GRM8, LY6G6D/F, SLCO1B3 and TLR4). Protein expression was confirmed by immunohistochemistry of patient tissues. Except for SLCO1B3, diffuse and low expression was observed for each marker in normal colon tissues. The three markers with the greatest protein overexpression were CLDN1, LY6G6D/F and TLR4, where at least one of these markers was overexpressed in 97% of the CRC samples. GPR56, LY6G6D/F and SLCO1B3 protein expression was significantly correlated with the proximal tumor location and with expression of mismatch repair genes. Marker expression was further validated in CRC cell lines. Hence, three cell-surface markers were discovered that distinguish CRC from surrounding normal tissues. These markers can be used to develop imaging or therapeutic agents targeted to the luminal surface of CRC.

Detection of novel and potentially actionable anaplastic lymphoma kinase (ALK) rearrangement in colorectal adenocarcinoma by immunohistochemistry screening

PubMed Central

Wang, Kai; Kim, Sun Young; Jang, Jiryeon; Kim, Seung Tae; Park, Joon Oh; Lim, Ho Yeong; Kang, Won Ki; Park, Young Suk; Lee, Jiyun; Lee, Woo Yong; Park, Yoon Ah; Huh, Jung Wook; Yun, Seong Hyeon; Do, In-Gu; Kim, Seok Hyung; Balasubramanian, Sohail; Stephens, Philip J.; Ross, Jeffrey S.; Li, Gang Gary; Hornby, Zachary; Ali, Siraj M.; Miller, Vincent A.; Kim, Kyoung-Mee; Ou, Sai-Hong Ignatius

2015-01-01

Purpose Anaplastic lymphoma kinase (ALK) rearrangement has been detected in colorectal carcinoma (CRC) using advanced molecular diagnostics tests including exon scanning, fluorescence in situ hybridization (FISH), and next generation sequencing (NGS). We investigated if immunohistochemistry (IHC) can be used to detect ALK rearrangement in gastrointestinal malignancies. Experimental designs Tissue microarrays (TMAs) from consecutive gastric carcinoma (GC) and CRC patients who underwent surgical resection at Samsung Medical Center, Seoul, Korea were screened by IHC using ALK monoclonal antibody 5A4. IHC positive cases were confirmed by FISH, nCounter assays, and NGS-based comprehensive genomic profiling (CGP). ALK IHC was further applied to CRC patients enrolled in a pathway-directed therapeutic trial. Results Four hundred thirty-two GC and 172 CRC cases were screened by IHC. No GC sample was ALK IHC positive. One CRC (0.6%) was ALK IHC positive (3+) that was confirmed by ALK FISH and a novel CAD-ALK (C35; A20) fusion variant that resulted from a paracentric inversion event inv(2)(p22–21p23) was identified by CGP. One out of 50 CRC patients enrolled in a pathway-directed therapeutic trial was ALK IHC positive (3+) confirmed by ALK FISH and found to harbor the EML4-ALK (E21, A20) fusion variant by CGP. Growth of a tumor cell line derived from this EML4-ALK CRC patient was inhibited by ALK inhibitors crizotinib and entrectinib. Conclusions ALK IHC is a viable screening strategy for identifying ALK rearrangement in CRC. ALK rearrangement is a potential actionable driver mutation in CRC based on survival inhibition of patient tumor-derived cell line by potent ALK inhibitors. PMID:26172300

Opposing roles of the aldo-keto reductases AKR1B1 and AKR1B10 in colorectal cancer.

PubMed

Taskoparan, Betul; Seza, Esin Gulce; Demirkol, Secil; Tuncer, Sinem; Stefek, Milan; Gure, Ali Osmay; Banerjee, Sreeparna

2017-12-01

Aldo-keto reductases (including AKR1B1 and AKR1B10) constitute a family of oxidoreductases that have been implicated in the pathophysiology of diabetes and cancer, including colorectal cancer (CRC). Available data indicate that, despite their similarities in structure and enzymatic functions, their roles in CRC may be divergent. Here, we aimed to determine the expression and functional implications of AKR1B1 and AKR1B10 in CRC. AKR1B1 and AKR1B10 gene expression levels were analyzed using publicly available microarray data and ex vivo CRC-derived cDNA samples. Gene Set Enrichment Analysis (GSEA), The Cancer Genome Atlas (TCGA) RNA-seq data and The Cancer Proteome Atlas (TCPA) proteome data were analyzed to determine the effect of high and low AKR1B1 and AKR1B10 expression levels in CRC patients. Proliferation, cell cycle progression, cellular motility, adhesion and inflammation were determined in CRC-derived cell lines in which these genes were either exogenously overexpressed or silenced. We found that the expression of AKR1B1 was unaltered, whereas that of AKR1B10 was decreased in primary CRCs. GSEA revealed that, while high AKR1B1 expression was associated with increased cell cycle progression, cellular motility and inflammation, high AKR1B10 expression was associated with a weak inflammatory phenotype. Functional studies carried out in CRC-derived cell lines confirmed these data. Microarray data analysis indicated that high expression levels of AKR1B1 and AKR1B10 were significantly associated with shorter and longer disease-free survival rates, respectively. A combined gene expression signature of AKR1B10 (low) and AKR1B1 (high) showed a better prognostic stratification of CRC patients independent of confounding factors. Despite their similarities, the expression levels and functions of AKR1B1 and AKR1B10 are highly divergent in CRC, and they may have prognostic implications.

IN-VITRO evidence for the protective properties of the main components of the Mediterranean diet against colorectal cancer: A systematic review.

PubMed

Rotelli, M T; Bocale, D; De Fazio, M; Ancona, P; Scalera, I; Memeo, R; Travaglio, E; Zbar, A P; Altomare, D F

2015-09-01

Epidemiological studies have shown that the incidence and mortality rates of colorectal cancer (CRC) vary over 10-fold worldwide where within Westernized societies lower rates are observed amongst populations living within the Mediterranean basin, suggesting a significant influence of environment and dietary style in CRC carcinogenesis. Interpretation of the data concerning the benefits of mediterranean (MD) diet is difficult in vivo because of the variability of alimentary regimens used, the differing compliance with dietary supplementation and because of the non-uniform duration of patient cohort observation. Therefore, the aim of this review is to evaluate the in-vitro effects on colorectal cancer cell lines. the literature concerning the in-vitro effects of 4 of the principal components symbolizing the MD such as olive oil (polyphenol), red chili (capsaicin), tomato (lycopene) and red grapes (resveratrol) have been systematically reviewed. Several studies have demonstrated that polyphenols form olive oil, lycopene, resveratrol and capsaicin have multiple anticancer properties affecting several metabolic pathways involved in cancerogenesis, apoptosis, and metastasis in CRC cell lines. This review summarizes some of the most recent data potentially supportive of the use of MD in CRC chemoprevention, analyzing the in vitro effects of individual components of the MD on CRC cell development, progression, metastasis and apoptosis. Copyright © 2015 Elsevier Ltd. All rights reserved.

ShaoYao decoction ameliorates colitis-associated colorectal cancer by downregulating proinflammatory cytokines and promoting epithelial-mesenchymal transition

PubMed Central

2014-01-01

Abstracts Background Shaoyao decoction (SYD) is a traditional Chinese medicine prescription formulated by Liu Wan-Su, a master of traditional Chinese medicine in Jin-Yuan Dynasty. SYD is effective in treating ulcerative colitis. Paeonol, a component of SYD, inhibits colorectal cancer (CRC) cell proliferation and induces CRC cell apoptosis. In this study, azoxymethane (AOM)/dextran sodium sulfate (DSS)–induced colitis-associated CRC (caCRC) model and CRC cell lines were used to examine the effects of SYD on CRC in vivo and in vitro. Methods A translational medicine strategy based on phytomics quality control was adopted. Liquid chromatography was employed for the chemical characterization and chemical fingerprinting of SYD. Protein expression and macrophage existence were determined by immunohistochemistry and western blot. Serum cytokines were quantified by Luminex assay. Results AOM/DSS-induced caCRC phenotypically resembled human caCRC. SYD significantly increased the survival rate of the mice, ameliorated the general well-being of the mice, and reduced the incidence and multiplicity of colonic neoplasms. SYD inhibited epithelial–mesenchymal transition (EMT), as indicated by upregulated epithelia cadherin and downregulated neuronal cadherin, fibronectin, vimentin, and transcription factor Snail. SYD reduced the expression levels of serum interleukin 1β, interleukin-6, tumor necrosis factor α, tumor-associated macrophages, and p65. These results showed that SYD can attenuate proinflammatory cytokines and inhibit EMT. Conclusions SYD ameliorates caCRC by suppressing inflammation and inhibiting EMT. SYD might be an alternative therapy for caCRC. PMID:24766737

Umbilical cord blood-derived natural killer cells combined with Bevacizumab for colorectal cancer treatment.

PubMed

Xu, Chen; Liu, Dongning; Chen, Zhixin; Zhuo, Fan; Sun, Huankui; Hu, Jiaping; Li, Taiyuan

2018-06-19

Colorectal cancer (CRC) is among cancers with highest incidence globally and currently ranks fourth as the leading cause of cancer-related deaths worldwide. It remains an urgent need for novel strategies in the management of patients with advanced CRC. Adoptive transfer of allogeneic natural killer (NK) cells represent an attractive option in the treatment of patients with CRC. In this study, we successfully expanded NK cells from umbilical cord blood (UCB) with membrane-bound IL-21, termed eUCB-NK cells. eUCB-NK cells efficiently lysed CRC cell lines in vitro and secreted significantly higher levels of IFN-γ, TNF-α, GM-CSF and CCL3 compared with IL-2 stimulated NK cells. Adoptive transfer of these NK cells significantly inhibited the growth of HT29 xenografts, whereas LoVo tumors were not effectively controlled with eUCB-NK cells. More NK cells inside HT29 tumors, not seen in LoVo tumors, might contribute to the differences in response to eUCB-NK cells. Combination of bevacizumab can increase extravasation of adoptively transferred NK cells into the LoVo tumors and improve the therapeutic activity of eUCB-NK cells. These results justified clinical translation of this UCB-derived NK cell-based therapeutics, either used alone or combined with bevacizumab, as a novel treatment option for patients with CRC.

CRHR2/Ucn2 signaling is a novel regulator of miR-7/YY1/Fas circuitry contributing to reversal of colorectal cancer cell resistance to Fas-mediated apoptosis.

PubMed

Pothoulakis, Charalabos; Torre-Rojas, Monica; Duran-Padilla, Marco A; Gevorkian, Jonathan; Zoras, Odysseas; Chrysos, Emmanuel; Chalkiadakis, George; Baritaki, Stavroula

2018-01-15

Colorectal cancer (CRC) responds poorly to immuno-mediated cytotoxicity. Underexpression of corticotropin-releasing-hormone-receptor-2 (CRHR2) in CRC, promotes tumor survival, growth and Epithelial to Mesenchymal Transition (EMT), in vitro and in vivo. We explored the role of CRHR2 downregulation in CRC cell resistance to Fas/FasL-mediated apoptosis and the underlying molecular mechanism. CRC cell sensitivity to CH11-induced apoptosis was compared between Urocortin-2 (Ucn2)-stimulated parental and CRHR2-overexpressing CRC cell lines and targets of CRHR2/Ucn2 signaling were identified through in vitro and ex vivo analyses. Induced CRHR2/Ucn2 signaling in SW620 and DLD1 cells increased specifically their sensitivity to CH11-mediated apoptosis, via Fas mRNA and protein upregulation. CRC compared to control tissues had reduced Fas expression that was associated with lost CRHR2 mRNA, poor tumor differentiation and high risk for distant metastasis. YY1 silencing increased Fas promoter activity in SW620 and re-sensitized them to CH11-apoptosis, thus suggesting YY1 as a putative transcriptional repressor of Fas in CRC. An inverse correlation between Fas and YY1 expression was confirmed in CRC tissue arrays, while elevated YY1 mRNA was clinically relevant with advanced CRC grade and higher risk for distant metastasis. CRHR2/Ucn2 signaling downregulated specifically YY1 expression through miR-7 elevation, while miR-7 modulation in miR-7 high SW620-CRHR2+ and miR-7 low HCT116 cells, had opposite effects on YY1 and Fas expressions and cell sensitivity to CH11-killing. CRHR2/Ucn2 signaling is a negative regulator of CRC cell resistance to Fas/FasL-apoptosis via targeting the miR-7/YY1/Fas circuitry. CRHR2 restoration might prove effective in managing CRC response to immune-mediated apoptotic stimuli. © 2017 UICC.

Tumor-Associated Macrophages Recruit CCR6+ Regulatory T Cells and Promote the Development of Colorectal Cancer via Enhancing CCL20 Production in Mice

PubMed Central

Li, Qun; Zhang, Weiwei; Ke, Fang; Leng, Qibin; Wang, Hong; Chen, Jinfei; Wang, Honglin

2011-01-01

Background Tumor-associated macrophages (TAMs) remodel the colorectal cancer (CRC) microenvironment. Yet, findings on the role of TAMs in CRC seem to be contradictory compared with other cancers. FoxP3+ regulatory T (Treg)-cells dominantly infiltrate CRC. However, the underlying molecular mechanism in which TAMs may contribute to the trafficking of Treg-cells to the tumor mass remains unknown. Methodology/Principal Findings CRC was either induced by N-methyl-N-nitrosourea (MNU) and H. pylori or established by subcutaneous injection of mouse colorectal tumor cell line (CMT93) in mice. CMT93 cells were co-cultured with primary macrophages in a transwell apparatus. Recruitment of FoxP3 green fluorescence protein positive (FoxP3GFP+) Treg-cells was assessed using the IVIS Imaging System or immunofluorescence staining. A role for macrophages in trafficking of Treg-cells and in the development of CRC was investigated in CD11b diphtheria toxin receptor (CD11b-DTR) transgenic C57BL/6J mice in which macrophages can be selectively depleted. Treg-cells remarkably infiltrated solid tumor, and predominantly expressed the homing chemokine receptor (CCR) 6 in the induced CRC model. Both CMT93 cancer cells and macrophages produced a large amount of CCL20, the sole ligand of CCR6 in vitro and in vivo. Injection of recombinant mouse CCL20 into tumor sites promoted its development with a marked recruitment of Treg-cells in the graft CRC model. Conditional macrophage ablation decreased CCL20 levels, blocked Treg-cell recruitment and inhibited tumor growth in CD11b-DTR mice grafted with CMT93. Conclusions/Significance TAMs recruit CCR6+ Treg-cells to tumor mass and promote its development via enhancing the production of CCL20 in a CRC mouse model. PMID:21559338

Cytoskeleton-centric protein transportation by exosomes transforms tumor-favorable macrophages.

PubMed

Chen, Zhipeng; Yang, Lijuan; Cui, Yizhi; Zhou, Yanlong; Yin, Xingfeng; Guo, Jiahui; Zhang, Gong; Wang, Tong; He, Qing-Yu

2016-10-11

The exosome is a key initiator of pre-metastatic niche in numerous cancers, where macrophages serve as primary inducers of tumor microenvironment. However, the proteome that can be exosomally transported from cancer cells to macrophages has not been sufficiently characterized so far. Here, we used colorectal cancer (CRC) exosomes to educate tumor-favorable macrophages. With a SILAC-based mass spectrometry strategy, we successfully traced the proteome transported from CRC exosomes to macrophages. Such a proteome primarily focused on promoting cytoskeleton rearrangement, which was biologically validated with multiple cell lines. We reproduced the exosomal transportation of functional vimentin as a proof-of-concept example. In addition, we found that some CRC exosomes could be recognized by macrophages via Fc receptors. Therefore, we revealed the active and necessary role of exosomes secreted from CRC cells to transform cancer-favorable macrophages, with the cytoskeleton-centric proteins serving as the top functional unit.

EphA2 expression is a key driver of migration and invasion and a poor prognostic marker in colorectal cancer

PubMed Central

Blayney, Jaine K.; McArt, Darragh G.; Redmond, Keara L.; Weir, Jessica-Anne; Bradley, Conor A.; Sasazuki, Takehiko; Shirasawa, Senji; Wang, Tingting; Srivastava, Supriya; Ong, Chee Wee; Arthur, Ken; Salto-Tellez, Manuel; Wilson, Richard H.

2015-01-01

Purpose EphA2, a member of the Eph receptor tyrosine kinases family, is an important regulator of tumour initiation, neo-vascularization and metastasis in a wide range of epithelial and mesenchymal cancers, however its role in colorectal cancer (CRC) recurrence and progression is unclear. Experimental Design EphA2 expression was determined by immunohistochemistry in stage II/III colorectal tumours (N=338), and findings correlated with clinical outcome. The correlation between EphA2 expression and stem cell markers CD44 and Lgr5 was examined. The role of EphA2 in migration/invasion was assessed using a panel of KRAS wild-type (WT) and mutant (MT) parental and invasive CRC cell line models. Results Colorectal tumours displayed significantly higher expression levels of EphA2 compared with matched normal tissue, which positively correlated with high CD44 and Lgr5 expression levels. Moreover, high EphA2 mRNA and protein expression were found to be associated with poor overall survival in stage II/III CRC tissues, in both univariate and multivariate analyses. Pre-clinically, we found that EphA2 was highly expressed in KRASMT CRC cells and that EphA2 levels are regulated by the KRAS-driven MAPK and RalGDS-RalA pathways. Moreover, EphA2 levels were elevated in several invasive daughter cell lines and down-regulation of EphA2 using RNAi or recombinant EFNA1, suppressed migration and invasion of KRASMT CRC cells. Conclusions These data show that EpHA2 is a poor prognostic marker in stage II/III CRC, which may be due to its ability to promote cell migration and invasion, providing support for the further investigation of EphA2 as a novel prognostic biomarker and therapeutic target. PMID:26283684

Tumor-selective proteotoxicity of verteporfin inhibits colon cancer progression independently of YAP1#

PubMed Central

Zhang, Huabing; Ramakrishnan, Sadeesh K.; Triner, Daniel; Centofanti, Brook; Maitra, Dhiman; Győrffy, Balázs; Sebolt-Leopold, Judith S.; Dame, Michael K.; Varani, James; Brenner, Dean E.; Fearon, Eric R.; Omary, M. Bishr; Shah, Yatrik M.

2016-01-01

Yes-associated protein 1 (YAP1) is a transcriptional coactivator in the Hippo signaling pathway. Increased YAP1- activity promotes the growth of tumors, including that of colorectal cancer (CRC). Verteporfin, a drug that enhances phototherapy to treat neovascular macular degeneration, is an inhibitor of YAP1. Here, we found that verteporfin inhibited tumor growth independently of its effects on YAP1 or the related protein TAZ in genetic or chemical-induced mouse models of CRC, in patient-derived xenografts and in enteroid models of CRC. Instead, verteporfin exhibited in vivo selectivity for killing tumor cells in part by impairing the global clearance of high molecular weight oligomerized proteins, particularly p62 (a sequestrome involved in autophagy) and STAT3 (a transcription factor). Verteporfin inhibited cytokine-induced STAT3 activity and cell proliferation and reduced the viabilty of cultured CRC cells. Although verteporfin accumulated to a greater extent in normal cells than in tumor cells in vivo, experiments with cultured cells indicated that the normal cells efficiently cleared verteporfin-induced protein oligomers through autophagic and proteasomal pathways. Culturing CRC cells in hypoxic or nutrient-deprived conditions (modeling a typical CRC microenvironment) impaired the clearance of protein oligomers and resulted in cell death; whereas culturing cells in normoxic or glucose-replete conditions protected cell viability and proliferation in the presence of verteporfin. Furthermore, verteporfin suppressed the proliferation of other cancer cell lines even in the absence of YAP1, suggesting that verteporfin may be effective against multiple types of solid cancers. PMID:26443705

Preclinical rationale for combination of crizotinib with mitomycin C for the treatment of advanced colorectal cancer.

PubMed

Lev, Avital; Deihimi, Safoora; Shagisultanova, Elena; Xiu, Joanne; Lulla, Amriti R; Dicker, David T; El-Deiry, Wafik S

2017-09-02

Colorectal cancer (CRC) is a leading cause of cancer-related deaths in the United States. We analyzed 26 MSI-High and 558 non-MSI-High CRC tumors. BRCA2 mutations were highly enriched (50%) in MSI-High CRC. Immunohistochemistry showed that BRCA2-mutated MSI-High CRC had high c-MET (64%) expression compared with BRCA-WT (17%). We hypothesized a mechanistic link between BRCA2-deficiency and c-MET overexpression and synergistic interaction between drugs that treat BRCA-deficient tumors (mitomycin C (MMC) or PARP inhibitors) and c-MET inhibitors (crizotinib). We tested CRC cell lines for sensitivity to MMC plus crizotinib or other drug combinations including PARP-inhibitors. Combined treatment of tumor cells with crizotinib and MMC led to increased apoptosis as compared with each drug alone. Additionally, combination treatment with increasing concentrations of both drugs demonstrated a synergistic anti-cancer effect (CI = 0.006-0.74). However, we found no evidence for c-MET upregulation upon effective BRCA2 knockdown in tumor cells -/+DNA damage. Although we found no mechanistic link between BRCA2 deficiency and c-MET overexpression, c-MET is frequently overexpressed in CRC and BRCA2 is mutated especially in MSI-H CRC. The combination of crizotinib with MMC appeared synergistic regardless of MSI or BRCA2 status. Using an in-vivo CRC xenograft model we found reduced tumor growth with combined crizotinib and MMC therapy (p = 0.0088). Our preclinical results support clinical testing of the combination of MMC and crizotinib in advanced CRC. Targeting cell survival mediated by c-MET in combination with targeting DNA repair may be a reasonable strategy for therapy development in CRC or other cancers.

Novel synthetic curcumin analogs as potent antiangiogenic agents in colorectal cancer.

PubMed

Rajitha, Balney; Nagaraju, Ganji Purnachandra; Shaib, Walid L; Alese, Olatunji B; Snyder, James P; Shoji, Mamoru; Pattnaik, Subasini; Alam, Afroz; El-Rayes, Bassel F

2017-01-01

The transcription factor NF-κB plays a central role in angiogenesis in colorectal cancer (CRC). Curcumin is a natural dietary product that inhibits NF-κB. The objective of this study is to evaluate the antiangiogenic effects of curcumin and two potent synthetic analogues (EF31 and UBS109) in CRC. IC 50 values for curcumin, EF31, and UBS109 were determined in the HCT116 and HT-29 cell lines. HUVEC tube formation, egg CAM assay, and matrigel plug assays revealed decreased angiogenesis in cell lines treated with curcumin, EF31, or UBS109. Curcumin and its analogues significantly inhibited VEGF-A synthesis and secretion in both cell lines in association with loss of HIF-1α, COX-2, and p-STAT-3 expression. Nuclear NF-κB expression was inhibited by curcumin, EF31, and UBS109. Transfection of p65-NF-κB in HCT116 and HT-29 cells resulted in increased expression of HIF-1α, COX-2, STAT-3, and VEGF-A. Treatment with curcumin, EF31, or UBS109 inhibited these effects in transfected cell lines. In mice carrying HCT116 and HT-29 cell xenografts, EF31 and UBS109 inhibited subcutaneous tumor growth and potentiated the effects of oxaliplatin and 5-FU. Tumors from treated animals revealed inhibition of HIF-1α, COX-2, p-STAT-3, and VEGF expression. Our findings suggest that inhibition of NF-κB leading to decreased transcription and expression of HIF-1α, COX-2, STAT-3, and VEGF is a rational approach for antiangiogenic therapy in CRC. The distinctive properties of EF31 and UBS109 make them promising therapeutic agents for development in CRC as single agents or as part of combination chemotherapy regimens. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

The cAMP analogs have potent anti-proliferative effects on medullary thyroid cancer cell lines.

PubMed

Dicitore, Alessandra; Grassi, Elisa Stellaria; Caraglia, Michele; Borghi, Maria Orietta; Gaudenzi, Germano; Hofland, Leo J; Persani, Luca; Vitale, Giovanni

2016-01-01

The oncogenic activation of the rearranged during transfection (RET) proto-oncogene has a main role in the pathogenesis of medullary thyroid cancer (MTC). Several lines of evidence suggest that RET function could be influenced by cyclic AMP (cAMP)-dependent protein kinase A (PKA) activity. We evaluated the in vitro anti-tumor activity of 8-chloroadenosine-3',5'-cyclic monophosphate (8-Cl-cAMP) and PKA type I-selective cAMP analogs [equimolar combination of the 8-piperidinoadenosine-3',5'-cyclic monophosphate (8-PIP-cAMP) and 8-hexylaminoadenosine-3',5'-cyclic monophosphate (8-HA-cAMP) in MTC cell lines (TT and MZ-CRC-1)]. 8-Cl-cAMP and the PKA I-selective cAMP analogs showed a potent anti-proliferative effect in both cell lines. In detail, 8-Cl-cAMP blocked significantly the transition of TT cell population from G2/M to G0/G1 phase and from G0/G1 to S phase and of MZ-CRC-1 cells from G0/G1 to S phase. Moreover, 8-Cl-cAMP induced apoptosis in both cell lines, as demonstrated by FACS analysis for annexin V-FITC/propidium iodide, the activation of caspase-3 and PARP cleavage. On the other hand, the only effect induced by PKA I-selective cAMP analogs was a delay in G0/G1-S and S-G2/M progression in TT and MZ-CRC-1 cells, respectively. In conclusion, these data demonstrate that cAMP analogs, particularly 8-Cl-cAMP, significantly suppress in vitro MTC proliferation and provide rationale for a potential clinical use of cAMP analogs in the treatment of advanced MTC.

SCF/C-Kit/JNK/AP-1 Signaling Pathway Promotes Claudin-3 Expression in Colonic Epithelium and Colorectal Carcinoma.

PubMed

Wang, Yaxi; Sun, Tingyi; Sun, Haimei; Yang, Shu; Li, Dandan; Zhou, Deshan

2017-04-06

Claudin-3 is a major protein of tight junctions (TJs) in the intestinal epithelium and is critical for maintaining cell-cell adhesion, barrier function, and epithelium polarity. Recent studies have shown high claudin-3 levels in several solid tumors, but the regulation mechanism of claudin-3 expression remains poorly understood. In the present study, colorectal cancer (CRC) tissues, HT-29 and DLD-1 CRC cell lines, CRC murine model (C57BL/6 mice) and c-kit loss-of-function mutant mice were used. We demonstrated that elevated claudin-3 levels were positively correlated with highly expressed c-kit in CRC tissues based upon analysis of protein expression. In vitro, claudin-3 expression was clearly increased in CRC cells by overexpressed c-kit or stimulated by exogenous recombinant human stem cell factor (rhSCF), while significantly decreased by the treatment with c-kit or c-Jun N-terminal kinase (JNK) inhibitors. Chromatin immunoprecipitation (ChIP) and luciferase reporter assay showed that SCF/c-kit signaling significantly promoted activator protein-1 (AP-1) binding with CLDN-3 promoter and enhanced its transcription activity. Furthermore, decreased expression of claudin-3 was obtained in the colonic epithelium from the c-Kit loss-of-function mutant mice. In conclusion, SCF/c-kit-JNK/AP-1 signaling pathway significantly promoted claudin-3 expression in colonic epithelium and CRC, which could contribute to epithelial barrier function maintenance and to CRC development.

Chemokine Receptor CXCR4 Expression in Patients With Melanoma and Colorectal Cancer Liver Metastases and the Association With Disease Outcome

PubMed Central

Kim, Joseph; Mori, Takuji; Chen, Steven L.; Amersi, Farin F.; Martinez, Steve R.; Kuo, Christine; Turner, Roderick R.; Ye, Xing; Bilchik, Anton J.; Morton, Donald L.; Hoon, Dave S. B.

2006-01-01

Objective: To determine the role of chemokine receptor (CR) expression in patients with melanoma and colorectal cancer (CRC) liver metastases. Summary Background Data: Murine and in vitro models have identified CR as potential factors in organ-specific metastasis of multiple cancers. Chemokines via their respective receptors have been shown to promote cell migration to distant organs. Methods: Patients who underwent hepatic surgery for melanoma or CRC liver metastases were assessed. Screening cDNA microarrays of melanoma/CRC cell lines and tumor specimens were analyzed to identify CR. Microarray data were validated by quantitative real-time RT-PCR (qRT) in paraffin-embedded liver metastases. Migration assays and immunohistochemistry were performed to verify CR function and confirm CR expression, respectively. Results: Microarray analysis identified CXCR4 as the most common CR expressed by both cancers. qRT demonstrated CXCR4 expression in 24 of 27 (89%) melanoma and 28 of 29 (97%) CRC liver metastases. In vitro treatment of melanoma or CRC cells with CXCL12, the ligand for CXCR4, significantly increased cell migration (P < 0.001). Low versus high CXCR4 expression in CRC liver metastases correlated with a significant difference in overall survival (median 27 months vs. 10 months, respectively; P = 0.036). In melanoma, low versus high CXCR4 expression in liver metastases demonstrated no difference in overall survival (median 11 months vs. 8 months, respectively; P = not significant). Conclusions: CXCR4 is expressed and functional on melanoma and CRC cells. The ligand for CXCR4 is highly expressed in liver and may specifically attract melanoma and CRC CXCR4 (+) cells. Quantitative analysis of CXCR4 gene expression in patients with liver metastases has prognostic significance for disease outcome. PMID:16794396

Epigenetic down regulation of G protein-coupled estrogen receptor (GPER) functions as a tumor suppressor in colorectal cancer.

PubMed

Liu, Qiao; Chen, Zhuojia; Jiang, Guanmin; Zhou, Yan; Yang, Xiangling; Huang, Hongbin; Liu, Huanliang; Du, Jun; Wang, Hongsheng

2017-05-05

Estrogenic signals are suggested to have protection roles in the development of colorectal cancer (CRC). The G protein-coupled estrogen receptor (GPER) has been reported to mediate non-genomic effects of estrogen in hormone related cancers except CRC. Its expression and functions in CRC were investigated. The expression of GPER and its associations with clinicopathological features were examined. The mechanisms were further investigated using cells, mouse xenograft models, and clinical human samples. GPER was significantly (p < 0.01) down regulated in CRC tissues compared with their matched adjacent normal tissues in our two cohorts and three independent investigations from Oncomine database. Patients whose tumors expressing less (n = 36) GPER showed significant (p < 0.01) poorer survival rate as compared with those with greater levels of GPER (n = 54). Promoter methylation and histone H3 deacetylation were involved in the down regulation of GPER in CRC cell lines and clinical tissues. Activation of GPER by its specific agonist G-1 inhibited proliferation, induced cell cycle arrest, mitochondrial-related apoptosis and endoplasmic reticulum (ER) stress of CRC cells. The upregulation of reactive oxygen species (ROS) induced sustained ERK1/2 activation participated in G-1 induced cell growth arrest. Further, G-1 can inhibit the phosphorylation, nuclear localization, and transcriptional activities of NF-κB via both canonical IKKα/ IκBα pathways and phosphorylation of GSK-3β. Xenograft model based on HCT-116 cells confirmed that G-1 can suppress the in vivo progression of CRC. Epigenetic down regulation of GPER acts as a tumor suppressor in colorectal cancer and its specific activation might be a potential approach for CRC treatment.

Cetuximab intensifies the ADCC activity of adoptive NK cells in a nude mouse colorectal cancer xenograft model.

PubMed

Chen, Shanshan; Li, Xuechun; Chen, Rongming; Yin, Mingang; Zheng, Qiuhong

2016-09-01

Natural killer (NK) cells, discovered ~40 years ago, are believed to be the most effective cytotoxic lymphocytes to counteract cancer; however, adoptive NK cell therapy in vivo has encountered certain limitations, including a lack of specificity. The drug cetuximab can mediate antibody dependent cell mediated cytotoxicity (ADCC) activity through NK cells in vivo , and has been approved for the first-line treatment of epidermal growth factor receptor (EGFR)-positive metastatic colorectal cancer (CRC). However, the ADCC activity of adoptive NK cells, induced by cetuximab in a nude mouse CRC xenograft model, has not been previously reported. The aim of the present study was to explore the ADCC activity of cetuximab combined with adoptive NK cells in CRC xenograft models with various EGFR expressions. The nude mouse xenograft models were established by subcutaneously injecting LOVO or SW620 cells. The mice were then randomly divided into 6 groups: Phosphate-buffered saline, cetuximab, human immunoglobulin G (hIgG), NK cells, hIgG plus NK cells and cetuximab plus NK cells. The ADCC antitumor activity was evaluated in these CRC models. The results indicated that the cetuximab plus NK cells group showed the greatest tumor inhibition effect compared with the NK cells group in LOVO xenograft tumor models with positive EGFR expression. However, the combination of cetuximab and NK cells did not show a stronger tumor inhibitory effect against the SW620 xenograft tumor models compared with the efficiency of NK cells. In conclusion, cetuximab could intensify the ADCC antitumor activity of adoptive NK cells towards CRC with an increased EGFR expression. The combination of cetuximab and NK cells may be a potential immunotherapy for metastatic CRC patients with positive EGFR expression.

Colorectal cancer cells display chaperone dependency for the unconventional prefoldin URI1

PubMed Central

Christinat, Yann; Frischknecht, Lukas; Krek, Wilhelm

2016-01-01

Chaperone dependency of cancer cells is an emerging trait that relates to the need of transformed cells to cope with the various stresses associated with the malignant state. URI1 (unconventional prefoldin RPB5 interactor 1) encodes a member of the prefoldin (PFD) family of molecular chaperones that acts as part of a heterohexameric PFD complex, the URI1 complex (URI1C), to promote assembly of multiprotein complexes involved in cell signaling and transcription processes. Here, we report that human colorectal cancer (CRCs) cell lines demonstrate differential dependency on URI1 and on the URI1 partner PFD STAP1 for survival, suggesting that this differential vulnerability of CRC cells is directly linked to URI1C chaperone function. Interestingly, in URI1-dependent CRC cells, URI1 deficiency is associated with non-genotoxic p53 activation and p53-dependent apoptosis. URI1-independent CRC cells do not exhibit such effects even in the context of wildtype p53. Lastly, in tumor xenografts, the conditional depletion of URI1 in URI1-dependent CRC cells was, after tumor establishment, associated with severe inhibition of subsequent tumor growth and activation of p53 target genes. Thus, a subset of CRC cells has acquired a dependency on the URI1 chaperone system for survival, providing an example of ‘non-oncogene addiction’ and vulnerability for therapeutic targeting. PMID:27105489

Rational drug design for anti-cancer chemotherapy: multi-target QSAR models for the in silico discovery of anti-colorectal cancer agents.

PubMed

Speck-Planche, Alejandro; Kleandrova, Valeria V; Luan, Feng; Cordeiro, M Natália D S

2012-08-01

The discovery of new and more potent anti-cancer agents constitutes one of the most active fields of research in chemotherapy. Colorectal cancer (CRC) is one of the most studied cancers because of its high prevalence and number of deaths. In the current pharmaceutical design of more efficient anti-CRC drugs, the use of methodologies based on Chemoinformatics has played a decisive role, including Quantitative-Structure-Activity Relationship (QSAR) techniques. However, until now, there is no methodology able to predict anti-CRC activity of compounds against more than one CRC cell line, which should constitute the principal goal. In an attempt to overcome this problem we develop here the first multi-target (mt) approach for the virtual screening and rational in silico discovery of anti-CRC agents against ten cell lines. Here, two mt-QSAR classification models were constructed using a large and heterogeneous database of compounds. The first model was based on linear discriminant analysis (mt-QSAR-LDA) employing fragment-based descriptors while the second model was obtained using artificial neural networks (mt-QSAR-ANN) with global 2D descriptors. Both models correctly classified more than 90% of active and inactive compounds in training and prediction sets. Some fragments were extracted from the molecules and their contributions to anti-CRC activity were calculated using mt-QSAR-LDA model. Several fragments were identified as potential substructural features responsible for the anti-CRC activity and new molecules designed from those fragments with positive contributions were suggested and correctly predicted by the two models as possible potent and versatile anti-CRC agents. Copyright © 2012 Elsevier Ltd. All rights reserved.

Peptostreptococcus anaerobius Induces Intracellular Cholesterol Biosynthesis in Colon Cells to Induce Proliferation and Causes Dysplasia in Mice.

PubMed

Tsoi, Ho; Chu, Eagle S H; Zhang, Xiang; Sheng, Jianqiu; Nakatsu, Geicho; Ng, Siew C; Chan, Anthony W H; Chan, Francis K L; Sung, Joseph J Y; Yu, Jun

2017-05-01

Stool samples from patients with colorectal cancer (CRC) have a higher abundance of Peptostreptococcus anaerobius than stool from individuals without CRC, based on metagenome sequencing. We investigated whether P anaerobius contributes to colon tumor formation in mice and its possible mechanisms of carcinogenesis. We performed quantitative polymerase chain reaction analyses to measure P anaerobius in 112 stool samples and 255 colon biopsies from patients with CRC or advanced adenoma and from healthy individuals (controls) undergoing colonoscopy examination at hospitals in Hong Kong and Beijing. C57BL/6 mice were given broad-spectrum antibiotics, followed by a single dose of azoxymethane, to induce colon tumor formation. Three days later, mice were given P anaerobius or Esherichia coli MG1655 (control bacteria), via gavage, for 6 weeks. Some mice were also given the nicotinamide adenine dinucleotide phosphate oxidase inhibitor apocynin. Intestine tissues were collected and analyzed histologically. The colon epithelial cell line NCM460 and colon cancer cell lines HT-29 and Caco-2 were exposed to P anaerobius or control bacteria; cells were analyzed by immunoblot, proliferation, and bacterial attachment analyses and compared in gene expression profiling studies. Gene expression was knocked down in these cell lines with small interfering RNAs. P anaerobius was significantly enriched in stool samples from patients with CRC and in biopsies from patients with colorectal adenoma or CRC compared with controls. Mice depleted of bacteria and exposed to azoxymethane and P anaerobius had a higher incidence of intestinal dysplasia (63%) compared with mice not given the bacteria (8.3%; P < .01). P anaerobius mainly colonized the colon compared with the rest of the intestine. Colon cells exposed to P anaerobius had significantly higher levels of proliferation than control cells. We found genes that regulate cholesterol biosynthesis, Toll-like receptor (TLR) signaling, and AMP-activated protein kinase signaling to be significantly up-regulated in cells exposed to P anaerobius. Total cholesterol levels were significantly increased in colon cell lines exposed to P anaerobius via activation of sterol regulatory element-binding protein 2. P anaerobius interacted with TLR2 and TLR4 to increase intracellular levels of reactive oxidative species, which promoted cholesterol synthesis and cell proliferation. Depletion of reactive oxidative species by knockdown of TLR2 or TLR4, or incubation of cells with an antioxidant, prevented P anaerobius from inducing cholesterol biosynthesis and proliferation. Levels of P anaerobius are increased in human colon tumor tissues and adenomas compared with non-tumor tissues; this bacteria increases colon dysplasia in a mouse model of CRC. P anaerobius interacts with TLR2 and TLR4 on colon cells to increase levels of reactive oxidative species, which promotes cholesterol synthesis and cell proliferation. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Crataegus azarolus Leaves Induce Antiproliferative Activity, Cell Cycle Arrest, and Apoptosis in Human HT-29 and HCT-116 Colorectal Cancer Cells.

PubMed

Mustapha, Nadia; Pinon, Aline; Limami, Youness; Simon, Alain; Ghedira, Kamel; Hennebelle, Thierry; Chekir-Ghedira, Leila

2016-05-01

Limited success has been achieved in extending the survival of patients with metastatic colorectal cancer (CRC). There is a strong need for novel agents in the treatment and prevention of CRC. Therefore, in the present study we evaluated the antiproliferative and pro-apoptotic potential of Crataegus azarolus ethyl acetate extract in HCT-116 and HT-29 human colorectal cancer cell lines. Moreover, we attempted to investigate the signaling pathways that should be involved in its cytotoxic effect. The Crataegus azarolus ethyl acetate extract-induced growth inhibitory effect was associated with DNA fragmentation, sub-G1 peak, loss of mitochondrial potential, and poly (ADP-ribose) polymerase (PARP) cleavage. In addition, ethyl acetate extract of Crataegus azarolus induced the cleavage of caspase-8. It has no effect on steady-state levels of total Bcl-2 protein. Whereas Bax levels decreased significantly in a dose-dependent manner in both tested cell lines. Taken together, these findings confirm the involvement of the extrinsic pathway of apoptosis. The apoptotic cell death induced by ethyl acetate extract of Crataegus azarolus was accompanied by an enhancement of the p21 expression but not through p53 activation in human colorectal cancer cells. The above-mentioned data provide insight into the molecular mechanisms of Crataegus azarolus ethyl acetate extract-induced apoptosis in CRC. Therefore, this compound should be a potential anticancer agent for the treatment of CRC. © 2015 Wiley Periodicals, Inc.

Long-term Culture and Cloning of Primary Human Bronchial Basal Cells that Maintain Multipotent Differentiation Capacity and CFTR Channel Function.

PubMed

Peters-Hall, Jennifer Ruth; Coquelin, Melissa L; Torres, Michael J; LaRanger, Ryan; Alabi, Busola Ruth; Sho, Sei; Calva-Moreno, Jose Francisco; Thomas, Philip J; Shay, Jerry William

2018-05-03

While primary cystic fibrosis (CF) and non-CF human bronchial epithelial basal cells (HBECs) accurately represent in vivo phenotypes, one barrier to their wider use has been a limited ability to clone and expand cells in sufficient numbers to produce rare genotypes using genome editing tools. Recently, conditional reprogramming of cells (CRC) with a ROCK inhibitor and culture on an irradiated fibroblast feeder layer resulted in extension of the lifespan of HBECs, but differentiation capacity and CF transmembrane conductance regulator (CFTR) function decreased as a function of passage. This report details modifications to the standard HBEC CRC protocol (Mod CRC), including the use of bronchial epithelial growth medium instead of F-medium and 2% oxygen instead of 21% oxygen, that extend HBEC lifespan while preserving multipotent differentiation capacity and CFTR function. Critically, Mod CRC conditions support clonal growth of primary HBECs from a single cell and the resulting clonal HBEC population maintains multipotent differentiation capacity, including CFTR function, permitting gene editing of these cells. As a proof of concept, CRISPR/Cas9 genome editing and cloning was used to introduce insertions/deletions in CFTR exon 11. Mod CRC conditions overcome many barriers to the expanded use of HBECs for basic research and drug screens. Importantly, Mod CRC conditions support the creation of isogenic cell lines in which CFTR is mutant or wild-type in the same genetic background with no history of CF to enable determination of the primary defects of mutant CFTR.

Association between hMLH1 hypermethylation and JC virus (JCV) infection in human colorectal cancer (CRC).

PubMed

Vilkin, Alex; Niv, Yaron

2011-04-01

Incorporation of viral DNA may interfere with the normal sequence of human DNA bases on the genetic level or cause secondary epigenetic changes such as gene promoter methylation or histone acetylation. Colorectal cancer (CRC) is the second leading cause of cancer mortality in the USA. Chromosomal instability (CIN) was established as the key mechanism in cancer development. Later, it was found that CRC results not only from the progressive accumulation of genetic alterations but also from epigenetic changes. JC virus (JCV) is a candidate etiologic factor in sporadic CRC. It may act by stabilizing β-catenin, facilitating its entrance to the cell nucleus, initialing proliferation and cancer development. Diploid CRC cell lines transfected with JCV-containing plasmids developed CIN. This result provides direct experimental evidence for the ability of JCV T-Ag to induce CIN in the genome of colonic epithelial cells. The association of CRC hMLH1 methylation and tumor positivity for JCV was recently documented. JC virus T-Ag DNA sequences were found in 77% of CRCs and are associated with promoter methylation of multiple genes. hMLH1 was methylated in 25 out of 80 CRC patients positive for T-Ag (31%) in comparison with only one out of 11 T-Ag negative cases (9%). Thus, JCV can mediate both CIN and aberrant methylation in CRC. Like other viruses, chronic infection with JCV may induce CRC by different mechanisms which should be further investigated. Thus, gene promoter methylation induced by JCV may be an important process in CRC and the polyp-carcinoma sequence.

The novel long intergenic noncoding RNA UCC promotes colorectal cancer progression by sponging miR-143

PubMed Central

Huang, Feng-Ting; Chen, Wen-Ying; Gu, Zhi-Qiang; Zhuang, Yan-Yan; Li, Chu-Qiang; Wang, Ling-Yun; Peng, Juan-Fei; Zhu, Zhe; Luo, Xin; Li, Yuan-Hua; Yao, He-Rui; Zhang, Shi-Neng

2017-01-01

The human genome contains thousands of long intergenic noncoding RNAs (lincRNAs). However, the functional roles of these transcripts and the mechanisms responsible for their deregulation in colorectal cancer (CRC) remain elusive. A novel lincRNA termed upregulated in CRC (UCC) was found to be highly expressed in human CRC tissues and cell lines. UCC levels correlated with lymph node metastasis, Dukes’ stage, and patient outcomes. In SW480 and SW620 cells, knockdown of UCC inhibited proliferation, invasion, and cell cycle progression and induced apoptosis in vitro. Xenograft tumors grown from UCC-silenced SW620 cells had smaller mean volumes and formed more slowly than xenograft tumors grown from control cells. Inversely, overexpression of UCC in HCT116 promoted cell growth and invasion in vitro. Bioinformatics analysis, dual-luciferase reporter assays, and RNA immunoprecipitation assays showed that miR-143 can interact with UCC, and we found that UCC expression inversely correlates with miR-143 expression in CRC specimens. Moreover, mechanistic investigations showed that UCC may act as an endogenous sponge by competing for miR-143, thereby regulating the targets of this miRNA. Our results suggest that UCC and miR-143 may be promising molecular targets for CRC therapy. PMID:28492554

The novel long intergenic noncoding RNA UCC promotes colorectal cancer progression by sponging miR-143.

PubMed

Huang, Feng-Ting; Chen, Wen-Ying; Gu, Zhi-Qiang; Zhuang, Yan-Yan; Li, Chu-Qiang; Wang, Ling-Yun; Peng, Juan-Fei; Zhu, Zhe; Luo, Xin; Li, Yuan-Hua; Yao, He-Rui; Zhang, Shi-Neng

2017-05-11

The human genome contains thousands of long intergenic noncoding RNAs (lincRNAs). However, the functional roles of these transcripts and the mechanisms responsible for their deregulation in colorectal cancer (CRC) remain elusive. A novel lincRNA termed upregulated in CRC (UCC) was found to be highly expressed in human CRC tissues and cell lines. UCC levels correlated with lymph node metastasis, Dukes' stage, and patient outcomes. In SW480 and SW620 cells, knockdown of UCC inhibited proliferation, invasion, and cell cycle progression and induced apoptosis in vitro. Xenograft tumors grown from UCC-silenced SW620 cells had smaller mean volumes and formed more slowly than xenograft tumors grown from control cells. Inversely, overexpression of UCC in HCT116 promoted cell growth and invasion in vitro. Bioinformatics analysis, dual-luciferase reporter assays, and RNA immunoprecipitation assays showed that miR-143 can interact with UCC, and we found that UCC expression inversely correlates with miR-143 expression in CRC specimens. Moreover, mechanistic investigations showed that UCC may act as an endogenous sponge by competing for miR-143, thereby regulating the targets of this miRNA. Our results suggest that UCC and miR-143 may be promising molecular targets for CRC therapy.

GPER mediates differential effects of estrogen on colon cancer cell proliferation and migration under normoxic and hypoxic conditions

PubMed Central

Bustos, Viviana; Nolan, Áine M.; Nijhuis, Anke; Harvey, Harry; Parker, Alexandra; Poulsom, Richard; McBryan, Jean; Thomas, Warren; Silver, Andrew; Harvey, Brian J.

2017-01-01

The estrogen receptor ERβ is the predominant ER subtype expressed in normal well-differentiated colonic epithelium. However, ERβ expression is lost under the hypoxic microenvironment as colorectal cancer (CRC) malignancy progresses. This raises questions about the role of signalling through other estrogen receptors such as ERα or G-protein coupled estrogen receptor (GPER, GPR30) by the estrogen 17β-estradiol (E2) under hypoxic conditions after ERβ is lost in CRC progression. We tested the hypothesis that E2 or hypoxia can act via GPER to contribute to the altered phenotype of CRC cells. GPER expression was found to be up-regulated by hypoxia and E2 in a panel of CRC cell lines. The E2-modulated gene, Ataxia telangiectasia mutated (ATM), was repressed in hypoxia via GPER signalling. E2 treatment enhanced hypoxia-induced expression of HIF1-α and VEGFA, but repressed HIF1-α and VEGFA expression under normoxic conditions. The expression and repression of VEGFA by E2 were mediated by a GPER-dependent mechanism. E2 treatment potentiated hypoxia-induced CRC cell migration and proliferation, whereas in normoxia, cell migration and proliferation were suppressed by E2 treatment. The effects of E2 on these cellular responses in normoxia and hypoxia were mediated by GPER. In a cohort of 566 CRC patient tumor samples, GPER expression significantly associated with poor survival in CRC Stages 3-4 females but not in the stage-matched male population. Our findings support a potentially pro-tumorigenic role for E2 in ERβ-negative CRC under hypoxic conditions transduced via GPER and suggest a novel route of therapeutic intervention through GPER antagonism. PMID:29137421

GPER mediates differential effects of estrogen on colon cancer cell proliferation and migration under normoxic and hypoxic conditions.

PubMed

Bustos, Viviana; Nolan, Áine M; Nijhuis, Anke; Harvey, Harry; Parker, Alexandra; Poulsom, Richard; McBryan, Jean; Thomas, Warren; Silver, Andrew; Harvey, Brian J

2017-10-13

The estrogen receptor ERβ is the predominant ER subtype expressed in normal well-differentiated colonic epithelium. However, ERβ expression is lost under the hypoxic microenvironment as colorectal cancer (CRC) malignancy progresses. This raises questions about the role of signalling through other estrogen receptors such as ERα or G-protein coupled estrogen receptor (GPER, GPR30) by the estrogen 17β-estradiol (E2) under hypoxic conditions after ERβ is lost in CRC progression. We tested the hypothesis that E2 or hypoxia can act via GPER to contribute to the altered phenotype of CRC cells. GPER expression was found to be up-regulated by hypoxia and E2 in a panel of CRC cell lines. The E2-modulated gene, Ataxia telangiectasia mutated ( ATM ), was repressed in hypoxia via GPER signalling. E2 treatment enhanced hypoxia-induced expression of HIF1-α and VEGFA, but repressed HIF1-α and VEGFA expression under normoxic conditions. The expression and repression of VEGFA by E2 were mediated by a GPER-dependent mechanism. E2 treatment potentiated hypoxia-induced CRC cell migration and proliferation, whereas in normoxia, cell migration and proliferation were suppressed by E2 treatment. The effects of E2 on these cellular responses in normoxia and hypoxia were mediated by GPER. In a cohort of 566 CRC patient tumor samples, GPER expression significantly associated with poor survival in CRC Stages 3-4 females but not in the stage-matched male population. Our findings support a potentially pro-tumorigenic role for E2 in ERβ-negative CRC under hypoxic conditions transduced via GPER and suggest a novel route of therapeutic intervention through GPER antagonism.

E2F1 promote the aggressiveness of human colorectal cancer by activating the ribonucleotide reductase small subunit M2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fang, Zejun; Gong, Chaoju; Liu, Hong

2015-08-21

As the ribonucleotide reductase small subunit, the high expression of ribonucleotide reductase small subunit M2 (RRM2) induces cancer and contributes to tumor growth and invasion. In several colorectal cancer (CRC) cell lines, we found that the expression levels of RRM2 were closely related to the transcription factor E2F1. Mechanistic studies were conducted to determine the molecular basis. Ectopic overexpression of E2F1 promoted RRM2 transactivation while knockdown of E2F1 reduced the levels of RRM2 mRNA and protein. To further investigate the roles of RRM2 which was activated by E2F1 in CRC, CCK-8 assay and EdU incorporation assay were performed. Overexpression ofmore » E2F1 promoted cell proliferation in CRC cells, which was blocked by RRM2 knockdown attenuation. In the migration and invasion tests, overexpression of E2F1 enhanced the migration and invasion of CRC cells which was abrogated by silencing RRM2. Besides, overexpression of RRM2 reversed the effects of E2F1 knockdown partially in CRC cells. Examination of clinical CRC specimens demonstrated that both RRM2 and E2F1 were elevated in most cancer tissues compared to the paired normal tissues. Further analysis showed that the protein expression levels of E2F1 and RRM2 were parallel with each other and positively correlated with lymph node metastasis (LNM), TNM stage and distant metastasis. Consistently, the patients with low E2F1 and RRM2 levels have a better prognosis than those with high levels. Therefore, we suggest that E2F1 can promote CRC proliferation, migration, invasion and metastasis by regulating RRM2 transactivation. Understanding the role of E2F1 in activating RRM2 transcription will help to explain the relationship between E2F1 and RRM2 in CRC and provide a novel predictive marker for diagnosis and prognosis of the disease. - Highlights: • E2F1 promotes RRM2 transactivation in CRC cells. • E2F1 promotes the proliferation of CRC cells by activating RRM2. • E2F1 promotes the migration and invasion of CRC cells by activating RRM2. • E2F1 is upregulated in CRC tissues and positively associated with RRM2 level. • E2F1-mediated RRM2 transcription will provide a new strategy in CRC.« less

Heat-shock protein 27 (HSP27, HSPB1) is synthetic lethal to cells with oncogenic activation of MET, EGFR and BRAF.

PubMed

Konda, John D; Olivero, Martina; Musiani, Daniele; Lamba, Simona; Di Renzo, Maria F

2017-06-01

The small heat-shock protein of 27 kDa (HSP27) is highly expressed in many cancers and is associated with aggressive tumour behaviour, metastasis, poor prognosis and resistance to chemotherapy. We aimed at assessing the role of HSP27 in modulating responses to target therapies. We selected several oncogene-addicted cancer cell lines, which undergo either cell cycle blockade or cell death in response to agents that target the specific oncogene. Surprisingly, HSP27 suppression alone resulted in the apoptotic death of MET-addicted EBC-1 lung cancer cells, epidermal growth factor receptor (EGFR)-addicted colorectal carcinoma (CRC) DiFi cells and BRAF-addicted CRC COLO205 and OXCO-1 and melanoma COLO741 cells, all of which also undergo death when treated with the specific targeted agent. In other cell lines, such as MET-addicted gastric carcinoma MKN45 and EGFR-addicted CRC SW48 lines, where oncogene inhibition only blocked proliferation, HSP27 knockdown made targeted agents switch from cytostatic to cytotoxic activity. Mechanistically, the more the cells were susceptible to HSP27 suppression, the more they were primed for death, as demonstrated by increased levels of mitochondrial outer membrane permeabilization. Priming for death was accompanied by the increase in pro-apoptotic proteins of the BCL2 family and of active caspase-3 and lamin B. Together, these data suggest that oncogene-addicted cells require HSP27 for survival and that HSP27 might interfere with the effectiveness of targeted agents. © 2017 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

Identification of G-protein-coupled receptor 120 as a tumor-promoting receptor that induces angiogenesis and migration in human colorectal carcinoma.

PubMed

Wu, Q; Wang, H; Zhao, X; Shi, Y; Jin, M; Wan, B; Xu, H; Cheng, Y; Ge, H; Zhang, Y

2013-12-05

G-protein-coupled receptor 120 (GPR120) functions as a receptor for unsaturated long-chain free fatty acids and has an important role in regulating lipid and glucose metabolism. However, a role for GPR120 in the development of tumors has not yet been clarified. Here, we show that GPR120 signaling promotes angiogenic switching and motility of human colorectal carcinoma (CRC) cells. We show that the expression of GPR120 is significantly induced in CRC tissues and cell lines, which is associated with tumor progression. Activation of GPR120 signaling in human CRC promotes angiogenesis in vitro and in vivo, largely by inducing the expression and secretion of proangiogenic mediators such as vascular endothelial growth factor (VEGF), interleukin-8 and cyclooxygenase-2-derived prostaglandin E2. The PI3K/Akt-NF-κB pathway is activated by GPR120 signaling and is required for GPR120 signaling-induced angiogenic switching in CRC cells. And, GPR120 activation enhances the motility of CRC cells and induces epithelial-mesenchymal transition. Furthermore, in vivo study shows that activation of GPR120 promotes angiogenesis and tumor growth. Finally, we find that GPR120 expression is positively correlated with VEGF expression and inversely correlated with the epithelial marker E-cadherin in CRC tissues. Collectively, our results demonstrate that GPR120 functions as a tumor-promoting receptor in CRC and, therefore, shows promise as a new potential target for cancer therapeutics.

Apelin13/APJ promotes proliferation of colon carcinoma by activating Notch3 signaling pathway.

PubMed

Chen, Tong; Liu, Ning; Xu, Guang-Meng; Liu, Tong-Jun; Liu, Ying; Zhou, Yan; Huo, Si-Bo; Zhang, Kai

2017-11-24

The link between Apelin (APL)/APL receptor (APJ) and Jagged (JAG)/Notch signaling pathways in colorectal cancer (CRC) has been poorly investigated. APL/APJ system, a potent angiogenic factor, is up-regulated in a variety of cancers. It contributes to tumor angiogenesis, and correlates with progression of malignancy. JAG/Notch signaling also contributes to progression, proliferation and metastasis of multiple cancers, including CRC. Here we tested the hypothesis that APL/APJ system promotes CRC proliferation by up-regulating Notch3, thus allowing further binding of JAG1 to Notch3. We used a variety of methods including Western blot, RT-qPCR, gene silencing, ELISA, immunofluorescence staining, to investigate the interaction between APL/APJ system and Notch3 signaling pathway in both surgically-resected specimens and CRC cell line LS180. We show that the expression of APL13, APJ, and Notch3 is elevated in CRC. We further demonstrate that APL13 can be secreted into culture media of LS180 cells, suggesting the existence of autocrine loop in CRC. Moreover, we found that APL13 stimulated expression of Notch3. Finally, we found that inhibition of either APJ or Notch3 prevents proliferation of LS180 cells. Our results suggest that APL13/APJ and JAG1/Notch3 signaling pathways are linked in CRC. These findings provide a new direction to the efforts targeting effective therapeutic and management approaches in the treatment of CRC.

Celecoxib enhances the radiosensitivity of HCT116 cells in a COX-2 independent manner by up-regulating BCCIP

PubMed Central

Xu, Xiao-Ting; Hu, Wen-Tao; Zhou, Ju-Ying; Tu, Yu

2017-01-01

It has been reported that celecoxib, a cyclooxygenase-2 (COX-2)-selective nonsteroidal anti-inflammatory drug (NSAID), regulates the radiosensitivity of several cancer cells. BCCIP (BRCA2 and CDKN1A interacting protein) plays a critical role in maintaining the critical functions of p53 in tumor suppression and response to therapy. However, whether the effect of celecoxib on the radiosensitivity of colorectal cancer (CRC) cells is dependent on BCCIP is largely unclear. In this study, we found that celecoxib enhanced the radiosensitivity of HeLa (a human cervical carcinoma cell line), A549 (a human lung carcinoma cell line), and HCT116 cells (a human CRC cells line). Among these cells, COX-2 expression was undetected in HCT116 cells. Treatment with celecoxib significantly increased BCCIP expression in COX-2 negative HCT116 cells. Knockdown of BCCIP obviously abrogated the enhanced radiosensitivity of HCT116 cells induced by celecoxib. A combination of celecoxib and irradiation treatment induced much more γ-H2AX foci formation, higher levels of radiation injury-related proteins phosphorylation, G2/M arrest, apoptosis, and p53 and p21 expression, and lower levels of Cyclin B1 in HCT116 cells than those in cells treated with irradiation alone. However, these changes were undetected in BCCIP-silenced HCT116 cells. Therefore, these data suggest that BCCIP gene may be a radiosensitivity-related gene in CRC. Celecoxib affects the functions of p53 and inhibits the recovery from the irradiation-induced injury by up-regulating the expression of BCCIP, and subsequently regulates the expressions of genes such as p21 and Cyclin B1 to enhance the radiosensitivity of HCT116 cells in a COX-2 independent manner. PMID:28386336

Regulatory network analysis of LINC00472, a long noncoding RNA downregulated by DNA hypermethylation in colorectal cancer.

PubMed

Chen, L; Zhang, W; Li, D Y; Wang, X; Tao, Y; Zhang, Y; Dong, C; Zhao, J; Zhang, L; Zhang, X; Guo, J; Zhang, X; Liao, Q

2018-06-01

Colorectal cancer (CRC), one of the common malignant cancers in the world, is caused by accumulated alterations of genetic and epigenetic factors over a long period of time. Along with that protein-coding genes being identified as oncogenes or tumor suppressors in CRC, a number of lncRNAs have also been found to be associated with CRC. Considering the important regulatory role of lncRNAs, the first goal of this study was to identify CRC-associated lncRNAs from a public database. One such lncRNA, LINC00472, was verified to be downregulated in CRC cell lines and cancer tissues compared with adjacent tissues. In addition, the down-regulation of LINC00472 seemed to be caused by DNA hypermethylation at its promoter region. Furthermore, the expression of LINC00472 and DNA methylation of promoter were significantly correlated with clinicopathological features. And DNA hypermethylation of LINC00472 may serve as a better diagnostic biomarker than its expression for CRC. Finally, we predicted the functions of LINC00472 and constructed a regulatory network and found LINC00472 may be involved in cell cycle and cell proliferation processes. Our results may provide a clue to further research into the function and regulatory mechanism of LINC00472 in CRC. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Combination of tolfenamic acid and curcumin induces colon cancer cell growth inhibition through modulating specific transcription factors and reactive oxygen species.

PubMed

Sankpal, Umesh T; Nagaraju, Ganji Purnachandra; Gottipolu, Sriharika R; Hurtado, Myrna; Jordan, Christopher G; Simecka, Jerry W; Shoji, Mamoru; El-Rayes, Bassel; Basha, Riyaz

2016-01-19

Curcumin (Cur) has been extensively studied in several types of malignancies including colorectal cancer (CRC); however its clinical application is greatly affected by low bioavailability. Several strategies to improve the therapeutic response of Cur are being pursued, including its combination with small molecules and drugs. We investigated the therapeutic efficacy of Cur in combination with the small molecule tolfenamic acid (TA) in CRC cell lines. TA has been shown to inhibit the growth of human cancer cells in vitro and in vivo, via targeting the transcription factor specificity protein1 (Sp1) and suppressing survivin expression. CRC cell lines HCT116 and HT29 were treated with TA and/or Cur and cell viability was measured 24-72 hours post-treatment. While both agents caused a steady reduction in cell viability, following a clear dose/ time-dependent response, the combination of TA+Cur showed higher growth inhibition when compared to either single agent. Effects on apoptosis were determined using flow cytometry (JC-1 staining to measure mitochondrial membrane potential), Western blot analysis (c-PARP expression) and caspase 3/7 activity. Reactive oxygen species (ROS) levels were measured by flow cytometry and the translocation of NF-kB into the nucleus was determined using immunofluorescence. Results showed that apoptotic markers and ROS activity were significantly upregulated following combination treatment, when compared to the individual agents. This was accompanied by decreased expression of Sp1, survivin and NF-kB translocation. The combination of TA+Cur was more effective in HCT116 cells than HT29 cells. These results demonstrate that TA may enhance the anti-proliferative efficacy of Cur in CRC cells.

MicroRNA-375 inhibits colorectal cancer growth by targeting PIK3CA

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yihui; Tang, Qingchao; Li, Mingqi

2014-02-07

Highlights: • miR-375 is downregulated in colorectal cancer cell lines and tissues. • miR-375 inhibits colorectal cancer cell growth by targeting PIK3CA. • miR-375 inhibits colorectal cancer cell growth in xenograft nude mice model. - Abstract: Colorectal cancer (CRC) is the second most common cause of death from cancer. MicroRNAs (miRNAs) represent a class of small non-coding RNAs that control gene expression by triggering RNA degradation or interfering with translation. Aberrant miRNA expression is involved in human disease including cancer. Herein, we showed that miR-375 was frequently down-regulated in human colorectal cancer cell lines and tissues when compared to normalmore » human colon tissues. PIK3CA was identified as a potential miR-375 target by bioinformatics. Overexpression of miR-375 in SW480 and HCT15 cells reduced PIK3CA protein expression. Subsequently, using reporter constructs, we showed that the PIK3CA untranslated region (3′-UTR) carries the directly binding site of miR-375. Additionally, miR-375 suppressed CRC cell proliferation and colony formation and led to cell cycle arrest. Furthermore, miR-375 overexpression resulted in inhibition of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. SiRNA-mediated silencing of PIK3CA blocked the inhibitory effect of miR-375 on CRC cell growth. Lastly, we found overexpressed miR-375 effectively repressed tumor growth in xenograft animal experiments. Taken together, we propose that overexpression of miR-375 may provide a selective growth inhibition for CRC cells by targeting PI3K/Akt signaling pathway.« less

MicroRNA-320 suppresses colorectal cancer by targeting SOX4, FOXM1, and FOXQ1

PubMed Central

Vishnubalaji, Radhakrishnan; Hamam, Rimi; Shijun, Yue; Al-Obeed, Omar; Kassem, Moustapha; Liu, Fei-Fei; Aldahmash, Abdullah; Alajez, Nehad M.

2016-01-01

Colorectal cancer (CRC) is the third most common cancer causing high mortality rates world-wide. Delineating the molecular mechanisms leading to CRC development and progression, including the role of microRNAs (miRNAs), are currently being unravelled at a rapid rate. Here, we report frequent downregulation of the microRNA miR-320 family in primary CRC tissues and cell lines. Lentiviral-mediated re-expression of miR-320c (representative member of the miR-320 family) inhibited HCT116 CRC growth and migration in vitro, sensitized CRC cells to 5-Fluorouracil (5-FU), and inhibited tumor formation in SCID mice. Global gene expression analysis in CRC cells over-expressing miR-320c, combined with in silico prediction identified 84 clinically-relevant potential gene targets for miR-320 in CRC. Using a series of biochemical assays and functional validation, SOX4, FOXM1, and FOXQ1 were validated as novel gene targets for the miR-320 family. Inverse correlation between the expression of miR-320 members with SOX4, FOXM1, and FOXQ1 was observed in primary CRC patients' specimens, suggesting that these genes are likely bona fide targets for the miR-320 family. Interestingly, interrogation of the expression levels of this gene panel (SOX4, FOXM1, and FOXQ1) in The Cancer Genome Atlas (TCGA) colorectal cancer data set (319 patients) revealed significantly poor disease-free survival in patients with elevated expression of this gene panel (P-Value: 0.0058). Collectively, our data revealed a novel role for the miR-320/SOX4/FOXM1/FOXQ1 axes in promoting CRC development and progression and suggest targeting those networks as potential therapeutic strategy for CRC. PMID:27119506

Tcf3 and cell cycle factors contribute to butyrate resistance in colorectal cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chiaro, Christopher, E-mail: cchiaro@tcmedc.org; Lazarova, Darina L., E-mail: dlazarova@tcmedc.org; Bordonaro, Michael, E-mail: mbordonaro@tcmedc.org

2012-11-09

Highlights: Black-Right-Pointing-Pointer We investigate mechanisms responsible for butyrate resistance in colon cancer cells. Black-Right-Pointing-Pointer Tcf3 modulates butyrate's effects on Wnt activity and cell growth in resistant cells. Black-Right-Pointing-Pointer Tcf3 modulation of butyrate's effects differ by cell context. Black-Right-Pointing-Pointer Cell cycle factors are overexpressed in the resistant cells. Black-Right-Pointing-Pointer Reversal of altered gene expression can enhance the anti-cancer effects of butyrate. -- Abstract: Butyrate, a fermentation product of dietary fiber, inhibits clonal growth in colorectal cancer (CRC) cells dependent upon the fold induction of Wnt activity. We have developed a CRC cell line (HCT-R) that, unlike its parental cell line, HCT-116,more » does not respond to butyrate exposure with hyperactivation of Wnt signaling and suppressed clonal growth. PCR array analyses revealed Wnt pathway-related genes, the expression of which differs between butyrate-sensitive HCT-116 CRC cells and their butyrate-resistant HCT-R cell counterparts. We identified overexpression of Tcf3 as being partially responsible for the butyrate-resistant phenotype, as this DNA-binding protein suppresses the hyperinduction of Wnt activity by butyrate. Consequently, Tcf3 knockdown in HCT-R cells restores their sensitivity to the effects of butyrate on Wnt activity and clonal cell growth. Interestingly, the effects of overexpressed Tcf3 differ between HCT-116 and HCT-R cells; thus, in HCT-116 cells Tcf3 suppresses proliferation without rendering the cells resistant to butyrate. In HCT-R cells, however, the overexpression of Tcf3 inhibits Wnt activity, and the cells are still able to proliferate due to the higher expression levels of cell cycle factors, particularly those driving the G{sub 1} to S transition. Knowledge of the molecular mechanisms determining the variable sensitivity of CRC cells to butyrate may assist in developing approaches that prevent or reverse butyrate resistance.« less

Loss of Smad4 in colorectal cancer induces resistance to 5-fluorouracil through activating Akt pathway.

PubMed

Zhang, B; Zhang, B; Chen, X; Bae, S; Singh, K; Washington, M K; Datta, P K

2014-02-18

Higher frequency of Smad4 inactivation or loss of expression is observed in metastasis of colorectal cancer (CRC) leading to unfavourable survival and contributes to chemoresistance. However, the molecular mechanism of how Smad4 regulates chemosensitivity of CRC is unknown. We evaluated how the loss of Smad4 in CRC enhanced chemoresistance to 5-fluorouracil (5-FU) using two CRC cell lines in vitro and in vivo. Immunoblotting with cell and tumour lysates and immunohistochemical analyses with tissue microarray were performed. Knockdown or loss of Smad4 induced tumorigenicity, migration, invasion, angiogenesis, metastasis, and 5-FU resistance. Smad4 expression in mouse tumours regulated cell-cycle regulatory proteins leading to Rb phosphorylation. Loss of Smad4 activated Akt pathway that resulted in upregulation of anti-apoptotic proteins, Bcl-2 and Bcl-w, and Survivin. Suppression of phosphatidylinositol-3-kinase (PI3K)/Akt pathway by LY294002 restored chemosensitivity of Smad4-deficient cells to 5-FU. Vascular endothelial growth factor-induced angiogenesis in Smad4-deficient cells might also lead to chemoresistance. Low levels of Smad4 expression in CRC tissues correlated with higher levels of Bcl-2 and Bcl-w and with poor overall survival as observed in immunohistochemical staining of tissue microarrays. Loss of Smad4 in CRC patients induces resistance to 5-FU-based therapy through activation of Akt pathway and inhibitors of this pathway may sensitise these patients to 5-FU.

Loss of Smad4 in colorectal cancer induces resistance to 5-fluorouracil through activating Akt pathway

PubMed Central

Zhang, B; Zhang, B; Chen, X; Bae, S; Singh, K; Washington, M K; Datta, P K

2014-01-01

Background: Higher frequency of Smad4 inactivation or loss of expression is observed in metastasis of colorectal cancer (CRC) leading to unfavourable survival and contributes to chemoresistance. However, the molecular mechanism of how Smad4 regulates chemosensitivity of CRC is unknown. Methods: We evaluated how the loss of Smad4 in CRC enhanced chemoresistance to 5-fluorouracil (5-FU) using two CRC cell lines in vitro and in vivo. Immunoblotting with cell and tumour lysates and immunohistochemical analyses with tissue microarray were performed. Results: Knockdown or loss of Smad4 induced tumorigenicity, migration, invasion, angiogenesis, metastasis, and 5-FU resistance. Smad4 expression in mouse tumours regulated cell-cycle regulatory proteins leading to Rb phosphorylation. Loss of Smad4 activated Akt pathway that resulted in upregulation of anti-apoptotic proteins, Bcl-2 and Bcl-w, and Survivin. Suppression of phosphatidylinositol-3-kinase (PI3K)/Akt pathway by LY294002 restored chemosensitivity of Smad4-deficient cells to 5-FU. Vascular endothelial growth factor-induced angiogenesis in Smad4-deficient cells might also lead to chemoresistance. Low levels of Smad4 expression in CRC tissues correlated with higher levels of Bcl-2 and Bcl-w and with poor overall survival as observed in immunohistochemical staining of tissue microarrays. Conclusion: Loss of Smad4 in CRC patients induces resistance to 5-FU-based therapy through activation of Akt pathway and inhibitors of this pathway may sensitise these patients to 5-FU. PMID:24384683

In vivo analysis of insulin-like growth factor type 1 receptor humanized monoclonal antibody MK-0646 and small molecule kinase inhibitor OSI-906 in colorectal cancer.

PubMed

Leiphrakpam, Premila D; Agarwal, Ekta; Mathiesen, Michelle; Haferbier, Katie L; Brattain, Michael G; Chowdhury, Sanjib

2014-01-01

The development and characterization of effective anticancer drugs against colorectal cancer (CRC) is of urgent need since it is the second most common cause of cancer death. The study was designed to evaluate the effects of two IGF-1R antagonists, MK-0646, a recombinant fully humanized monoclonal antibody and OSI-906, a small molecule tyrosine kinase inhibitor on CRC cells. Xenograft study was performed on IGF-1R-dependent CRC cell lines for analyzing the antitumor activity of MK-0646 and OSI-906. Tumor proliferation and apoptosis were assessed using Ki67 and TUNEL assays, respectively. We also performed in vitro characterization of MK-0646 and OSI-906 treatment on CRC cells to identify mechanisms associated with drug-induced cell death. Exposure of the GEO and CBS tumor xenografts to MK-0646 or OSI-906 led to a decrease in tumor growth. TUNEL analysis showed an increase of approximately 45-55% in apoptotic cells in both MK-0646 and OSI-906 treated tumor samples. We report the novel finding that treatment with IGF-1R antagonists led to downregulation of X-linked inhibitor of apoptosis (XIAP) protein involved in cell survival and inhibition of cell death. In conclusion, IGF-1R antagonists (MK-0646 and OSI-906) demonstrated single agent inhibition of subcutaneous CRC xenograft growth. This was coupled to pro-apoptotic effects resulting in downregulation of XIAP and inhibition of cell survival. We report a novel mechanism by which MK-0646 and OSI-906 elicits cell death in vivo and in vitro. Moreover, these results indicate that MK-0646 and OSI-906 may be potential anticancer candidates for the treatment of patients with IGF-1R-dependent CRC.

In vivo analysis of insulin-like growth factor type 1 receptor humanized monoclonal antibody MK-0646 and small molecule kinase inhibitor OSI-906 in colorectal cancer

PubMed Central

LEIPHRAKPAM, PREMILA D.; AGARWAL, EKTA; MATHIESEN, MICHELLE; HAFERBIER, KATIE L.; BRATTAIN, MICHAEL G.; CHOWDHURY, SANJIB

2014-01-01

The development and characterization of effective anticancer drugs against colorectal cancer (CRC) is of urgent need since it is the second most common cause of cancer death. The study was designed to evaluate the effects of two IGF-1R antagonists, MK-0646, a recombinant fully humanized monoclonal antibody and OSI-906, a small molecule tyrosine kinase inhibitor on CRC cells. Xenograft study was performed on IGF-1R-dependent CRC cell lines for analyzing the antitumor activity of MK-0646 and OSI-906. Tumor proliferation and apoptosis were assessed using Ki67 and TUNEL assays, respectively. We also performed in vitro characterization of MK-0646 and OSI-906 treatment on CRC cells to identify mechanisms associated with drug-induced cell death. Exposure of the GEO and CBS tumor xenografts to MK-0646 or OSI-906 led to a decrease in tumor growth. TUNEL analysis showed an increase of approximately 45–55% in apoptotic cells in both MK-0646 and OSI-906 treated tumor samples. We report the novel finding that treatment with IGF-1R antagonists led to downregulation of X-linked inhibitor of apoptosis (XIAP) protein involved in cell survival and inhibition of cell death. In conclusion, IGF-1R antagonists (MK-0646 and OSI-906) demonstrated single agent inhibition of subcutaneous CRC xenograft growth. This was coupled to pro-apoptotic effects resulting in downregulation of XIAP and inhibition of cell survival. We report a novel mechanism by which MK-0646 and OSI-906 elicits cell death in vivo and in vitro. Moreover, these results indicate that MK-0646 and OSI-906 may be potential anticancer candidates for the treatment of patients with IGF-1R-dependent CRC. PMID:24173770

Modulation of transcription factor binding and epigenetic regulation of the MLH1 CpG island and shore by polymorphism rs1800734 in colorectal cancer

PubMed Central

Savio, Andrea J.; Bapat, Bharati

2017-01-01

ABSTRACT The MLH1 promoter polymorphism rs1800734 is associated with MLH1 CpG island hypermethylation and expression loss in colorectal cancer (CRC). Conversely, variant rs1800734 is associated with MLH1 shore, but not island, hypomethylation in peripheral blood mononuclear cell DNA. To explore these distinct patterns, MLH1 CpG island and shore methylation was assessed in CRC cell lines stratified by rs1800734 genotype. Cell lines containing the variant A allele demonstrated MLH1 shore hypomethylation compared to wild type (GG). There was significant enrichment of transcription factor AP4 at the MLH1 promoter in GG and GA cell lines, but not the AA cell line, by chromatin immunoprecipitation studies. Preferential binding to the G allele was confirmed by sequencing in the GA cell line. The enhancer-associated histone modification H3K4me1 was enriched at the MLH1 shore; however, H3K27ac was not, indicating the shore is an inactive enhancer. These results demonstrate the role of variant rs1800734 in altering transcription factor binding as well as epigenetics at regions beyond the MLH1 CpG island in which it is located. PMID:28304185

Modulation of transcription factor binding and epigenetic regulation of the MLH1 CpG island and shore by polymorphism rs1800734 in colorectal cancer.

PubMed

Savio, Andrea J; Bapat, Bharati

2017-06-03

The MLH1 promoter polymorphism rs1800734 is associated with MLH1 CpG island hypermethylation and expression loss in colorectal cancer (CRC). Conversely, variant rs1800734 is associated with MLH1 shore, but not island, hypomethylation in peripheral blood mononuclear cell DNA. To explore these distinct patterns, MLH1 CpG island and shore methylation was assessed in CRC cell lines stratified by rs1800734 genotype. Cell lines containing the variant A allele demonstrated MLH1 shore hypomethylation compared to wild type (GG). There was significant enrichment of transcription factor AP4 at the MLH1 promoter in GG and GA cell lines, but not the AA cell line, by chromatin immunoprecipitation studies. Preferential binding to the G allele was confirmed by sequencing in the GA cell line. The enhancer-associated histone modification H3K4me1 was enriched at the MLH1 shore; however, H3K27ac was not, indicating the shore is an inactive enhancer. These results demonstrate the role of variant rs1800734 in altering transcription factor binding as well as epigenetics at regions beyond the MLH1 CpG island in which it is located.

PKM2 Subcellular Localization Is Involved in Oxaliplatin Resistance Acquisition in HT29 Human Colorectal Cancer Cell Lines

PubMed Central

Ginés, Alba; Bystrup, Sara; Ruiz de Porras, Vicenç; Guardia, Cristina; Musulén, Eva; Martínez-Cardús, Anna; Manzano, José Luis; Layos, Laura; Abad, Albert; Martínez-Balibrea, Eva

2015-01-01

Chemoresistance is the main cause of treatment failure in advanced colorectal cancer (CRC). However, molecular mechanisms underlying this phenomenon remain to be elucidated. In a previous work we identified low levels of PKM2 as a putative oxaliplatin-resistance marker in HT29 CRC cell lines and also in patients. In order to assess how PKM2 influences oxaliplatin response in CRC cells, we silenced PKM2 using specific siRNAs in HT29, SW480 and HCT116 cells. MTT test demonstrated that PKM2 silencing induced resistance in HT29 and SW480 cells and sensitivity in HCT116 cells. Same experiments in isogenic HCT116 p53 null cells and double silencing of p53 and PKM2 in HT29 cells failed to show an influence of p53. By using trypan blue stain and FITC-Annexin V/PI tests we detected that PKM2 knockdown was associated with an increase in cell viability but not with a decrease in apoptosis activation in HT29 cells. Fluorescence microscopy revealed PKM2 nuclear translocation in response to oxaliplatin in HCT116 and HT29 cells but not in OXA-resistant HTOXAR3 cells. Finally, by using a qPCR Array we demonstrated that oxaliplatin and PKM2 silencing altered cell death gene expression patterns including those of BMF, which was significantly increased in HT29 cells in response to oxaliplatin, in a dose and time-dependent manner, but not in siPKM2-HT29 and HTOXAR3 cells. BMF gene silencing in HT29 cells lead to a decrease in oxaliplatin-induced cell death. In conclusion, our data report new non-glycolytic roles of PKM2 in response to genotoxic damage and proposes BMF as a possible target gene of PKM2 to be involved in oxaliplatin response and resistance in CRC cells. PMID:25955657

Circular RNA circITGA7 inhibits colorectal cancer growth and metastasis by modulating the Ras pathway and upregulating transcription of its host gene ITGA7.

PubMed

Li, Xiaomin; Wang, Jianjun; Zhang, Chao; Lin, Chun; Zhang, Jianming; Zhang, Wei; Zhang, Wenjuan; Lu, Yanxia; Zheng, Lin; Li, Xuenong

2018-06-26

Circular RNAs (circRNAs) are significantly dysregulated in various cancer types. However, the roles and mechanisms of circRNAs in cancer remain largely unknown. In this study, we demonstrated that a novel circRNA (circITGA7) and its linear host gene ITGA7 are both significantly downregulated in colorectal cancer (CRC) tissues and cell lines. These decreased expression levels correlated with CRC progression. Functional assays demonstrated that ectopic circITGA7 expression suppressed the growth and metastasis of CRC cell in vitro and in vivo. Knockdown of circITGA7 or ITGA7 promoted the proliferation and migration of CRC cells in vitro and enhanced CRC growth in vivo. Mechanistically, we found that circITGA7 is a negative regulator of the Ras signalling pathway and ITGA7 is associated with cytokine-related signalling pathways through RNA-seq and KEGG enrichment analysis. In addition, circITGA7 binds to miR-370-3p to antagonize its suppression of NF1, which is a well-known negative regulator of the Ras pathway. Finally, circITGA7 upregulates the transcription of ITGA7 by suppressing RREB1 via the Ras pathway. In conclusion, our findings indicate a suppressor role of circITGA7 and ITGA7 in CRC and reveal that circITGA7 inhibits proliferation and metastasis of CRC cells by suppressing the Ras signalling pathway and promoting the transcription of ITGA7, suggesting that circITGA7 is a potential target for CRC treatment. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

EphA2 receptor activation with ephrin-A1 ligand restores cetuximab efficacy in NRAS-mutant colorectal cancer cells.

PubMed

Cuyàs, Elisabet; Queralt, Bernardo; Martin-Castillo, Begoña; Bosch-Barrera, Joaquim; Menendez, Javier A

2017-07-01

Patients with wild-type KRAS metastatic colorectal cancer (mCRC) that harbors NRAS activating mutations do not benefit from anti-EGFR therapies. Very little is known about oncogenic NRAS signaling driving mCRC unresponsiveness to the EGFR-directed antibody cetuximab. Using a system of paired NRAS-mutant and wild-type isogenic mCRC cell lines to explore signaling pathways engaged by the common oncogenic NRAS Q61K variant upon challenge with cetuximab, we uncovered an unexpected mechanism of resistance to cetuximab involving dysregulation of the ephrin-A1/EphA2 signaling axis. Parental NRAS+/+ cells, but not NRASQ61K/+ cells, activated the ephrin receptor ephA1 in response to cetuximab treatment. Moreover, whereas cetuximab treatment significantly downregulated EPHA2 gene expression in NRAS+/+ cells, EPHA2 expression in NRASQ61K/+ cells was refractory to cetuximab. Remarkably, pharmacologically mimicked ephrin-A1 engagement to ephA2 converted NRAS-mutant into RAS wild-type mCRC cells in terms of cetuximab efficacy. Accordingly, activation of the ephA2 receptor by bioactive recombinant human ephrin-A1/Fc-fusion protein suppressed the cetuximab-unresponsive hyperactivation of MAPK and AKT and fully restored cetuximab activity in NRAS-mutant colorectal cells. Collectively, these findings reveal that the clinical benefit of cetuximab in mCRC might necessarily involve the suppression of the ligandless oncogenic signaling of the ephA2 receptor. Hence, ligand-dependent tumor suppressor signaling using therapeutic ephA2 agonists might offer new therapeutic opportunities to clinically widen the use of cetuximab in NRAS-mutated and/or ephA2-dependent mCRC tumors.

Hepatocytes Determine the Hypoxic Microenvironment and Radiosensitivity of Colorectal Cancer Cells Through Production of Nitric Oxide That Targets Mitochondrial Respiration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiang, Heng; Verovski, Valeri N.; Leonard, Wim

2013-03-01

Purpose: To determine whether host hepatocytes may reverse hypoxic radioresistance through nitric oxide (NO)-induced oxygen sparing, in a model relevant to colorectal cancer (CRC) liver metastases. Methods and Materials: Hepatocytes and a panel of CRC cells were incubated in a tissue-mimetic coculture system with diffusion-limited oxygenation, and oxygen levels were monitored by an oxygen-sensing fluorescence probe. To activate endogenous NO production, cocultures were exposed to a cytokine mixture, and the expression of inducible nitric oxide synthase was analyzed by reverse transcription–polymerase chain reaction, Western blotting, and NO/nitrite production. The mitochondrial targets of NO were examined by enzymatic activity. To assessmore » hypoxic radioresponse, cocultures were irradiated and reseeded for colonies. Results: Resting hepatocytes consumed 10-40 times more oxygen than mouse CT26 and human DLD-1, HT29, HCT116, and SW480 CRC cells, and thus seemed to be the major effectors of hypoxic conditioning. As a result, hepatocytes caused uniform radioprotection of tumor cells at a 1:1 ratio. Conversely, NO-producing hepatocytes radiosensitized all CRC cell lines more than 1.5-fold, similar to the effect of selective mitochondrial inhibitors. The radiosensitizing effect was associated with a respiratory self-arrest of hepatocytes at the level of aconitase and complex II, which resulted in profound reoxygenation of tumor cells through oxygen sparing. Nitric oxide–producing hepatocytes were at least 10 times more active than NO-producing macrophages to reverse hypoxia-induced radioresistance. Conclusions: Hepatocytes were the major determinants of the hypoxic microenvironment and radioresponse of CRC cells in our model of metabolic hypoxia. We provide evidence that reoxygenation and radiosensitization of hypoxic CRC cells can be achieved through oxygen sparing induced by endogenous NO production in host hepatocytes.« less

Overexpression of phosphoserine aminotransferase PSAT1 stimulates cell growth and increases chemoresistance of colon cancer cells

PubMed Central

Vié, Nadia; Copois, Virginie; Bascoul-Mollevi, Caroline; Denis, Vincent; Bec, Nicole; Robert, Bruno; Fraslon, Caroline; Conseiller, Emmanuel; Molina, Franck; Larroque, Christian; Martineau, Pierre; Del Rio, Maguy; Gongora, Céline

2008-01-01

Background Colorectal cancer (CRC) is one of the most common causes of cancer death throughout the world. In this work our aim was to study the role of the phosphoserine aminotransferase PSAT1 in colorectal cancer development. Results We first observed that PSAT1 is overexpressed in colon tumors. In addition, we showed that after drug treatment, PSAT1 expression level in hepatic metastases increased in non responder and decreased in responder patients. In experiments using human cell lines, we showed that ectopic PSAT1 overexpression in colon carcinoma SW480 cell line resulted in an increase in its growth rate and survival. In addition, SW480-PSAT1 cells presented a higher tumorigenic potential than SW480 control cells in xenografted mice. Moreover, the SW480-PSAT1 cell line was more resistant to oxaliplatin treatment than the non-transfected SW480 cell line. This resistance resulted from a decrease in the apoptotic response and in the mitotic catastrophes induced by the drug treatment. Conclusion These results show that an enzyme playing a role in the L-serine biosynthesis could be implicated in colon cancer progression and chemoresistance and indicate that PSAT1 represents a new interesting target for CRC therapy. PMID:18221502

IL-33 promotes growth and liver metastasis of colorectal cancer in mice by remodeling the tumor microenvironment and inducing angiogenesis.

PubMed

Zhang, Yu; Davis, Celestia; Shah, Sapana; Hughes, Daniel; Ryan, James C; Altomare, Diego; Peña, Maria Marjorette O

2017-01-01

Liver metastasis is the major cause of death from colorectal cancer (CRC). Understanding its mechanisms is necessary for timely diagnosis and development of effective therapies. Interleukin-33 (IL-33) is an IL-1 cytokine family member that uniquely functions as a cytokine and nuclear factor. It is released by necrotic epithelial cells and activated innate immune cells, functioning as an alarmin or an early danger signal. Its role in invoking type 2 immune response has been established; however, it has contrasting roles in tumor development and metastasis. We identified IL-33 as a potently upregulated cytokine in a highly metastatic murine CRC cell line and examined its role in tumor growth and metastasis to the liver. IL-33 was transgenically expressed in murine and human adenocarcinoma and carcinoma cell lines and their growth and spontaneous metastasis to the liver were assessed in orthotopic models of CRC in wild-type C57Bl/6 and Il33 knockout mice. The results showed that increased expression of IL-33 in CRC cells enhanced their tumor take, growth, and liver metastasis. Tumor- rather than host-derived IL-33 induced the enhanced recruitment of CD11b + GR1 + and CD11b + F4/80 + myeloid cells to remodel the tumor microenvironment by increased expression of mobilizing cytokines, and tumor angiogenesis by activating endothelial cells. IL-33 expression was elevated in patient tumor tissues, induced early in adenoma development, and activated by pro-inflammatory cytokines derived from the tumor microenvironment. The data suggest that tumor-derived IL-33 modulates the tumor microenvironment to potently promote colon carcinogenesis and liver metastasis, underscoring its potential as a therapeutic target. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Loss of AXIN1 drives acquired resistance to WNT pathway blockade in colorectal cancer cells carrying RSPO3 fusions.

PubMed

Picco, Gabriele; Petti, Consalvo; Centonze, Alessia; Torchiaro, Erica; Crisafulli, Giovanni; Novara, Luca; Acquaviva, Andrea; Bardelli, Alberto; Medico, Enzo

2017-03-01

In colorectal cancer (CRC), WNT pathway activation by genetic rearrangements of RSPO3 is emerging as a promising target. However, its low prevalence severely limits availability of preclinical models for in-depth characterization. Using a pipeline designed to suppress stroma-derived signal, we find that RSPO3 "outlier" expression in CRC samples highlights translocation and fusion transcript expression. Outlier search in 151 CRC cell lines identified VACO6 and SNU1411 cells as carriers of, respectively, a canonical PTPRK(e1)-RSPO3(e2) fusion and a novel PTPRK(e13)-RSPO3(e2) fusion. Both lines displayed marked in vitro and in vivo sensitivity to WNT blockade by the porcupine inhibitor LGK974, associated with transcriptional and morphological evidence of WNT pathway suppression. Long-term treatment of VACO6 cells with LGK974 led to the emergence of a resistant population carrying two frameshift deletions of the WNT pathway inhibitor AXIN1, with consequent protein loss. Suppression of AXIN1 in parental VACO6 cells by RNA interference conferred marked resistance to LGK974. These results provide the first mechanism of secondary resistance to WNT pathway inhibition. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

BAG3 regulates cell proliferation, migration, and invasion in human colorectal cancer.

PubMed

Shi, Huiyong; Xu, Haidong; Li, Zengjun; Zhen, Yanan; Wang, Bin; Huo, Shoujun; Xiao, Ruixue; Xu, Zhongfa

2016-04-01

Bcl2-associated athanogene 3 (BAG3) has been reported to be elevated in various tumors. However, it is unclear whether BAG3 has a functional role in the initiation and progression of colorectal cancer (CRC). Here, we collected CRC samples and cell lines to validate the pathway by using gene and protein assays. RT-PCR showed that the expression of BAG3 mRNA in CRC tissues was obviously higher than that in non-tumor tissues (p < 0.001). Immunohistochemical analysis showed that immunoreactivity of BAG3 was found in most CRC tissues and strongly correlated with TNM stage (p = 0.001), differentiation (p = 0.003), and metastasis (p = 0.010). Low expression of BAG3 in HCT-8 significantly reduced cellular proliferation, migration, and invasion. The analysis of in vitro cell showed that HCT-8 cells were exposed to si-BAG3, and its growth was inhibited depending on modulation of cell cycle G1/S checkpoints and cell cycle regulators, involving cyclin D1, cyclin A2, and cyclin B1. Furthermore, suppression of the epithelial-mesenchymal transition (EMT) by si-BAG3 is linked to the decreased expression of E-cadherin and the increased expression of N-cadherin, vimentin, and MMP9. In conclusion, in the present study, we demonstrated that BAG3 overexpression plays a critical role in cell proliferation, migration, and invasion of colorectal cancer. Our data suggests targeted inhibition of BAG3 may be useful for patients with CRC.

Bax/Tubulin/Epithelial-Mesenchymal Pathways Determine the Efficacy of Silybin Analog HM015k in Colorectal Cancer Cell Growth and Metastasis.

PubMed

Amawi, Haneen; Hussein, Noor A; Ashby, Charles R; Alnafisah, Rawan; Sanglard, Leticia M; Manivannan, Elangovan; Karthikeyan, Chandrabose; Trivedi, Piyush; Eisenmann, Kathryn M; Robey, Robert W; Tiwari, Amit K

2018-01-01

The inhibition of apoptosis, disruption of cellular microtubule dynamics, and over-activation of the epithelial mesenchymal transition (EMT), are involved in the progression, metastasis, and resistance of colorectal cancer (CRC) to chemotherapy. Therefore, the design of a molecule that can target these pathways could be an effective strategy to reverse CRC progression and metastasis. In this study, twelve novel silybin derivatives, HM015a-HM015k (15a-15k) and compound 17, were screened for cytotoxicity in CRC cell lines. Compounds HM015j and HM015k (15k and 15j) significantly decreased cell proliferation, inhibited colony formation, and produced cell cycle arrest in CRC cells. Furthermore, 15k significantly induced the formation of reactive oxygen species and apoptosis. It induced the cleavage of the intrinsic apoptotic protein (Bax p21) to its more efficacious fragment, p18. Compound 15k also inhibited tubulin expression and disrupted its structure. Compound 15k significantly decreased metastatic LOVO cell migration and invasion. Furthermore, 15k reversed mesenchymal morphology in HCT116 and LOVO cells. Additionally, 15k significantly inhibited the expression of the mesenchymal marker N-cadherin and upregulated the expression of the epithelial marker, E-cadherin. Compound 15k inhibited the expression of key proteins known to induce EMT (i.e., DVL3, β-catenin, c-Myc) and upregulated the anti-metastatic protein, cyclin B1. Overall, in vitro , 15k significantly inhibited CRC progression and metastasis by inhibiting apoptosis, tubulin activity and the EMT pathways. Overall, these data suggest that compound 15k should be tested in vivo in a CRC animal model for further development.

MicroRNA-627 Mediates the Epigenetic Mechanisms of Vitamin D to Suppress Proliferation of Human Colorectal Cancer Cells and Growth of Xenograft Tumors in Mice

PubMed Central

Padi, Sathish K.R.; Zhang, Qunshu; Rustum, Youcef M; Morrison, Carl; Guo, Bin

2013-01-01

Background & Aims Vitamin D protects against colorectal cancer by unclear mechanisms. We investigated the effects of calcitriol (1α,25-dihydroxyvitamin D3, the active form of vitamin D) on levels of different microRNAs (miRs) in colorectal cancer (CRC) cells from humans and xenograft tumors in mice. Methods Expression of microRNAs in CRC cell lines was examined using the Ambion mirVana miRNA Bioarray. The effects of calcitriol on expression of miR-627 and cell proliferation were determined by real-time PCR and WST-1 assay, respectively; growth of colorectal xenograft tumors was examined in nude mice. Real-time PCR was used to analyze levels of miR-627 in human colon adenocarcinoma samples and non-tumor colon mucosa tissues (controls). Results In HT-29 cells, miR-627 was the only microRNA significantly upregulated by calcitriol. Jumonji domain containing 1A (JMJD1A), which encodes a histone demethylase, was found to be a target of miR-627. By downregulating JMJD1A, miR-627 increased methylation of histone H3K9 and suppressed expression of proliferative factors such as GDF15. Calcitriol induced expression of miR-627, which downregulated JMJD1A and suppressed growth of xenograft tumors from HCT-116 cells in nude mice. Overexpression of miR-627 prevented proliferation of CRC cell lines in culture and growth of xenograft tumors in mice. Conversely, blocking the activity of miR-627 inhibited the tumor suppressive effects of calcitriol in cultured CRC cells and in mice. Levels of miR-627 were decreased in human colon adenocarcinoma samples, compared with controls. Conclusions miR-627 mediates tumor-suppressive epigenetic activities of vitamin D on CRC cells and xenograft tumors in mice. The mRNA that encodes the histone demethylase JMJD1A is a direct target of miR-627. Reagents designed to target JMJD1A or its mRNA, or increase the function of miR-627, might have the same antitumor activities of vitamin D without the hypercalcemic side effects. PMID:23619147

Experimental verification of a predicted novel microRNA located in human PIK3CA gene with a potential oncogenic function in colorectal cancer.

PubMed

Saleh, Ali Jason; Soltani, Bahram M; Dokanehiifard, Sadat; Medlej, Abdallah; Tavalaei, Mahmoud; Mowla, Seyed Javad

2016-10-01

PI3K/AKT signaling is involved in cell survival, proliferation, and migration. In this pathway, PI3Kα enzyme is composed of a regulatory protein encoded by p85 gene and a catalytic protein encoded by PIK3CA gene. Human PIK3CA locus is amplified in several cancers including lung and colorectal cancer (CRC). Therefore, microRNAs (miRNAs) that are encoded within the PIK3CA gene might have a role in cancer development. Here, we report a novel microRNA named PIK3CA-miR1 (EBI accession no. LN626315), which is located within PIK3CA gene. A DNA segment corresponding to PIK3CA-premir1 sequence was transfected in human cell lines that resulted in generation of mature exogenous PIK3CA-miR1. Following the overexpression of PIK3CA-miR1, its predicted target genes (APPL1 and TrkC) were significantly downregulated in the CRC-originated HCT116 and SW480 cell lines, detected by qRT-PCR. Then, dual luciferase assay supported the interaction of PIK3CA-miR1 with APPL1 and TrkC transcripts. Endogenous PIK3CA-miR1 expression was also detected in several cell lines (highly in HCT116 and SW480) and highly in CRC specimens. Consistently, overexpression of PIK3CA-premir1 in HCT116 and SW480 cells resulted in significant reduction of the sub-G1 cell distribution and apoptotic cell rate, as detected by flowcytometry, and resulted in increased cell proliferation, as detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. PIK3CA-miR1 overexpression also resulted in Wnt signaling upregulation detected by Top/Fop assay. Overall, accumulative evidences indicated the presence of a bona fide novel onco-miRNA encoded within the PIK3CA oncogene, which is highly expressed in colorectal cancer and has a survival effect in CRC-originated cells.

Functional role of a long non-coding RNA LIFR-AS1/miR-29a/TNFAIP3 axis in colorectal cancer resistance to pohotodynamic therapy.

PubMed

Liu, Kuijie; Yao, Hongliang; Wen, Yu; Zhao, Hua; Zhou, Nanjiang; Lei, Sanlin; Xiong, Li

2018-05-25

Colorectal Cancer (CRC) is one of the most common digestive system malignant tumors. Recently, PDT has been used as a first-line treatment for colon cancer; however, limited curative effect was obtained due to resistance of CRC to PDT. During the past decades, accumulating CRC-related long non-coding RNAs (lncRNAs), microRNAs (miRNAs) and mRNAs have been reported to exert diverse functions through various biological processes; their dysregulation might trigger and/or promote the pathological changes. Herein, we performed microarrays analysis to identify dysregulated lncRNAs, miRNAs and mRNAs in PDT-treated HCT116 cells to figure out the lncRNA-miRNA interactions related to the resistance of CRC to PDT treatment, and the downstream mRNA target, as well as the molecular mechanism. We found a total of 1096 lncRNAs dysregulated in PDT-treated CRC HCT116 cells; among them, LIFR-AS1 negatively interacted with miR-29a, one of the dysregulated miRNAs in PDT-treated CRC cells, to affect the resistance of CRC to PDT. LIFR-AS1 knockdown attenuated, whereas miR-29a inhibition enhanced the cellular effect of PDT on HCT116 cell proliferation and apoptosis. Furthermore, among the dysregulated mRNAs, TNFAIP3 was confirmed to be a direct target of miR-29a and exerted a similar effect to LIFR-AS1 on the cellular effects of PDT. In summary, LIFR-AS1 serves as a competitive endogenous RNA (ceRNA) for miR-29a to inhibit its expression and up-regulate downstream target TNFAIP3 expression, finally modulating the resistance of CRC to PDT. We provide an experimental basis for this lncRNA/miRNA/mRNA network being a promising target in CRC resistance to PDT treatment. Copyright © 2018. Published by Elsevier B.V.

Piceatannol promotes apoptosis via up-regulation of microRNA-129 expression in colorectal cancer cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Haogang; Jia, Ruichun; Wang, Chunjing

2014-09-26

Highlights: • Piceatannol induces apoptosis in cultured CRC cells. • Piceatannol promotes expression of miR-129. • miR-129 mediates proapoptotic effects of piceatannol. - Abstract: Piceatannol, a naturally occurring analog of resveratrol, has been confirmed as an antitumor agent by inhibiting proliferation, migration, and metastasis in diverse cancer. However, the effect and mechanisms of piceatannol on colorectal cancer (CRC) have not been well understood. This study aimed to test whether piceatannol could inhibit growth of CRC cells and reveal its underlying molecular mechanism. MTT assay was used to detect the cell viability in HCT116 and HT29 cells. Flow cytometry analysis wasmore » employed to measure apoptosis of CRC cells. Bcl-2, Bax and caspase-3 levels were analyzed by Western blot and miR-129 levels were determined by real-time RT-PCR. Our study showed that piceatannol inhibited HCT116 and HT29 cells growth in a concentration- and time-dependent manner. Piceatannol induced apoptosis by promoting expression of miR-129, and then inhibiting expression of Bcl-2, an known target for miR-129. Moreover, knock down of miR-129 could reverse the reduction of cell viability induced by piceatannol in HCT116 and HT29 cells. Taken together, our study unraveled the ability of piceatannol to suppress colorectal cancer growth and elucidated the participation of miR-129 in the anti-cancer action of piceatannol. Our findings suggest that piceatannol can be considered to be a promising anticancer agent for CRC.« less

The chemokines CCR1 and CCRL2 have a role in colorectal cancer liver metastasis.

PubMed

Akram, Israa G; Georges, Rania; Hielscher, Thomas; Adwan, Hassan; Berger, Martin R

2016-02-01

C-C chemokine receptor type 1 (CCR1) and chemokine C-C motif receptor-like 2 (CCRL2) have not yet been sufficiently investigated for their role in colorectal cancer (CRC). Here, we investigated their expression in rat and human CRC samples, their modulation of expression in a rat liver metastasis model, as well as the effects on cellular properties resulting from their knockdown. One rat and five human colorectal cancer cell lines were used. CC531 rat colorectal cells were injected via the portal vein into rats and re-isolated from rat livers after defined periods. Following mRNA isolation, the gene expression was investigated by microarray. In addition, all cell lines were screened for mRNA expression of CCR1 and CCRL2 by reverse transcription polymerase chain reaction (RT-PCR). Cell lines with detectable expression were used for knockdown experiments; and the respective influence was determined on the cells' proliferation, scratch closure, and colony formation. Finally, specimens from the primaries of 50 patients with CRC were monitored by quantitative RT-PCR for CCR1 and CCRL2 expression levels. The microarray studies showed peak increases of CCR1 and CCRL2 in the early phase of liver colonization. Knockdown was sufficient at mRNA but only moderate at protein levels and resulted in modest but significant inhibition of proliferation (p < 0.05), scratch closure, and colony formation (p < 0.05). All human CRC samples were positive for CCR1 and CCRL2 and showed a significant pairwise correlation (p < 0.0004), but there was no correlation with tumor stage or age of patients. In summary, the data point to an important role of CCR1 and CCRL2 under conditions of organ colonization and both chemokine receptors qualify as targets of treatment during early colorectal cancer liver metastasis.

Efficacy of anti-RON antibody Zt/g4-drug maytansinoid conjugation (Anti-RON ADC) as a novel therapeutics for targeted colorectal cancer therapy.

PubMed

Feng, Liang; Yao, Hang-Ping; Wang, Wei; Zhou, Yong-Qing; Zhou, Jianwei; Zhang, Ruiwen; Wang, Ming-Hai

2014-12-01

The receptor tyrosine kinase RON is critical in epithelial tumorigenesis and a drug target for cancer therapy. Here, we report the development and therapeutic efficacy of a novel anti-RON antibody Zt/g4-maytansinoid (DM1) conjugates for targeted colorectal cancer (CRC) therapy. Zt/g4 (IgG1a/κ) was conjugated to DM1 via thioether linkage to form Zt/g4-DM1 with a drug-antibody ratio of 4:1. CRC cell lines expressing different levels of RON were tested in vitro to determine Zt/g4-DM1-induced RON endocytosis, cell-cycle arrest, and cytotoxicity. Efficacy of Zt/g4-DM1 in vivo was evaluated in mouse xenograft CRC tumor model. Zt/g4-DM1 rapidly induced RON endocytosis, arrested cell cycle at G2-M phase, reduced cell viability, and caused massive cell death within 72 hours. In mouse xenograft CRC models, Zt/g4-DM1 at a single dose of 20 mg/kg body weight effectively delayed CRC cell-mediated tumor growth up to 20 days. In a multiple dose-ranging study with a five injection regimen, Zt/g4-DM1 inhibited more than 90% tumor growth at doses of 7, 10, and 15 mg/kg body weight. The minimal dose achieving 50% of tumor inhibition was approximately 5.0 mg/kg. The prepared Zt/g4-DM1 is stable at 37°C for up to 30 days. At 60 mg/kg, Zt/g4-DM1 had a moderate toxicity in vivo with an average of 12% reduction in mouse body weight. Zt/g4-DM1 is highly effective in targeted inhibition of CRC cell-derived tumor growth in mouse xenograft models. This work provides the basis for development of humanized Zt/g4-DM1 for RON-targeted CRC therapy in the future. ©2014 American Association for Cancer Research.

Knockdown of astrocyte elevated gene-1 inhibits tumor growth and modifies microRNAs expression profiles in human colorectal cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Sujun; Southern Medical University, Guangzhou, Guangdong 510515; Wu, Binwen, E-mail: wubinwengd@aliyun.com

2014-02-14

Highlights: • AEG-1 expression in CRC cell lines and down-regulation or upregulation of AEG-1 in vitro. • Knockdown of AEG-1 inhibits cell proliferation, colony formation and invasion. • Upregulation of AEG-1 enhances proliferation, invasion and colony formation. • Knockdown of AEG-1 accumulates G0/G1-phase cells and promotes apoptosis in CRC cells. • AEG-1 knockdown increases 5-FU cytotoxicity. - Abstract: Astrocyte elevated gene-1 (AEG-1), upregulated in various types of malignancies including colorectal cancer (CRC), has been reported to be associated with the carcinogenesis. MicroRNAs (miRNAs) are widely involved in the initiation and progression of cancer. However, the functional significance of AEG-1 andmore » the relationship between AEG-1 and microRNAs in human CRC remains unclear. The aim of this study was to investigate whether AEG-1 could serve as a potential therapeutic target of human CRC and its possible mechanism. We adopted a strategy of ectopic overexpression or RNA interference to upregulate or downregulate expression of AEG-1 in CRC models. Their phenotypic changes were analyzed by Western blot, MTT and transwell matrix penetration assays. MicroRNAs expression profiles were performed using microarray analysis followed by validation using qRT-PCR. Knockdown of AEG-1 could significantly inhibit colon cancer cell proliferation, colony formation, invasion and promotes apoptosis. Conversely, upregulation of AEG-1 could significantly enhance cell proliferation, invasion and reduced apoptisis. AEG-1 directly contributes to resistance to chemotherapeutic drug. Targeted downregulation of AEG-1 might improve the expression of miR-181a-2{sup ∗}, -193b and -193a, and inversely inhibit miR-31 and -9{sup ∗}. Targeted inhibition of AEG-1 can lead to modification of key elemental characteristics, such as miRNAs, which may become a potential effective therapeutic strategy for CRC.« less

Identifying clinically relevant drug resistance genes in drug-induced resistant cancer cell lines and post-chemotherapy tissues.

PubMed

Tong, Mengsha; Zheng, Weicheng; Lu, Xingrong; Ao, Lu; Li, Xiangyu; Guan, Qingzhou; Cai, Hao; Li, Mengyao; Yan, Haidan; Guo, You; Chi, Pan; Guo, Zheng

2015-12-01

Until recently, few molecular signatures of drug resistance identified in drug-induced resistant cancer cell models can be translated into clinical practice. Here, we defined differentially expressed genes (DEGs) between pre-chemotherapy colorectal cancer (CRC) tissue samples of non-responders and responders for 5-fluorouracil and oxaliplatin-based therapy as clinically relevant drug resistance genes (CRG5-FU/L-OHP). Taking CRG5-FU/L-OHP as reference, we evaluated the clinical relevance of several types of genes derived from HCT116 CRC cells with resistance to 5-fluorouracil and oxaliplatin, respectively. The results revealed that DEGs between parental and resistant cells, when both were treated with the corresponding drug for a certain time, were significantly consistent with the CRG5-FU/L-OHP as well as the DEGs between the post-chemotherapy CRC specimens of responders and non-responders. This study suggests a novel strategy to extract clinically relevant drug resistance genes from both drug-induced resistant cell models and post-chemotherapy cancer tissue specimens.

Targeted delivery of anti-miR-155 by functionalized mesoporous silica nanoparticles for colorectal cancer therapy

PubMed Central

Bi, Jiangang; Zeng, Xiaowei; Mei, Lin; Bao, Shiyun; He, Lisheng; Shan, Aijun; Zhang, Yue; Yu, Xiaofang

2018-01-01

Introduction MicroRNA-155 (miR-155) is an oncogenic microRNA, which is upregulated in many human cancers including colorectal cancer (CRC). Overexpression of miR-155 has been found to regulate several cancer-related pathways, and therefore, targeting miR-155 may be an effective strategy for cancer therapy. However, effective and safe delivery of anti-miR-155 to tumors remains challenging for the clinical applications of anti-miR-155-based therapeutics. Methods In this study, we explored the expression of miR-155 and the transcription factor nuclear factor kappa B (NF-κB) in CRC tissues and cell lines, and the possible relationship between miR-155 and NF-κB. We further report on anti-miR-155-loaded mesoporous silica nanoparticles (MSNs) modified with polymerized dopamine (PDA) and AS1411 aptamer (MSNs-anti-miR-155@PDA-Apt) for the targeted treatment of CRC. Results Results showed that miR-155 is overexpressed in CRC tissues and cell lines, and there is a positive feedback loop between NF-κB and miR-155. Compared to the control groups, MSNs-anti-miR-155@PDA-Apt could efficiently downregulate miR-155 expression in SW480 cells and achieve significantly high targeting efficiency and enhanced therapeutic effects in both in vivo and in vitro experiments. Furthermore, inhibition of miR-155 by MSNs-anti-miR-155@PDA-Apt can enhance the sensitivity of SW480 to 5-fluorouracil chemotherapy. Conclusion Thus, our results suggested that MSNs-anti-miR-155@PDA-Apt is a promising nanoformulation for CRC treatment. PMID:29535520

Up-regulation of lncRNA SNHG1 indicates poor prognosis and promotes cell proliferation and metastasis of colorectal cancer by activation of the Wnt/β-catenin signaling pathway

PubMed Central

Zhu, Yuping; Li, Bo; Liu, Zhuo; Jiang, Lai; Wang, Gang; Lv, Min; Li, Dechuan

2017-01-01

Recently, the lncRNA small nucleolar RNA host gene (SNHG1) has been exhibited to be upregulated, which plays a crucial role in the development and prognosis of several cancers. However, the role of the biology and clinical significance of SNHG1 in the tumorigenesis of colorectal cancer (CRC) has rarely been reported. In this work, we firstly found that SNHG1 expression levels were upregulated aberrantly in colorectal cancer tissues and colorectal cancer cell lines. By Kaplan-Meier survival analysis, patients with high SNHG1 expression level had poorer overall survival (OS) and progression-free survival (PFS) than those with low SNHG1 expression. In multivariate analysis, increased SNHG1 expression was proved to be an independent unfavorable prognostic indicator for CRC. In vitro experiments revealed that SNHG1 silencing inhibited the growth and metastasis and induced apoptosis of CRC cell lines. Finally, we found that SNHG1 may induce the activation of the WNT/β-catenin pathway through regulating β-catenin expression and transcription factor-4 (TCF-4), cyclin D1 and MMP-9. Altogether, our findings demonstrated that lncRNA SNHG1, was high expressed in colorectal cancer tissues and may serve as a tumor oncogene through regulating WNT/β-catenin signal pathway, which provided a candidate diagnostic biomarker and a promising therapeutic target for patients with CRC. PMID:29340086

Role of serine/threonine kinase 33 methylation in colorectal cancer and its clinical significance

PubMed Central

Yin, Ming-Di; Ma, Si-Ping; Liu, Fang; Chen, Yu-Ze

2018-01-01

Serine/threonine kinase 33 (STK33) is a novel protein that has been the focus of an increasing number of studies in recent years; however, the role of STK33 in tumorigenesis remains controversial. Previous studies have demonstrated that STK33 is overexpressed in several human cancers and exerts a pro-tumorigenic effect through the promotion of cell proliferation. However, the role of STK33 in colorectal cancer (CRC), which is one of the most aggressive human malignancies, remains unclear. The aim of the current study was to investigate the methylation status of STK33 in CRC and to determine its clinical significance. The results demonstrated that STK33 was hypermethylated in CRC cell lines and promoted the proliferation of CRC cells. In addition, the methylation status and expression of STK33 in 94 pairs of cancer and noncancerous tissues obtained from patients with CRC was investigated. STK33 methylation was significantly increased in cancer tissues when compared with adjacent noncancerous tissues (P<0.001). STK33 methylation was associated with lymph node metastasis (P<0.05), tumor invasion (P<0.05), distant metastases (P<0.01) and tumor stage (P<0.01). Reduced STK33 mRNA and protein expression in CRC was associated with STK33 hypermethylation (P<0.001). In addition, patients with hypermethylated STK33 exhibited shorter overall survival rates when compared with those with unmethylated STK33 (P<0.01). In conclusion, the results of the current study suggest that STK33 hypermethylation may be a promising novel biomarker for the diagnosis, prognosis and suitable treatment of CRC. PMID:29434919

Overexpression of secretagogin promotes cell apoptosis and inhibits migration and invasion of human SW480 human colorectal cancer cells.

PubMed

Yang, Xiang-Yi; Liu, Qiao-Rui; Wu, Li-Ming; Zheng, Xu-Lei; Ma, Cong; Na, Ri-Su

2018-05-01

In order to investigate the effect of secretagogin (SCGN) on colorectal cancer (CRC) cells apoptosis, invasion and migration in vitro. Expression of SCGN in CRC tissues and the paired adjacent non-tumorous tissues (n = 36) and four human CRC cell lines (HT29, HCT116, SW480 and SW620) were detected. SW480 cells were transfected with the SCGN overexpression plasmid (eGFP-SCGN), si-SCGN-773, and the corresponding negative controls (NCs). Then, cell-cycle distribution, cell apoptosis, migration, invasion and expression of apoptosis- and metastasis-related proteins were detected. SCGN was significantly downregulated in CRC tissues as compared with the adjacent non-tumorous tissues. The expression of SCGN in HT29 and SW480 cells were lower than those in HT116 and SW620 cells. We transfected SW480 cells with SCGN overexpression plasmid eGFP-SCGN and found the increased cell apoptosis, with cell arresting at G0/G1 phase. SW480 cells with SCGN overexpression showed wider wound width and fewer invaded cells than control and blank cells, with upregulated Bax, cleaved Caspase 3 and E-cadherin, and downregulated Bcl-2 and Vimentin. We also transfected SW480 cells with si-SCGN-773 and found si-SCGN increased cell migration and invasion, but did not affect cell apoptosis and expression of related proteins. We concluded that the overexpression of SCGN in SW480 cells promoted cell apoptosis and inhibited cell migration and invasion. Copyright © 2018. Published by Elsevier Masson SAS.

TET1 Depletion Induces Aberrant CpG Methylation in Colorectal Cancer Cells

PubMed Central

Yamamoto, Eiichiro; Harada, Taku; Aoki, Hironori; Maruyama, Reo; Toyota, Mutsumi; Sasaki, Yasushi; Sugai, Tamotsu; Tokino, Takashi; Nakase, Hiroshi

2016-01-01

Aberrant DNA methylation is commonly observed in colorectal cancer (CRC), but the underlying mechanism is not fully understood. 5-hydroxymethylcytosine levels and TET1 expression are both reduced in CRC, while epigenetic silencing of TET1 is reportedly associated with the CpG island methylator phenotype. In the present study, we aimed to clarify the relationship between loss of TET1 and aberrant DNA methylation in CRC. Stable TET1 knockdown clones were established using Colo320DM cells, which express high levels of TET1, and HCT116 cells, which express TET1 at a level similar to that in normal colonic tissue. Infinium HumanMethylation450 BeadChip assays revealed increased levels of 5-methylcytosine at more than 10,000 CpG sites in TET1-depleted Colo320DM cells. Changes in DNA methylation were observed at various positions within the genome, including promoters, gene bodies and intergenic regions, and the altered methylation affected expression of a subset of genes. By contrast, TET1 knockdown did not significantly affect DNA methylation in HCT116 cells. However, TET1 depletion was associated with attenuated effects of 5-aza-2’-deoxycytidine on gene expression profiles in both cell lines. These results suggest that loss of TET1 may induce aberrant DNA methylation and may attenuate the effect of 5-aza-2’-deoxycytidine in CRC cells. PMID:27977763

Pre-45s rRNA promotes colon cancer and is associated with poor survival of CRC patients.

PubMed

Tsoi, H; Lam, K C; Dong, Y; Zhang, X; Lee, C K; Zhang, J; Ng, S C; Ng, S S M; Zheng, S; Chen, Y; Fang, J; Yu, J

2017-11-02

One characteristic of cancer cells is the abnormally high rate of cell metabolism to sustain their enhanced proliferation. However, the behind mechanism of this phenomenon is still elusive. Here we find that enhanced precursor 45s ribosomal RNA (pre-45s rRNA) is one of the core mechanisms in promoting the pathogenesis of colorectal cancer (CRC). Pre-45s rRNA expression is significantly higher in primary CRC tumor tissues samples and cancer cell lines compared with the non-tumorous colon tissues, and is associated with tumor sizes. Knockdown of pre-45s rRNA inhibits G1/S cell-cycle transition by stabilizing p53 through inducing murine double minute 2 (MDM2) and ribosomal protein L11 (RpL11) interaction. In addition, we revealed that high rate of cancer cell metabolism triggers the passive release of calcium ion from endoplasmic reticulum to the cytoplasm. The elevated calcium ion in the cytoplasm activates the signaling cascade of calcium/calmodulin-dependent protein kinase II, ribosomal S6 kinase (S6K) and ribosomal S6K (CaMKII-S6K-UBF). The activated UBF promotes the transcription of rDNA, which therefore increases pre-45s rRNA. Disruption of CaMKII-S6K-UBF axis by either RNAi or pharmaceutical approaches leads to reduction of pre-45s rRNA expression, which subsequently suppresses cell proliferation in colon cancer cells by causing cell-cycle arrest. Knockdown of APC activates CaMKII-S6K-UBF cascade and thus enhances pre-45s rRNA expression. Moreover, the high expression level of pre-45s rRNA is associated with poor survival of CRC patients in two independent cohorts. Our study identifies a novel mechanism in CRC pathogenesis mediated by pre-45s rRNA and a prognostic factor of pre-45s rRNA in CRC patients.

Systematic identification and validation of candidate genes for detection of circulating tumor cells in peripheral blood specimens of colorectal cancer patients.

PubMed

Findeisen, Peter; Röckel, Matthias; Nees, Matthias; Röder, Christian; Kienle, Peter; Von Knebel Doeberitz, Magnus; Kalthoff, Holger; Neumaier, Michael

2008-11-01

The presence of tumor cells in peripheral blood is being regarded increasingly as a clinically relevant prognostic factor for colorectal cancer patients. Current molecular methods are very sensitive but due to low specificity their diagnostic value is limited. This study was undertaken in order to systematically identify and validate new colorectal cancer (CRC) marker genes for improved detection of minimal residual disease in peripheral blood mononuclear cells of colorectal cancer patients. Marker genes with upregulated gene expression in colorectal cancer tissue and cell lines were identified using microarray experiments and publicly available gene expression data. A systematic iterative approach was used to reduce a set of 346 candidate genes, reportedly associated with CRC to a selection of candidate genes that were then further validated by relative quantitative real-time RT-PCR. Analytical sensitivity of RT-PCR assays was determined by spiking experiments with CRC cells. Diagnostic sensitivity as well as specificity was tested on a control group consisting of 18 CRC patients compared to 12 individuals without malignant disease. From a total of 346-screened genes only serine (or cysteine) proteinase inhibitor, clade B (ovalbumin), member 5 (SERPINB5) showed significantly elevated transcript levels in peripheral venous blood specimens of tumor patients when compared to the nonmalignant control group. These results were confirmed by analysis of an enlarged collective consisting of 63 CRC patients and 36 control individuals without malignant disease. In conclusion SERPINB5 seems to be a promising marker for detection of circulating tumor cells in peripheral blood of colorectal cancer patients.

T-oligo as an anticancer agent in colorectal cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wojdyla, Luke; Stone, Amanda L.; Sethakorn, Nan

Highlights: • T-oligo induces cell cycle arrest, senescence, apoptosis, and differentiation in CRC. • Treatment with T-oligo downregulates telomere-associated proteins. • T-oligo combined with an EGFR-TKI additively inhibits cellular proliferation. • T-oligo has potential as an effective therapeutic agent for CRC. - Abstract: In the United States, there will be an estimated 96,830 new cases of colorectal cancer (CRC) and 50,310 deaths in 2014. CRC is often detected at late stages of the disease, at which point there is no effective chemotherapy. Thus, there is an urgent need for effective novel therapies that have minimal effects on normal cells. T-oligo,more » an oligonucleotide homologous to the 3′-telomere overhang, induces potent DNA damage responses in multiple malignant cell types, however, its efficacy in CRC has not been studied. This is the first investigation demonstrating T-oligo-induced anticancer effects in two CRC cell lines, HT-29 and LoVo, which are highly resistant to conventional chemotherapies. In this investigation, we show that T-oligo may mediate its DNA damage responses through the p53/p73 pathway, thereby inhibiting cellular proliferation and inducing apoptosis or senescence. Additionally, upregulation of downstream DNA damage response proteins, including E2F1, p53 or p73, was observed. In LoVo cells, T-oligo induced senescence, decreased clonogenicity, and increased expression of senescence associated proteins p21, p27, and p53. In addition, downregulation of POT1 and TRF2, two components of the shelterin protein complex which protects telomeric ends, was observed. Moreover, we studied the antiproliferative effects of T-oligo in combination with an EGFR tyrosine kinase inhibitor, Gefitinib, which resulted in an additive inhibitory effect on cellular proliferation. Collectively, these data provide evidence that T-oligo alone, or in combination with other molecularly targeted therapies, has potential as an anti-cancer agent in CRC.« less

Curcumin Chemosensitizes 5-Fluorouracil Resistant MMR-Deficient Human Colon Cancer Cells in High Density Cultures

PubMed Central

Shakibaei, Mehdi; Buhrmann, Constanze; Kraehe, Patricia; Shayan, Parviz; Lueders, Cora; Goel, Ajay

2014-01-01

Objective Treatment of colorectal cancer (CRC) remains a clinical challenge, as more than 15% of patients are resistant to 5-Fluorouracil (5-FU)-based chemotherapeutic regimens, and tumor recurrence rates can be as high as 50–60%. Cancer stem cells (CSC) are capable of surviving conventional chemotherapies that permits regeneration of original tumors. Therefore, we investigated the effectiveness of 5-FU and plant polyphenol (curcumin) in context of DNA mismatch repair (MMR) status and CSC activity in 3D cultures of CRC cells. Methods High density 3D cultures of CRC cell lines HCT116, HCT116+ch3 (complemented with chromosome 3) and their corresponding isogenic 5-FU-chemo-resistant derivative clones (HCT116R, HCT116+ch3R) were treated with 5-FU either without or with curcumin in time- and dose-dependent assays. Results Pre-treatment with curcumin significantly enhanced the effect of 5-FU on HCT116R and HCR116+ch3R cells, in contrast to 5-FU alone as evidenced by increased disintegration of colonospheres, enhanced apoptosis and by inhibiting their growth. Curcumin and/or 5-FU strongly affected MMR-deficient CRC cells in high density cultures, however MMR-proficient CRC cells were more sensitive. These effects of curcumin in enhancing chemosensitivity to 5-FU were further supported by its ability to effectively suppress CSC pools as evidenced by decreased number of CSC marker positive cells, highlighting the suitability of this 3D culture model for evaluating CSC marker expression in a close to vivo setting. Conclusion Our results illustrate novel and previously unrecognized effects of curcumin in enhancing chemosensitization to 5-FU-based chemotherapy on DNA MMR-deficient and their chemo-resistant counterparts by targeting the CSC sub-population. (246 words in abstract). PMID:24404205

Curcumin chemosensitizes 5-fluorouracil resistant MMR-deficient human colon cancer cells in high density cultures.

PubMed

Shakibaei, Mehdi; Buhrmann, Constanze; Kraehe, Patricia; Shayan, Parviz; Lueders, Cora; Goel, Ajay

2014-01-01

Treatment of colorectal cancer (CRC) remains a clinical challenge, as more than 15% of patients are resistant to 5-Fluorouracil (5-FU)-based chemotherapeutic regimens, and tumor recurrence rates can be as high as 50-60%. Cancer stem cells (CSC) are capable of surviving conventional chemotherapies that permits regeneration of original tumors. Therefore, we investigated the effectiveness of 5-FU and plant polyphenol (curcumin) in context of DNA mismatch repair (MMR) status and CSC activity in 3D cultures of CRC cells. High density 3D cultures of CRC cell lines HCT116, HCT116+ch3 (complemented with chromosome 3) and their corresponding isogenic 5-FU-chemo-resistant derivative clones (HCT116R, HCT116+ch3R) were treated with 5-FU either without or with curcumin in time- and dose-dependent assays. Pre-treatment with curcumin significantly enhanced the effect of 5-FU on HCT116R and HCR116+ch3R cells, in contrast to 5-FU alone as evidenced by increased disintegration of colonospheres, enhanced apoptosis and by inhibiting their growth. Curcumin and/or 5-FU strongly affected MMR-deficient CRC cells in high density cultures, however MMR-proficient CRC cells were more sensitive. These effects of curcumin in enhancing chemosensitivity to 5-FU were further supported by its ability to effectively suppress CSC pools as evidenced by decreased number of CSC marker positive cells, highlighting the suitability of this 3D culture model for evaluating CSC marker expression in a close to vivo setting. Our results illustrate novel and previously unrecognized effects of curcumin in enhancing chemosensitization to 5-FU-based chemotherapy on DNA MMR-deficient and their chemo-resistant counterparts by targeting the CSC sub-population. (246 words in abstract).

Comprehensive Analysis of miRNome Alterations in Response to Sorafenib Treatment in Colorectal Cancer Cells

PubMed Central

Pehserl, Anna-Maria; Ress, Anna Lena; Stanzer, Stefanie; Resel, Margit; Karbiener, Michael; Stadelmeyer, Elke; Stiegelbauer, Verena; Gerger, Armin; Mayr, Christian; Scheideler, Marcel; Hutterer, Georg C.; Bauernhofer, Thomas; Kiesslich, Tobias; Pichler, Martin

2016-01-01

MicroRNAs (miRNAs) are master regulators of drug resistance and have been previously proposed as potential biomarkers for the prediction of therapeutic response in colorectal cancer (CRC). Sorafenib, a multi-kinase inhibitor which has been approved for the treatment of liver, renal and thyroid cancer, is currently being studied as a monotherapy in selected molecular subtypes or in combination with other drugs in metastatic CRC. In this study, we explored sorafenib-induced cellular effects in Kirsten rat sarcoma viral oncogene homolog olog (KRAS) wild-type and KRAS-mutated CRC cell lines (Caco-2 and HRT-18), and finally profiled expression changes of specific miRNAs within the miRNome (>1000 human miRNAs) after exposure to sorafenib. Overall, sorafenib induced a time- and dose-dependent growth-inhibitory effect through S-phase cell cycle arrest in KRAS wild-type and KRAS-mutated CRC cells. In HRT-18 cells, two human miRNAs (hsa-miR-597 and hsa-miR-720) and two small RNAs (SNORD 13 and hsa-miR-3182) were identified as specifically sorafenib-induced. In Caco-2 cells, nine human miRNAs (hsa-miR-3142, hsa-miR-20a, hsa-miR-4301, hsa-miR-1290, hsa-miR-4286, hsa-miR-3182, hsa-miR-3142, hsa-miR-1246 and hsa-miR-720) were identified to be differentially regulated post sorafenib treatment. In conclusion, we confirmed sorafenib as a potential anti-neoplastic treatment strategy for CRC cells by demonstrating a growth-inhibitory and cell cycle–arresting effect of this drug. Changes in the miRNome indicate that some specific miRNAs might be relevant as indicators for sorafenib response, drug resistance and potential targets for combinatorial miRNA-based drug strategies. PMID:27916938

Terpinen-4-ol inhibits colorectal cancer growth via reactive oxygen species

PubMed Central

Nakayama, Ken; Murata, Soichiro; Ito, Hiromu; Iwasaki, Kenichi; Villareal, Myra Orlina; Zheng, Yun-Wen; Matsui, Hirofumi; Isoda, Hiroko; Ohkohchi, Nobuhiro

2017-01-01

Terpinen-4-ol (TP4O) is the main component of the essential oil extracted from Melaleuca alternifolia, known as the tea tree, of the botanical family Myrtaceae. The anticancer effects of TP4O have been reported in several cancer cell lines. Previous reports have demonstrated that TP4O exerts anticancer effects by inducing apoptotic cell death in several cell lines; however, the underlying molecular mechanisms of these effects remain unclear. In the present study, the anticancer effects of TP4O against the colorectal cancer (CRC) cell lines HCT116 and RKO were evaluated using WST-8 and bromodeoxyuridine assays. The mechanism of cell death was investigated by the measurement of caspase-3/7, Annexin V and lactate dehydrogenase release. Reactive oxygen species (ROS) levels induced by TP4O were evaluated by electron spin resonance and quantitative measurement of dihydroethidium. Localization of the ROS derived from mitochondria was observed by confocal inverted microscopy. Protein levels of ROS scavengers were assessed by western blotting analysis. To confirm the role of ROS, cell viability was measured in the presence of antioxidant reagents. In an in vivo xenograft model of ICR-SCID mice implanted with HCT116 cells, 200 mg/kg TP4O was injected locally, and tumor growth was compared with that of the control. TP4O induced apoptotic cell death in HCT116 and RKO cells in a dose-dependent manner, and TP4O also increased the levels of ROS generated by mitochondria. TP4O-induced cell death was rescued by administration of antioxidant regents. In vivo, TP4O inhibited the proliferation of HCT116 xenografts compared with that of the control group. The results of the present study suggest that TP4O induces apoptosis in CRC cells through ROS generation. Furthermore, TP4O is potentially useful for the development of novel therapies against CRC. PMID:28781645

Effects of PHA-665752 and vemurafenib combination treatment on in vitro and murine xenograft growth of human colorectal cancer cells with BRAFV600E mutations.

PubMed

Zhi, Jie; Li, Zhongxin; Lv, Jian; Feng, Bo; Yang, Donghai; Xue, Liang; Zhao, Zhaolong; Zhang, Yanni; Wu, Jianhua; Jv, Yingchao; Jia, Yitao

2018-03-01

It remains unknown whether blockade of B-Raf proto-oncogene, serine/threonine kinase (BRAF) V600E signaling and MET proto-oncogene, receptor tyrosine kinase (c-Met) signaling is effective in suppressing the growth of human colorectal cancer (CRC) cells. The present study investigated the effects of the vemurafenib alone and in combination with c-Met inhibitor PHA-665752 on the growth of human CRC cells in vitro and in mouse xenografts. HT-29 and RKO CRC cell lines with BRAF V600E mutations and mice bearing HT-29 xenografts were treated with vemurafenib in the absence or presence of PHA-665752. Cell viability and cycle phase were respectively examined by using the MTT and flow cytometry assay. Immunohistochemistry was conducted to detect the protein expression levels of hepatocyte growth factor (HGF), phosphorylated (p)-c-Met, p-AKT serine/threonine kinase (AKT) and p-extracellular signal-regulated kinase (p-ERK). The MTT assay demonstrated that the growth of RKO and HT-29 cells was inhibited by PHA-665752 in a time- and dose-dependent manner (P<0.05), however no significant suppressive effects were observed with vemurafenib. Relative to the PHA-665752 or vemurafenib stand-alone treatment groups, the combination of PHA-665752 and vemurafenib had a significant inhibitory effect on the proliferation of CRC cell lines (P<0.05). The mean tumor volume in mice treated with vemurafenib in combination with PHA-665752 was significantly smaller compared with those treated with only vemurafenib or PHA-665752 (P<0.05). Flow cytometry assay revealed that the G0/G1 phase frequency was significantly increased in the combination group compared with any other treatment groups (P<0.05). Immunohistochemistry demonstrated that vemurafenib in combination with PHA-665752 effectively induced the expression of p-c-Met, p-AKT and p-ERK, however had no effect on HGF.

Identifying clinically relevant drug resistance genes in drug-induced resistant cancer cell lines and post- chemotherapy tissues

PubMed Central

Tong, Mengsha; Zheng, Weicheng; Lu, Xingrong; Ao, Lu; Li, Xiangyu; Guan, Qingzhou; Cai, Hao; Li, Mengyao; Yan, Haidan; Guo, You; Chi, Pan; Guo, Zheng

2015-01-01

Until recently, few molecular signatures of drug resistance identified in drug-induced resistant cancer cell models can be translated into clinical practice. Here, we defined differentially expressed genes (DEGs) between pre-chemotherapy colorectal cancer (CRC) tissue samples of non-responders and responders for 5-fluorouracil and oxaliplatin-based therapy as clinically relevant drug resistance genes (CRG5-FU/L-OHP). Taking CRG5-FU/L-OHP as reference, we evaluated the clinical relevance of several types of genes derived from HCT116 CRC cells with resistance to 5-fluorouracil and oxaliplatin, respectively. The results revealed that DEGs between parental and resistant cells, when both were treated with the corresponding drug for a certain time, were significantly consistent with the CRG5-FU/L-OHP as well as the DEGs between the post-chemotherapy CRC specimens of responders and non-responders. This study suggests a novel strategy to extract clinically relevant drug resistance genes from both drug-induced resistant cell models and post-chemotherapy cancer tissue specimens. PMID:26515599

5-ASA affects cell cycle progression in colorectal cells by reversibly activating a replication checkpoint.

PubMed

Luciani, M Gloria; Campregher, Christoph; Fortune, John M; Kunkel, Thomas A; Gasche, Christoph

2007-01-01

Individuals with inflammatory bowel disease are at risk of developing colorectal cancer (CRC). Epidemiologic, animal, and laboratory studies suggest that 5-amino-salicylic acid (5-ASA) protects from the development of CRC by altering cell cycle progression and by inducing apoptosis. Our previous results indicate that 5-ASA improves replication fidelity in colorectal cells, an effect that is active in reducing mutations. In this study, we hypothesized that 5-ASA restrains cell cycle progression by activating checkpoint pathways in colorectal cell lines, which would prevent tumor development and improve genomic stability. CRC cells with different genetic backgrounds such as HT29, HCT116, HCT116(p53-/-), HCT116+chr3, and LoVo were treated with 5-ASA for 2-96 hours. Cell cycle progression, phosphorylation, and DNA binding of cell cycle checkpoint proteins were analyzed. We found that 5-ASA at concentrations between 10 and 40 mmol/L affects cell cycle progression by inducing cells to accumulate in the S phase. This effect was independent of the hMLH1, hMSH2, and p53 status because it was observed to a similar extent in all cell lines under investigation. Moreover, wash-out experiments demonstrated reversibility within 48 hours. Although p53 did not have a causative role, p53 Ser15 was strongly phosphorylated. Proteins involved in the ATM-and-Rad3-related kinase (ATR)-dependent S-phase checkpoint response (Chk1 and Rad17) were also phosphorylated but not ataxia telengectasia mutated kinase. Our data demonstrate that 5-ASA causes cells to reversibly accumulate in S phase and activate an ATR-dependent checkpoint. The activation of replication checkpoint may slow down DNA replication and improve DNA replication fidelity, which increases the maintenance of genomic stability and counteracts carcinogenesis.

The Ras effector RASSF2 is a novel tumor-suppressor gene in human colorectal cancer.

PubMed

Akino, Kimishige; Toyota, Minoru; Suzuki, Hiromu; Mita, Hiroaki; Sasaki, Yasushi; Ohe-Toyota, Mutsumi; Issa, Jean-Pierre J; Hinoda, Yuji; Imai, Kohzoh; Tokino, Takashi

2005-07-01

Activation of Ras signaling is a hallmark of colorectal cancer (CRC), but the roles of negative regulators of Ras are not fully understood. Our aim was to address that question by surveying genetic and epigenetic alterations of Ras-Ras effector genes in CRC cells. The expression and methylation status of 6 RASSF family genes were examined using RT-PCR and bisulfite PCR in CRC cell lines and in primary CRCs and colorectal adenomas. Colony formation assays and flow cytometry were used to assess the tumor suppressor activities of RASSF1 and RASSF2. Immunofluorescence microscopy was used to determine the effect of altered RASSF2 expression on cell morphology. Mutations of K- ras , BRAF, and p53 were identified using single-strand conformation analysis and direct sequencing. Aberrant methylation and histone deacetylation of RASSF2 was associated with the gene's silencing in CRC. The activities of RASSF2, which were distinct from those of RASSF1, included induction of morphologic changes and apoptosis; moreover, its ability to prevent cell transformation suggests that RASSF2 acts as a tumor suppressor in CRC. Primary CRCs that showed K- ras /BRAF mutations also frequently showed RASSF2 methylation, and inactivation of RASSF2 enhanced K- ras -induced oncogenic transformation. RASSF2 methylation was also frequently identified in colorectal adenomas. RASSF2 is a novel tumor suppressor gene that regulates Ras signaling and plays a pivotal role in the early stages of colorectal tumorigenesis.

Smad4 sensitizes colorectal cancer to 5-fluorouracil through cell cycle arrest by inhibiting the PI3K/Akt/CDC2/survivin cascade.

PubMed

Zhang, Binhao; Leng, Chao; Wu, Chao; Zhang, Zhanguo; Dou, Lei; Luo, Xin; Zhang, Bixiang; Chen, Xiaoping

2016-03-01

5-Fluorouracil (5-FU), a cell cycle-specific antimetabolite, is one of the most commonly used chemotherapeutic agents for colorectal cancer (CRC). Yet, resistance to 5-FU-based chemotherapy is still an obstacle to the treatment of this malignancy. Mutation or loss of Smad4 in CRC is pivotal for chemoresistance. However, the mechanism by which Smad4 regulates the chemosensitivity of CRC remains unclear. In the present study, we investigated the role of Smad4 in the chemosensitivity of CRC to 5-FU, and whether Smad4-regulated cell cycle arrest is involved in 5-FU chemoresistance. We used Smad4-expressing CT26 and Smad4-null SW620 cell lines as experimental models, by knockdown or transgenic overexpression. Cells or tumors were treated with 5-FU to determine chemosensitivity by cell growth, tumorigenicity assay and a mouse model. Cell cycle distribution was examined with flow cytometric analysis, and cell cycle-related proteins were examined by western blotting. Smad4 deficiency in CT26 and SW620 cells induced chemoresistance to 5-FU both in vitro and in vivo. Smad4 deficiency attenuated G1 or G2 cell cycle arrest by activating the PI3K/Akt/CDC2/survivin pathway. The PI3K inhibitor, LY294002, reversed the activation of the Akt/CDC2/survivin cascade in the Smad4-deficient cells, while it had little effect on cells with high Smad4 expression. In conclusion, we discovered a novel mechanism mediated by Smad4 to trigger 5-FU chemosensitivity through cell cycle arrest by inhibiting the PI3K/Akt/CDC2/survivin cascade. The present study also implies that LY294002 has potential therapeutic value to reverse the chemosensitivity of CRC with low Smad4 expression.

Single-cell genetic analysis validates cytopathological identification of circulating cancer cells in patients with clear cell renal cell carcinoma.

PubMed

Broncy, Lucile; Njima, Basma Ben; Méjean, Arnaud; Béroud, Christophe; Romdhane, Khaled Ben; Ilie, Marius; Hofman, Veronique; Muret, Jane; Hofman, Paul; Bouhamed, Habiba Chaabouni; Paterlini-Bréchot, And Patrizia

2018-04-13

Circulating Rare Cells (CRC) are non-haematological cells circulating in blood. They include Circulating Cancer Cells (CCC) and cells with uncertain malignant features (CRC-UMF) according to cytomorphology. Clear cell renal cell carcinomas frequently bear a mutated Von Hippel-Lindau (VHL) gene. To match blind genetic analysis of CRC and tumor samples with CRC cytopathological diagnosis. 29/30 patients harboured CRC (20 harboured CCC, 29 CRC-UMF) and 25/29 patients carried VHL mutations in their tumour. 205 single CRC (64 CCC, 141 CRC-UMF) provided genetic data. 57/57 CCC and 104/125 CRC-UMF from the 25 patients with VHL-mutated tumor carried the same VHL mutation detected in the tumor. Seven CCC and 16 CRC-UMF did not carry VHL mutations but were found in patients with wild-type VHL tumor tissue. All the CCC and 83,2% (104/125) of the CRC-UMF were found to carry the same VHL mutation identified in the corresponding tumorous tissue, validating cytopathological identification of CCC in patients with clear cell renal cell carcinoma. The blood of 30 patients with clear cell renal cell carcinoma was treated by ISET ® for CRC isolation, cytopathology and single-cell VHL mutations analysis, performed blindly and compared to VHL mutations of corresponding tumor tissues and leukocytes.

Genetic and epigenetic markers in colorectal cancer screening: recent advances.

PubMed

Singh, Manish Pratap; Rai, Sandhya; Suyal, Shradha; Singh, Sunil Kumar; Singh, Nand Kumar; Agarwal, Akash; Srivastava, Sameer

2017-07-01

Colorectal cancer (CRC) is a heterogenous disease which develops from benign intraepithelial lesions known as adenomas to malignant carcinomas. Acquired alterations in Wnt signaling, TGFβ, MAPK pathway genes and clonal propagation of altered cells are responsible for this transformation. Detection of adenomas or early stage cancer in asymptomatic patients and better prognostic and predictive markers is important for improving the clinical management of CRC. Area covered: In this review, the authors have evaluated the potential of genetic and epigenetic alterations as markers for early detection, prognosis and therapeutic predictive potential in the context of CRC. We have discussed molecular heterogeneity present in CRC and its correlation to prognosis and response to therapy. Expert commentary: Molecular marker based CRC screening methods still fail to gain trust of clinicians. Invasive screening methods, molecular heterogeneity, chemoresistance and low quality test samples are some key challenges which need to be addressed in the present context. New sequencing technologies and integrated omics data analysis of individual or population cohort results in GWAS. MPE studies following a GWAS could be future line of research to establish accurate correlations between CRC and its risk factors. This strategy would identify most reliable biomarkers for CRC screening and management.

Fusobacterium nucleatum Increases Proliferation of Colorectal Cancer Cells and Tumor Development in Mice by Activating Toll-Like Receptor 4 Signaling to Nuclear Factor-κB, and Up-regulating Expression of MicroRNA-21.

PubMed

Yang, Yongzhi; Weng, Wenhao; Peng, Junjie; Hong, Leiming; Yang, Lei; Toiyama, Yuji; Gao, Renyuan; Liu, Minfeng; Yin, Mingming; Pan, Cheng; Li, Hao; Guo, Bomin; Zhu, Qingchao; Wei, Qing; Moyer, Mary-Pat; Wang, Ping; Cai, Sanjun; Goel, Ajay; Qin, Huanlong; Ma, Yanlei

2017-03-01

Nearly 20% of the global cancer burden can be linked to infectious agents. Fusobacterium nucleatum promotes tumor formation by epithelial cells via unclear mechanisms. We aimed to identify microRNAs (miRNAs) induced by F nucleatum and evaluate their ability to promote colorectal carcinogenesis in mice. Colorectal cancer (CRC) cell lines were incubated with F nucleatum or control reagents and analyzed in proliferation and would healing assays. HCT116, HT29, LoVo, and SW480 CRC cell lines were incubated with F nucleatum or phosphate-buffered saline (PBS [control]) and analyzed for miRNA expression patterns and in chromatin immunoprecipitation assays. Cells were incubated with miRNAs mimics, control sequences, or small interfering RNAs; expression of reporter constructs was measured in luciferase assays. CRC cells were incubated with F nucleatum or PBS and injected into BALB/C nude mice; growth of xenograft tumors was measured. C57BL adenomatous polyposis coli min/+ , C57BL miR21a -/- , and C57BL mice with full-length miR21a (controls) were given F nucleatum by gavage; some mice were given azoxymethane and dextran sodium sulfate to induce colitis and colon tumors. Intestinal tissues were collected and tumors were counted. Serum samples from mice were analyzed for cytokine levels by enzyme-linked immunosorbent assay. We performed in situ hybridization analyses to detect enrichment of F nucleatum in CRC cells. Fusobacterium nucleatum DNA in 90 tumor and matched nontumor tissues from patients in China were explored for the expression correlation analysis; levels in 125 tumor tissues from patients in Japan were compared with their survival times. Fusobacterium nucleatum increased proliferation and invasive activities of CRC cell lines compared with control cells. CRC cell lines infected with F nucleatum formed larger tumors, more rapidly, in nude mice than uninfected cells. Adenomatous polyposis coli min/+ mice gavaged with F nucleatum developed significantly more colorectal tumors than mice given PBS and had shorter survival times. We found several inflammatory factors to be significantly increased in serum from mice given F nucleatum (interleukin 17F, interleukin 21, and interleukin 22, and MIP3A). We found 50 miRNAs to be significantly up-regulated and 52 miRNAs to be significantly down-regulated in CRCs incubated with F nucleatum vs PBS; levels of miR21 increased by the greatest amount (>4-fold). Inhibitors of miR21 prevented F nucleatum from inducing cell proliferation and invasion in culture. miR21a -/- mice had a later appearance of fecal blood and diarrhea after administration of azoxymethane and dextran sodium sulfate, and had longer survival times compared with control mice. The colorectum of miR21a -/- mice had fewer tumors, of smaller size, and the miR21a -/- mice survived longer than control mice. We found RASA1, which encodes an RAS GTPase, to be one of the target genes consistently down-regulated in cells that overexpressed miR21 and up-regulated in cells exposed to miR21 inhibitors. Infection of cells with F nucleatum increased expression of miR21 by activating Toll-like receptor 4 signaling to MYD88, leading to activation of the nuclear factor-κB. Levels of F nucleatum DNA and miR21 were increased in tumor tissues (and even more so in advanced tumor tissues) compared with non-tumor colon tissues from patients. Patients whose tumors had high amounts of F nucleatum DNA and miR21 had shorter survival times than patients whose tumors had lower amounts. We found infection of CRC cells with F nucleatum to increase their proliferation, invasive activity, and ability to form xenograft tumors in mice. Fusobacterium nucleatum activates Toll-like receptor 4 signaling to MYD88, leading to activation of the nuclear factor-κB and increased expression of miR21; this miRNA reduces levels of the RAS GTPase RASA1. Patients with both high amount of tissue F nucleatum DNA and miR21 demonstrated a higher risk for poor outcomes. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Preclinical Efficacy of [V4Q5]dDAVP, a Second Generation Vasopressin Analog, on Metastatic Spread and Tumor-Associated Angiogenesis in Colorectal Cancer.

PubMed

Garona, Juan; Sobol, Natasha T; Pifano, Marina; Segatori, Valeria I; Gomez, Daniel E; Ripoll, Giselle V; Alonso, Daniel F

2018-06-01

Control of metastatic spread of colorectal cancer (CRC) remains as a major therapeutic challenge. [V4Q5]dDAVP is a vasopressin peptide analog with previously reported anticancer activity against carcinoma tumors. By acting as a selective agonist of AVPR2 present in endothelial and tumor cells, [V4Q5]dDAVP is able to impair tumor aggressiveness and distant spread. Our aim was to evaluate the potential therapeutic benefits of [V4Q5]dDAVP on highly aggressive CRC disease using experimental models with translational relevance. Murine CT-26 and human Colo-205 AVPR2-expressing CRC cell lines were used to test the preclinical efficacy of [V4Q5]dDAVP, both in vitro and in vivo. In syngeneic mice surgically implanted with CT-26 cells in the spleen, sustained i.v. treatment with [V4Q5]dDAVP (0.3 µg/kg) dramatically impaired metastatic progression to liver without overt signs of toxicity, and also reduced experimental lung colonization. The compound inhibited in vivo angiogenesis driven by Colo-205 cells in athymic mice, as well as in vitro endothelial cell migration and capillary tube formation. [V4Q5]dDAVP exerted AVPR2-dependent cytostatic activity in vitro (IC50 1.08 µM) and addition to 5-FU resulted in synergistic antiproliferative effects both in CT-26 and Colo-205 cells. The present preclinical study establishes for the first time the efficacy of [V4Q5]dDAVP on CRC. These encouraging results suggest that the novel second generation vasopressin analog could be used for the management of aggressive CRC as an adjuvant agent during surgery or to complement standard chemotherapy, limiting tumor angiogenesis and metastasis and thus protecting the patient from CRC recurrence.

Colorectal Cancer Metastases Settle in the Hepatic Microenvironment Through α5β1 Integrin.

PubMed

Pelillo, Chiara; Bergamo, Alberta; Mollica, Hilaria; Bestagno, Marco; Sava, Gianni

2015-10-01

Colorectal cancer (CRC) metastasis dissemination to secondary sites represents the critical point for the patient's survival. The microenvironment is crucial to cancer progression, influencing tumour cell behaviour by modulating the expression and activation of molecules such as integrins, the cell-extracellular matrix interacting proteins participating in different steps of the tumour metastatic process. In this work, we investigated the role of α5β1 integrin and how the microenvironment influences this adhesion molecule, in a model of colon cancer progression to the liver. The culture medium conditioned by the IHH hepatic cell line, and the extracellular matrix (ECM) proteins, modulate the activation of α5β1 integrin in the colon cancer cell line HCT-116, and drives FAK phosphorylation during the process of cell adhesion to fibronectin, one of the main components of liver ECM. In these conditions, α5β1 modulates the expression/activity of another integrin, α2β1, involved in the cell adhesion to collagen I. These results suggest that α5β1 integrin holds a leading role in HCT-116 colorectal cancer cells adhesion to the ECM through the modulation of the intracellular focal adhesion kinase FAK and the α2β1 integrin activity. The driving role of the tumour microenvironment on CRC dissemination, here detected, and described, strengthens and adds new value to the concept that α5β1 integrin can be an appropriate and relevant therapeutic target for the control of CRC metastases. © 2015 Wiley Periodicals, Inc.

Blockade of LAG3 enhances responses of tumor-infiltrating T cells in mismatch repair-proficient liver metastases of colorectal cancer

PubMed Central

Noordam, Lisanne; Sprengers, Dave; Boor, Patrick P. C.; Mancham, Shanta; Menon, Anand G.; Lange, Johan F.; Burger, Pim J. W. A.; Brandt, Alexandra; Galjart, Boris; Kwekkeboom, Jaap; Bruno, Marco J.

2018-01-01

ABSTRACT Purpose: Liver metastasis develops in >50% of patients with colorectal cancer (CRC), and is a leading cause of CRC-related mortality. We aimed to identify which inhibitory immune checkpoint pathways can be targeted to enhance functionality of intra-tumoral T-cells in mismatch repair-proficient liver metastases of colorectal cancer (LM-CRC). Methodology: Intra-tumoral expression of multiple inhibitory molecules was compared among mismatch repair-proficient LM-CRC, peritoneal metastases of colorectal cancer (PM-CRC) and primary CRC. Expression of inhibitory molecules was also analyzed on leukocytes isolated from paired resected metastatic liver tumors, tumor-free liver tissues, and blood of patients with mismatch repair-proficient LM-CRC. The effects of blocking inhibitory pathways on tumor-infiltrating T-cell responses were studied in ex vivo functional assays. Results: Mismatch repair-proficient LM-CRC showed higher expression of inhibitory receptors on intra-tumoral T-cells and contained higher proportions of CD8+ T-cells, dendritic cells and monocytes than mismatch repair-proficient primary CRC and/or PM-CRC. Inhibitory receptors LAG3, PD-1, TIM3 and CTLA4 were higher expressed on CD8+ T-cells, CD4+ T-helper and/or regulatory T-cells in LM-CRC tumors compared with tumor-free liver and blood. Antibody blockade of LAG3 or PD-L1 increased proliferation and effector cytokine production of intra-tumoral T-cells isolated from LM-CRC in response to both polyclonal and autologous tumor-specific stimulations. Higher LAG3 expression on intra-tumoral CD8+ T-cells associated with longer progression-free survival of LM-CRC patients. Conclusion: Mismatch repair-proficient LM-CRC may be more sensitive to immune checkpoint inhibitors than mismatch repair-proficient primary CRC. Blocking LAG3 enhances tumor-infiltrating T-cell responses of mismatch repair-proficient LM-CRC, and therefore may be a new promising immunotherapeutic target for LM-CRC.

Temperature induces significant changes in both glycolytic reserve and mitochondrial spare respiratory capacity in colorectal cancer cell lines

PubMed Central

Mitov, Mihail I.; Harris, Jennifer W.; Alstott, Michael C.; Zaytseva, Yekaterina Y.; Evers, B. Mark; Butterfield, D. Allan

2017-01-01

Thermotherapy, as a method of treating cancer, has recently attracted considerable attention from basic and clinical investigators. A number of studies and clinical trials have shown that thermotherapy can be successfully used as a therapeutic approach for various cancers. However, the effects of temperature on cancer bioenergetics have not been studied in detail with a real time, in a microplate, label-free detection approach. This study investigate how changes in temperature affect the bioenergetics characteristics (mitochondrial function and glycolysis) of three colorectal cancer (CRC) cell lines utilizing the Seahorse XF96 technology. Experiments were performed at 32°C, 37°C and 42°C using assay medium conditions and equipment settings adjusted to produce equal oxygen and pH levels ubiquitously at the beginning of all experiments. The results suggest that temperature significantly changes multiple components of glycolytic and mitochondrial function of all cell lines tested. Under hypothermia conditions (32°C), the extracellular acidification rates (ECAR) of CRC cells were significantly lower compared to the same basal ECAR levels measured at 37°C. Mitochondrial stress test for SW480 cells at 37°C vs 42°C demonstrated increased proton leak while all other OCR components remained unchanged (similar results were detected also for the patient-derived xenograft cells Pt.93). Interestingly, the FCCP dose response at 37°C vs 42°C show significant shifts in profiles, suggesting that single dose FCCP experiments might not be sufficient to characterize the mitochondrial metabolic potential when comparing groups, conditions or treatments. These findings provide valuable insights for the metabolic and bioenergetic changes of CRC cells under hypo- and hyperthermia conditions that could potentially lead to development of better targeted and personalized strategies for patients undergoing combined thermotherapy with chemotherapy. PMID:28342898

Temperature induces significant changes in both glycolytic reserve and mitochondrial spare respiratory capacity in colorectal cancer cell lines.

PubMed

Mitov, Mihail I; Harris, Jennifer W; Alstott, Michael C; Zaytseva, Yekaterina Y; Evers, B Mark; Butterfield, D Allan

2017-05-15

Thermotherapy, as a method of treating cancer, has recently attracted considerable attention from basic and clinical investigators. A number of studies and clinical trials have shown that thermotherapy can be successfully used as a therapeutic approach for various cancers. However, the effects of temperature on cancer bioenergetics have not been studied in detail with a real time, microplate based, label-free detection approach. This study investigates how changes in temperature affect the bioenergetics characteristics (mitochondrial function and glycolysis) of three colorectal cancer (CRC) cell lines utilizing the Seahorse XF96 technology. Experiments were performed at 32°C, 37°C and 42°C using assay medium conditions and equipment settings adjusted to produce equal oxygen and pH levels ubiquitously at the beginning of all experiments. The results suggest that temperature significantly changes multiple components of glycolytic and mitochondrial function of all cell lines tested. Under hypothermia conditions (32°C), the extracellular acidification rates (ECAR) of CRC cells were significantly lower compared to the same basal ECAR levels measured at 37°C. Mitochondrial stress test for SW480 cells at 37°C vs 42°C demonstrated increased proton leak while all other OCR components remained unchanged (similar results were detected also for the patient-derived xenograft cells Pt.93). Interestingly, the FCCP dose response at 37°C vs 42°C show significant shifts in profiles, suggesting that single dose FCCP experiments might not be sufficient to characterize the mitochondrial metabolic potential when comparing groups, conditions or treatments. These findings provide valuable insights for the metabolic and bioenergetic changes of CRC cells under hypo- and hyperthermia conditions that could potentially lead to development of better targeted and personalized strategies for patients undergoing combined thermotherapy with chemotherapy. Published by Elsevier Inc.

Temperature induces significant changes in both glycolytic reserve and mitochondrial spare respiratory capacity in colorectal cancer cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mitov, Mihail I., E-mail: m.mitov@uky.edu; Harris, Jennifer W.; Alstott, Michael C.

Thermotherapy, as a method of treating cancer, has recently attracted considerable attention from basic and clinical investigators. A number of studies and clinical trials have shown that thermotherapy can be successfully used as a therapeutic approach for various cancers. However, the effects of temperature on cancer bioenergetics have not been studied in detail with a real time, microplate based, label-free detection approach. This study investigates how changes in temperature affect the bioenergetics characteristics (mitochondrial function and glycolysis) of three colorectal cancer (CRC) cell lines utilizing the Seahorse XF96 technology. Experiments were performed at 32 °C, 37 °C and 42 °Cmore » using assay medium conditions and equipment settings adjusted to produce equal oxygen and pH levels ubiquitously at the beginning of all experiments. The results suggest that temperature significantly changes multiple components of glycolytic and mitochondrial function of all cell lines tested. Under hypothermia conditions (32 °C), the extracellular acidification rates (ECAR) of CRC cells were significantly lower compared to the same basal ECAR levels measured at 37 °C. Mitochondrial stress test for SW480 cells at 37 °C vs 42 °C demonstrated increased proton leak while all other OCR components remained unchanged (similar results were detected also for the patient-derived xenograft cells Pt.93). Interestingly, the FCCP dose response at 37 °C vs 42 °C show significant shifts in profiles, suggesting that single dose FCCP experiments might not be sufficient to characterize the mitochondrial metabolic potential when comparing groups, conditions or treatments. These findings provide valuable insights for the metabolic and bioenergetic changes of CRC cells under hypo- and hyperthermia conditions that could potentially lead to development of better targeted and personalized strategies for patients undergoing combined thermotherapy with chemotherapy.« less

Synergistic Effect of Simvastatin Plus Radiation in Gastric Cancer and Colorectal Cancer: Implications of BIRC5 and Connective Tissue Growth Factor.

PubMed

Lim, Taekyu; Lee, Inkyoung; Kim, Jungmin; Kang, Won Ki

2015-10-01

We investigated the synergistic effect of simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor plus radiation therapy, on the proliferation and survival of gastric cancer (GC) and colorectal cancer (CRC) cells. We also studied several genes involved in the simvastatin/radiation-induced effects. Gastric cancer (AGS, SNU601, MKN1, and MKN28) and CRC (CoLo320, SW48, HT29, and HCT8) cell lines were treated with 0.2 μM simvastatin alone, or in combination with 0 to 4 Gy of radiation, and subjected to clonogenic survival and proliferation assays in vitro. To assess the molecular mechanism of the combination treatment, we performed microarray analysis, immunoblot assays, small interfering RNA knockdown experiments, and plasmid rescue assays. The antitumoral effects of simvastatin and radiation were evaluated in vivo using xenograft models. The combination therapy of simvastatin plus radiation inhibited basal clonogenic survival and proliferation of GC and CRC cells in vitro. Simvastatin suppressed the expression of BIRC5 and CTGF genes in these cancer cells. In vivo, the combined treatment with simvastatin and radiation significantly reduced the growth of xenograft tumors compared with treatment with radiation alone. We suggest that simvastatin has a synergistic effect with radiation on GC and CRC through the induction of apoptosis, which may be mediated by a simultaneous inhibition of BIRC5 and CTGF expression. A clinical trial of simvastatin in combination with radiation in patients with GC or CRC is warranted. Copyright © 2015 Elsevier Inc. All rights reserved.

Circulating tumour cells from patients with colorectal cancer have cancer stem cell hallmarks in ex vivo culture.

PubMed

Grillet, Fanny; Bayet, Elsa; Villeronce, Olivia; Zappia, Luke; Lagerqvist, Ebba Louise; Lunke, Sebastian; Charafe-Jauffret, Emmanuelle; Pham, Kym; Molck, Christina; Rolland, Nathalie; Bourgaux, Jean François; Prudhomme, Michel; Philippe, Claire; Bravo, Sophie; Boyer, Jean Christophe; Canterel-Thouennon, Lucile; Taylor, Graham Roy; Hsu, Arthur; Pascussi, Jean Marc; Hollande, Frédéric; Pannequin, Julie

2017-10-01

Although counting of circulating tumour cells (CTC) has attracted a broad interest as potential markers of tumour progression and treatment response, the lack of functional characterisation of these cells had become a bottleneck in taking these observations to the clinic. Our objective was to culture these cells in order to understand them and exploit their therapeutic potential to the full. Here, hypothesising that some CTC potentially have cancer stem cell (CSC) phenotype, we generated several CTC lines from the blood of patients with advanced metastatic colorectal cancer (CRC) based on their self-renewal abilities. Multiple standard tests were then employed to characterise these cells. Our CTC lines self-renew, express CSC markers and have multilineage differentiation ability, both in vitro and in vivo . Patient-derived CTC lines are tumorigenic in subcutaneous xenografts and are also able to colonise the liver after intrasplenic injection. RNA sequencing analyses strikingly demonstrate that drug metabolising pathways represent the most upregulated feature among CTC lines in comparison with primary CRC cells grown under similar conditions. This result is corroborated by the high resistance of the CTC lines to conventional cytotoxic compounds. Taken together, our results directly demonstrate the existence of patient-derived colorectal CTCs that bear all the functional attributes of CSCs. The CTC culture model described here is simple and takes <1 month from blood collection to drug testing, therefore, routine clinical application could facilitate access to personalised medicine. ClinicalTrial.gov NCT01577511. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

Xc- inhibitor sulfasalazine sensitizes colorectal cancer to cisplatin by a GSH-dependent mechanism.

PubMed

Ma, Ming-zhe; Chen, Gang; Wang, Peng; Lu, Wen-hua; Zhu, Chao-feng; Song, Ming; Yang, Jing; Wen, Shijun; Xu, Rui-hua; Hu, Yumin; Huang, Peng

2015-11-01

Sulfasalazine (SSZ) is an anti-inflammatory drug that has been demonstrated to induce apoptosis and tumor regression through inhibition of plasma membrane cystine transporter xc(-). Cysteine is a rate-limiting precursor for intracellular glutathione (GSH) synthesis, which is vital for compound detoxification and maintaining redox balance. Platinum-based chemotherapy is an important regimen used in clinics for various cancers including colorectal cancer (CRC). We hypothesized that targeting xc(-) transporter by SSZ may annihilate cellular detoxification through interruption of GSH synthesis and may enhance the anti-cancer activity of cisplatin (CDDP) by increasing drug transport. In the present study, we revealed that xCT, the active subunit of xc(-), is highly expressed in CRC cell lines and human colorectal carcinoma tissues compared with their normal counterparts. SSZ effectively depleted cellular GSH, leading to significant accumulation of reactive oxygen species and growth inhibition in CRC cells. In contrast, the normal epithelial cells of colon origin were less sensitive to SSZ, showing a moderate ROS elevation. Importantly, SSZ effectively enhanced the intracellular platinum level and cytotoxicity of CDDP in CRC cells. The synergistic effect of SSZ and CDDP was reversed by antioxidant N-acetyl-L-cysteine (NAC). Together, these results suggest that SSZ, a relatively non-toxic drug that targets cystine transporter, may, in combination with CDDP, have effective therapy for colorectal cancer. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Colon cancer chemopreventive effects of baicalein, an active enteric microbiome metabolite from baicalin

PubMed Central

WANG, CHONG-ZHI; ZHANG, CHUN-FENG; CHEN, LINA; ANDERSON, SAMANTHA; LU, FANG; YUAN, CHUN-SU

2015-01-01

Baicalin is a major constituent of Scutellaria baicalensis, which is a commonly used herbal medicine in many Asian countries. After oral ingestion, intestinal micro-biota metabolism may change parent compound's structure and its biological activities. However, whether baicalin can be metabolized by enteric microbiota and the related anti-cancer activity is not clear. In this study, using human enteric microbiome incubation and HPLC analysis, we observed that baicalin can be quickly converted to baicalein. We compared the antiproliferative effects of baicalin and baicalein using a panel of human cancer cell lines, including three human colorectal cancer (CRC) cell lines. In vitro antiproliferative effects on CRC cells were verified using an in vivo xenograft nude mouse model. Baicalin showed limited antiproliferative effects on some of these cancer cell lines. Baicalein, however, showed significant antiproliferative effects in all the tested cancer cell lines, especially on HCT-116 human colorectal cancer cells. In vivo antitumor results supported our in vitro data. We demonstrated that baicalein exerts potent S phase cell cycle arrest and pro-apoptotic effects in HCT-116 cells. Baicalein induced the activation of caspase 3 and 9. The in silico modeling suggested that baicalein forms hydrogen bonds with residues Ser251 and Asp253 at the active site of caspase 3, while interactions with residues Leu227 and Asp228 in caspase 9 through its hydroxyl groups. Data from this study suggested that baicalein is a potent anticancer metabolite derived from S. baicalensis. Enteric microbiota play a key role in the colon cancer chemoprevention of S. baicalensis. PMID:26398706

Conditionally reprogrammed cells (CRC) methodology does not allow the in vitro expansion of patient-derived primary and metastatic lung cancer cells.

PubMed

Sette, Giovanni; Salvati, Valentina; Giordani, Ilenia; Pilozzi, Emanuela; Quacquarini, Denise; Duranti, Enrico; De Nicola, Francesca; Pallocca, Matteo; Fanciulli, Maurizio; Falchi, Mario; Pallini, Roberto; De Maria, Ruggero; Eramo, Adriana

2018-07-01

Availability of tumor and non-tumor patient-derived models would promote the development of more effective therapeutics for non-small cell lung cancer (NSCLC). Recently, conditionally reprogrammed cells (CRC) methodology demonstrated exceptional potential for the expansion of epithelial cells from patient tissues. However, the possibility to expand patient-derived lung cancer cells using CRC protocols is controversial. Here, we used CRC approach to expand cells from non-tumoral and tumor biopsies of patients with primary or metastatic NSCLC as well as pulmonary metastases of colorectal or breast cancers. CRC cultures were obtained from both tumor and non-malignant tissues with extraordinary high efficiency. Tumor cells were tracked in vitro through tumorigenicity assay, monitoring of tumor-specific genetic alterations and marker expression. Cultures were composed of EpCAM+ lung epithelial cells lacking tumorigenic potential. NSCLC biopsies-derived cultures rapidly lost patient-specific genetic mutations or tumor antigens. Similarly, pulmonary metastases of colon or breast cancer generated CRC cultures of lung epithelial cells. All CRC cultures examined displayed epithelial lung stem cell phenotype and function. In contrast, brain metastatic lung cancer biopsies failed to generate CRC cultures. In conclusion, patient-derived primary and metastatic lung cancer cells were negatively selected under CRC conditions, limiting the expansion to non-malignant lung epithelial stem cells from either tumor or non-tumor tissue sources. Thus, CRC approach cannot be applied for direct therapeutic testing of patient lung tumor cells, as the tumor-derived CRC cultures are composed of (non-tumoral) airway basal cells. © 2018 UICC.

Characterization of acetate transport in colorectal cancer cells and potential therapeutic implications

PubMed Central

Ferro, Suellen; Azevedo-Silva, João; Casal, Margarida; Côrte-Real, Manuela; Baltazar, Fatima; Preto, Ana

2016-01-01

Acetate, together with other short chain fatty acids has been implicated in colorectal cancer (CRC) prevention/therapy. Acetate was shown to induce apoptosis in CRC cells. The precise mechanism underlying acetate transport across CRC cells membrane, that may be implicated in its selectivity towards CRC cells, is not fully understood and was addressed here. We also assessed the effect of acetate in CRC glycolytic metabolism and explored its use in combination with the glycolytic inhibitor 3-bromopyruvate (3BP). We provide evidence that acetate enters CRC cells by the secondary active transporters MCT1 and/or MCT2 and SMCT1 as well as by facilitated diffusion via aquaporins. CRC cell exposure to acetate upregulates the expression of MCT1, MCT4 and CD147, while promoting MCT1 plasma membrane localization. We also observed that acetate increases CRC cell glycolytic phenotype and that acetate-induced apoptosis and anti-proliferative effect was potentiated by 3BP. Our data suggest that acetate selectivity towards CRC cells might be explained by the fact that aquaporins and MCTs are found overexpressed in CRC clinical cases. Our work highlights the importance that acetate transport regulation has in the use of drugs such as 3BP as a new therapeutic strategy for CRC. PMID:28874966

Molecular mechanisms underlying the antitumor activity of (E)-N-hydroxy-3-(1-(4-methoxyphenylsulfonyl)-1,2,3,4-tetrahydroquinolin-6-yl)acrylamide in human colorectal cancer cells in vitro and in vivo

PubMed Central

Chen, Chun-Han; Lee, Chia-Hwa; Liou, Jing-Ping; Teng, Che-Ming; Pan, Shiow-Lin

2015-01-01

Upregulation of class I histone deacetylases (HDAC) correlates with poor prognosis in colorectal cancer (CRC) patients. Previous study revealed that (E)-N-hydroxy-3-(1-(4-methoxyphenylsulfonyl)-1,2,3,4-tetrahydroquinolin-6-yl)acrylamide (Compound 11) is a potent and selective class I HDAC inhibitor, exhibited significant anti-proliferative activity in various human cancer cell lines. In current study, we demonstrated that compound 11 exhibited significant anti-proliferative and cytotoxic activity in CRC cells. Notably, compound 11 was less potent than SAHA in inhibiting HDAC6 as evident from the lower expression of acetyl-α-tubulin, suggesting higher selectivity for class I HDACs. Mechanistically, compound 11 induced cell-cycle arrest at the G2/M phase, activated both intrinsic- and extrinsic-apoptotic pathways, altered the expression of Bcl-2 family proteins and exerted a potent inhibitory effect on survival signals (p-Akt, p-ERK) in CRC cells. Moreover, we provide evidence that compound 11 suppressed motility, decreased mesenchymal markers (N-cadherin and vimentin) and increased epithelial marker (E-cadherin) through down-regulation of Akt. The anti-tumor activity and underlying molecular mechanisms of compound 11 were further confirmed using the HCT116 xenograft model in vivo. Our findings provide evidence of the significant anti-tumor activity of compound 11 in a preclinical model, supporting its potential as a novel therapeutic agent for CRC. PMID:26462017

BDE-99 (2,2',4,4',5-pentabromodiphenyl ether) triggers epithelial-mesenchymal transition in colorectal cancer cells via PI3K/Akt/Snail signaling pathway.

PubMed

Wang, Fei; Ruan, Xin-Jian; Zhang, Hong-Yan

2015-01-01

The gut is in direct contact with BDE-99 (2,2',4,4',5-pentabromodiphenyl ether), one of the most abundant PBDE congeners in the environment and in human tissues. The objective of the present study was to investigate the effects of BDE-99 on colorectal cancer (CRC) cells. The effects of BDE-99 on cell proliferation were measured by CCK-8 assay in the CRC cell line HCT-116. Wound healing and transwell migration/invasion assays were used to test the migration and invasion of CRC cells. Factors related to epithelial-to-mesenchymal transition (EMT) were measured by real-time PCR and Western blot analysis for mRNA and protein levels, respectively. BDE-99 was found to increase migration and invasion and trigger EMT in HCT-116 cells; EMT was characterized by cells acquiring mesenchymal spindle-like morphology and by increased expression of N-cadherin with a concomitant decrease in E-cadherin. BDE-99 treatment also increased the protein and mRNA levels of the transcription factor Snail, but not Slug, Twist, and ZEB1. Knockdown of Snail by siRNA significantly attenuated BDE-99-induced EMT in HCT-116 cells, suggesting that Snail plays a crucial role in BDE-99-induced EMT. The PI3K/Akt inhibitor LY294002 completely blocked BDE-99-induced Snail and invasion of HCT-116 cells. Our results revealed that BDE-99 can trigger the EMT of colon cancer cells via the PI3K/AKT/Snail signaling pathway. This study provides new insight into the tumorigenesis and metastasis of CRC stimulated by BDE-99 and possibly other PBDE congeners.

Effect of aspirin on tumour cell colony formation and evolution.

PubMed

Wodarz, Dominik; Goel, Ajay; Boland, C Richard; Komarova, Natalia L

2017-09-01

Aspirin is known to reduce the risk of colorectal cancer (CRC) incidence, but the underlying mechanisms are not fully understood. In a previous study, we quantified the in vitro growth kinetics of different CRC tumour cell lines treated with varying doses of aspirin, measuring the rate of cell division and cell death. Here, we use these measured parameters to calculate the chances of successful clonal expansion and to determine the evolutionary potential of the tumour cell lines in the presence and absence of aspirin. The calculations indicate that aspirin increases the probability that a single tumour cell fails to clonally expand. Further, calculations suggest that aspirin increases the evolutionary potential of an expanding tumour cell colony. An aspirin-treated tumour cell population is predicted to result in the accumulation of more mutations (and is thus more virulent and more difficult to treat) than a cell population of the same size that grew without aspirin. This indicates a potential trade-off between delaying the onset of cancer and increasing its evolutionary potential through chemoprevention. Further work needs to investigate to what extent these findings apply to in vivo settings, and to what degree they contribute to the epidemiologically documented aspirin-mediated protection. © 2017 The Author(s).

MUC4 is negatively regulated through the Wnt/β-catenin pathway via the Notch effector Hath1 in colorectal cancer

PubMed Central

Pai, Priya; Rachagani, Satyanarayana; Dhawan, Punita; Sheinin, Yuri M.; Mallya, Kavita; Pothuraju, Ramesh; Batra, Surinder K.

2016-01-01

MUC4 is a transmembrane mucin lining the normal colonic epithelium. The aberrant/de novo over-expression of MUC4 is well documented in malignancies of the pancreas, ovary and breast. However, studies have reported the loss of MUC4 expression in the majority of colorectal cancers (CRCs). A MUC4 promoter analysis showed the presence of three putative TCF/LEF sites, implying a possible regulation by the Wnt/β-catenin pathway, which has been shown to drive CRC progression. Thus, the objective of our study was to determine whether MUC4 is regulated by β-catenin in CRC. We first knocked down (KD) β-catenin in three CRC cell lines; LS180, HCT-8 and HCT116, which resulted in increased MUC4 transcript and MUC4 protein. Additionally, the overexpression of stabilized mutant β-catenin in LS180 and HCT-8 resulted in a decrease in MUC4 expression. Immunohistochemistry (IHC) of mouse colon tissue harboring tubular adenomas and high grade dysplasia showed dramatically reduced Muc4 in lesions relative to adjacent normal tissue, with increased cytosolic/nuclear β-catenin. Luciferase assays with the complete MUC4 promoter construct p3778 showed increased MUC4 promoter luciferase activity in the absence of β-catenin (KD). Mutation of all three putative TCF/LEF sites showed that MUC4 promoter luciferase activity was increased relative to the un-mutated promoter. Interestingly, it was observed that MUC4 expressing CRC cell lines also expressed high levels of Hath1, a transcription factor repressed by both active Wnt/β-catenin and Notch signaling. The KD of β-catenin and/or treatment with a Notch γ-secretase inhibitor, Dibenzazepine (DBZ) resulted in increased Hath1 and MUC4 in LS180, HCT-8 and HCT116. Furthermore, overexpression of Hath1 in HCT-8 and LS180 caused increased MUC4 transcript and MUC4 protein. Taken together, our results indicate that the Wnt/β-catenin pathway suppresses the Notch pathway effector Hath1, resulting in reduced MUC4 in CRC. PMID:27551331

MUC4 is negatively regulated through the Wnt/β-catenin pathway via the Notch effector Hath1 in colorectal cancer.

PubMed

Pai, Priya; Rachagani, Satyanarayana; Dhawan, Punita; Sheinin, Yuri M; Macha, Muzafar A; Qazi, Asif Khurshid; Chugh, Seema; Ponnusamy, Moorthy P; Mallya, Kavita; Pothuraju, Ramesh; Batra, Surinder K

2016-05-01

MUC4 is a transmembrane mucin lining the normal colonic epithelium. The aberrant/de novo over-expression of MUC4 is well documented in malignancies of the pancreas, ovary and breast. However, studies have reported the loss of MUC4 expression in the majority of colorectal cancers (CRCs). A MUC4 promoter analysis showed the presence of three putative TCF/LEF sites, implying a possible regulation by the Wnt/β-catenin pathway, which has been shown to drive CRC progression. Thus, the objective of our study was to determine whether MUC4 is regulated by β-catenin in CRC. We first knocked down (KD) β-catenin in three CRC cell lines; LS180, HCT-8 and HCT116, which resulted in increased MUC4 transcript and MUC4 protein. Additionally, the overexpression of stabilized mutant β-catenin in LS180 and HCT-8 resulted in a decrease in MUC4 expression. Immunohistochemistry (IHC) of mouse colon tissue harboring tubular adenomas and high grade dysplasia showed dramatically reduced Muc4 in lesions relative to adjacent normal tissue, with increased cytosolic/nuclear β-catenin. Luciferase assays with the complete MUC4 promoter construct p3778 showed increased MUC4 promoter luciferase activity in the absence of β-catenin (KD). Mutation of all three putative TCF/LEF sites showed that MUC4 promoter luciferase activity was increased relative to the un-mutated promoter. Interestingly, it was observed that MUC4 expressing CRC cell lines also expressed high levels of Hath1, a transcription factor repressed by both active Wnt/β-catenin and Notch signaling. The KD of β-catenin and/or treatment with a Notch γ-secretase inhibitor, Dibenzazepine (DBZ) resulted in increased Hath1 and MUC4 in LS180, HCT-8 and HCT116. Furthermore, overexpression of Hath1 in HCT-8 and LS180 caused increased MUC4 transcript and MUC4 protein. Taken together, our results indicate that the Wnt/β-catenin pathway suppresses the Notch pathway effector Hath1, resulting in reduced MUC4 in CRC.

5-ASA Affects Cell Cycle Progression in Colorectal Cells by Reversibly Activating a Replication Checkpoint

PubMed Central

LUCIANI, M. GLORIA; CAMPREGHER, CHRISTOPH; FORTUNE, JOHN M.; KUNKEL, THOMAS A.; GASCHE, CHRISTOPH

2007-01-01

Background & Aims Individuals with inflammatory bowel disease are at risk of developing colorectal cancer (CRC). Epidemiologic, animal, and laboratory studies suggest that 5-amino-salicylic acid (5-ASA) protects from the development of CRC by altering cell cycle progression and by inducing apoptosis. Our previous results indicate that 5-ASA improves replication fidelity in colorectal cells, an effect that is active in reducing mutations. In this study, we hypothesized that 5-ASA restrains cell cycle progression by activating checkpoint pathways in colorectal cell lines, which would prevent tumor development and improve genomic stability. Methods CRC cells with different genetic backgrounds such as HT29, HCT116, HCT116p53−/−, HCT116+chr3, and LoVo were treated with 5-ASA for 2–96 hours. Cell cycle progression, phosphorylation, and DNA binding of cell cycle checkpoint proteins were analyzed. Results We found that 5-ASA at concentrations between 10 and 40 mmol/L affects cell cycle progression by inducing cells to accumulate in the S phase. This effect was independent of the hMLH1, hMSH2, and p53 status because it was observed to a similar extent in all cell lines under investigation. Moreover, wash-out experiments demonstrated reversibility within 48 hours. Although p53 did not have a causative role, p53 Ser15 was strongly phosphorylated. Proteins involved in the ATM-and-Rad3-related kinase (ATR)-dependent S-phase checkpoint response (Chk1 and Rad17) were also phosphorylated but not ataxia telengectasia mutated kinase. Conclusions Our data demonstrate that 5-ASA causes cells to reversibly accumulate in S phase and activate an ATR-dependent checkpoint. The activation of replication checkpoint may slow down DNA replication and improve DNA replication fidelity, which increases the maintenance of genomic stability and counteracts carcinogenesis. PMID:17241873

Gasdermin C Is Upregulated by Inactivation of Transforming Growth Factor β Receptor Type II in the Presence of Mutated Apc, Promoting Colorectal Cancer Proliferation.

PubMed

Miguchi, Masashi; Hinoi, Takao; Shimomura, Manabu; Adachi, Tomohiro; Saito, Yasufumi; Niitsu, Hiroaki; Kochi, Masatoshi; Sada, Haruki; Sotomaru, Yusuke; Ikenoue, Tsuneo; Shigeyasu, Kunitoshi; Tanakaya, Kohji; Kitadai, Yasuhiko; Sentani, Kazuhiro; Oue, Naohide; Yasui, Wataru; Ohdan, Hideki

2016-01-01

Mutations in TGFBR2, a component of the transforming growth factor (TGF)-β signaling pathway, occur in high-frequency microsatellite instability (MSI-H) colorectal cancer (CRC). In mouse models, Tgfbr2 inactivation in the intestinal epithelium accelerates the development of malignant intestinal tumors in combination with disruption of the Wnt-β-catenin pathway. However, no studies have further identified the genes influenced by TGFBR2 inactivation following disruption of the Wnt-β-catenin pathway. We previously described CDX2P-G19Cre;Apcflox/flox mice, which is stochastically null for Apc in the colon epithelium. In this study, we generated CDX2P-G19Cre;Apcflox/flox;Tgfbr2flox/flox mice, with simultaneous loss of Apc and Tgfbr2. These mice developed tumors, including adenocarcinoma in the proximal colon. We compared gene expression profiles between tumors of the two types of mice using microarray analysis. Our results showed that the expression of the murine homolog of GSDMC was significantly upregulated by 9.25-fold in tumors of CDX2P-G19Cre;Apcflox/flox;Tgfbr2flox/flox mice compared with those of CDX2P-G19Cre;Apcflox/flox mice. We then investigated the role of GSDMC in regulating CRC tumorigenesis. The silencing of GSDMC led to a significant reduction in the proliferation and tumorigenesis of CRC cell lines, whereas the overexpression of GSDMC enhanced cell proliferation. These results suggested that GSDMC functioned as an oncogene, promoting cell proliferation in colorectal carcinogenesis. In conclusion, combined inactivation of both Apc and Tgfbr2 in the colon epithelium of a CRC mouse model promoted development of adenocarcinoma in the proximal colon. Moreover, GSDMC was upregulated by TGFBR2 mutation in CRC and promoted tumor cell proliferation in CRC carcinogenesis, suggesting that GSDMC may be a promising therapeutic target.

Effect of DNA methylation profile on OATP3A1 and OATP4A1 transcript levels in colorectal cancer.

PubMed

Rawłuszko-Wieczorek, Agnieszka Anna; Horst, Nikodem; Horbacka, Karolina; Bandura, Artur Szymon; Świderska, Monika; Krokowicz, Piotr; Jagodziński, Paweł Piotr

2015-08-01

Epidemiological studies indicate that 17β-estradiol (E2) prevents colorectal cancer (CRC). Organic anion transporting polypeptides (OATPs) are involved in the cellular uptake of various endogenous and exogenous substrates, including hormone conjugates. Because transfer of estrone sulfate (E1-S) can contribute to intra-tissue conversion of estrone to the biologically active form -E2, it is evident that the expression patterns of OATPs may be relevant to the analysis of CRC incidence and therapy. We therefore evaluated DNA methylation and transcript levels of two members of the OATP family, OATP3A1 and OATP4A1, that may be involved in E1-S transport in colorectal cancer patients. We detected a significant reduction in OATP3A1 and a significant increase in OATP4A1 mRNA levels in cancerous tissue, compared with histopathologically unchanged tissue (n=103). Moreover, we observed DNA hypermethylation in the OATP3A1 promoter region in a small subset of CRC patients and in HCT116 and Caco-2 colorectal cancer cell lines. We also observed increased OATP3A1 transcript following treatment with 5-aza-2-deoxycytidine and sodium butyrate. The OATP4A1 promoter region was hypomethylated in analyzed tissues and CRC cell lines and was not affected by these treatments. Our results suggest a potential mechanism for OATP3A1 downregulation that involves DNA methylation during colorectal carcinogenesis. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Synergistic Effect of Simvastatin Plus Radiation in Gastric Cancer and Colorectal Cancer: Implications of BIRC5 and Connective Tissue Growth Factor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lim, Taekyu; Lee, Inkyoung; Kim, Jungmin

Purpose: We investigated the synergistic effect of simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor plus radiation therapy, on the proliferation and survival of gastric cancer (GC) and colorectal cancer (CRC) cells. We also studied several genes involved in the simvastatin/radiation-induced effects. Methods and Materials: Gastric cancer (AGS, SNU601, MKN1, and MKN28) and CRC (CoLo320, SW48, HT29, and HCT8) cell lines were treated with 0.2 μM simvastatin alone, or in combination with 0 to 4 Gy of radiation, and subjected to clonogenic survival and proliferation assays in vitro. To assess the molecular mechanism of the combination treatment, we performed microarray analysis, immunoblot assays, small interferingmore » RNA knockdown experiments, and plasmid rescue assays. The antitumoral effects of simvastatin and radiation were evaluated in vivo using xenograft models. Results: The combination therapy of simvastatin plus radiation inhibited basal clonogenic survival and proliferation of GC and CRC cells in vitro. Simvastatin suppressed the expression of BIRC5 and CTGF genes in these cancer cells. In vivo, the combined treatment with simvastatin and radiation significantly reduced the growth of xenograft tumors compared with treatment with radiation alone. Conclusion: We suggest that simvastatin has a synergistic effect with radiation on GC and CRC through the induction of apoptosis, which may be mediated by a simultaneous inhibition of BIRC5 and CTGF expression. A clinical trial of simvastatin in combination with radiation in patients with GC or CRC is warranted.« less

Fructose-bisphosphate aldolase A is a key regulator of hypoxic adaptation in colorectal cancer cells and involved in treatment resistance and poor prognosis.

PubMed

Kawai, Kenji; Uemura, Mamoru; Munakata, Koji; Takahashi, Hidekazu; Haraguchi, Naotsugu; Nishimura, Junichi; Hata, Taishi; Matsuda, Chu; Ikenaga, Masakazu; Murata, Kohei; Mizushima, Tsunekazu; Yamamoto, Hirofumi; Doki, Yuichiro; Mori, Masaki

2017-02-01

Hypoxia is an essential feature of cancer malignancy, but there are no methods for the routine detection of hypoxia-inducible prognostic factors and potential therapeutic targets. We reported previously that the hypoxic tumor cells of metastatic liver tissue from patients with colorectal cancer (CRC) could be used as an 'in vivo' hypoxia culture model. Several potential hypoxia-inducible genes were identified using this model. Among them, one glycolytic enzyme was of special interest. There is currently increasing attention on glycolytic enzymes as potential therapeutic targets due to their association with cancer-specific metabolism. To better understand the molecular mechanisms of cancer malignancy, we investigated the expression of fructose-bisphosphate aldolase A (ALDOA) and its relationship with cancer metabolism. We found that ALDOA was induced by hypoxia in CRC-derived cell lines, and univariate and multivariate analyses of microarray data from the resected CRC samples of 222 patients revealed that ALDOA was an independent prognostic factor for CRC. We also analyzed the malignant potential of ALDOA in vitro using overexpression and knockdown assays. We found that ALDOA was negatively related to chemosensitivity and radiosensitivity and positively associated with proliferation, sphere formation and invasion in both normoxia and hypoxia. These associations were due to the roles of ALDOA in regulating glycolysis, the epithelial-mesenchymal transition and the cell cycle. These findings demonstrate that ALDOA is a hypoxia-inducible prognostic factor that is closely related to CRC malignancy, and also provide new insights into the importance of ALDOA and glycolysis in cancer and suggest new targets for anticancer therapies.

The function of BTG3 in colorectal cancer cells and its possible signaling pathway.

PubMed

Lv, Chi; Wang, Heling; Tong, Yuxin; Yin, Hongzhuan; Wang, Dalu; Yan, Zhaopeng; Liang, Yichao; Wu, Di; Su, Qi

2018-02-01

B-cell translocation gene 3 (BTG3) has been identified as a candidate driver gene for various cancers, but its specific role in colorectal cancer (CRC) is poorly understood. We aimed to investigate the relationship between expression of BTG3 and clinicopathological features and prognosis, as well as to explore the effects and the role of a possible BTG3 molecular mechanism on aggressive colorectal cancer behavior. BTG3 expression was assessed by immunohistochemistry (IHC) on specimens from 140 patients with CRC. The association of BTG3 expression with clinicopathological features was examined. To confirm the biological role of BTG3 in CRC, two CRC cell lines expressing BTG3 were used and BTG3 expression was knocked down by shRNA. CCK-8, cell cycle, apoptosis, migration, and invasion assays were performed. The influence of BTG3 knockdown was further investigated by genomic microarray to uncover the potential molecular mechanisms underlying BTG3-mediated CRC development and progression. BTG3 was downregulated in colorectal cancer tissues and positively correlated with pathological classification (p = 0.037), depth of invasion (p = 0.016), distant metastasis (p = 0.024), TNM stage (p = 0.007), and overall survival (OS) and disease-free survival (DFS). BTG3 knockdown promoted cell proliferation, migration, invasion, relieved G2 arrest, and inhibited apoptosis in HCT116 and LoVo cells. A genomic microarray analysis showed that numerous tumor-associated signaling pathways and oncogenes were altered by BTG3 knockdown. At the mRNA level, nine genes referred to the extracellular-regulated kinase/mitogen-activated protein kinase pathway were differentially expressed. Western blotting revealed that BTG3 knockdown upregulated PAK2, RPS6KA5, YWHAB, and signal transducer and activator of transcription (STAT)3 protein levels, but downregulated RAP1A, DUSP6, and STAT1 protein expression, which was consistent with the genomic microarray data. BTG3 expression might contribute to CRC carcinogenesis. BTG3 knockdown might strengthen the aggressive colorectal cancer behavior.

Vitamins K2, K3 and K5 exert antitumor effects on established colorectal cancer in mice by inducing apoptotic death of tumor cells.

PubMed

Ogawa, Mutsumi; Nakai, Seiji; Deguchi, Akihiro; Nonomura, Takako; Masaki, Tsutomu; Uchida, Naohito; Yoshiji, Hitoshi; Kuriyama, Shigeki

2007-08-01

Although a number of studies have shown that vitamin K possesses antitumor activities on various neoplastic cell lines, there are few reports demonstrating in vivo antitumor effects of vitamin K, and the antitumor effect on colorectal cancer (CRC) remains to be examined. Therefore, antitumor effects of vitamin K on CRC were examined both in vitro and in vivo. Vitamins K2, K3 and K5 suppressed the proliferation of colon 26 cells in a dose-dependent manner, while vitamin K1 did not. On flow cytometry, induction of apoptosis by vitamins K2, K3 and K5 was suggested by population in sub-G1 phase of the cell cycle. Hoechst 33342 staining and a two-color flow cytometric assay using fluorescein isothiocyanate-conjugated annexin V and propidium iodide confirmed that vitamins K2, K3 and K5 induced apoptotic death of colon 26 cells. Enzymatic activity of caspase-3 in colon 26 cells was significantly up-regulated by vitamins K2, K3 and K5. The pan-caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone, substantially prevented vitamin K-mediated apoptosis. In vivo study using syngeneic mice with subcutaneously established colon 26 tumors demonstrated that intravenous administration of vitamins K2, K3 and K5 significantly suppressed the tumor growth. The number of apoptotic tumor cells was significantly larger in the vitamin K-treated groups than in the control group. These results suggest that vitamins K2, K3 and K5 exerted effective antitumor effects on CRC in vitro and in vivo by inducing caspase-dependent apoptotic death of tumor cells, suggesting that these K vitamins may be promising agents for the treatment of patients with CRC.

B-cell translocation gene 3 overexpression inhibits proliferation and invasion of colorectal cancer SW480 cells via Wnt/β-catenin signaling pathway.

PubMed

Mao, D; Qiao, L; Lu, H; Feng, Y

2016-01-01

Increasing evidences have shown that B-cell translocation gene 3 (BTG3) inhibits metastasis of multiple cancer cells. However, the role of BTG3 in colorectal cancer (CRC) and its possible mechanism have not yet been reported. In our study, we evaluated BTG3 expression in several CRC cell lines. Then, pcDNA3.1-BTG3 was transfected into SW480 cells. We found that BTG3 was upregulated in SW480 cells after overexpression plasmid transfection. BTG3 overexpression significantly inhibited cell growth and decreased PCNA (proliferating cell nuclear antigen) and Ki67 levels. BTG3 overexpression markedly downregulated Cyclin D1 and Cyclin E1 levels, whereas elevated p27. Overexpression of BTG3 arrested the cell cycle at G1 phase, which was abrogated by p27 silencing. Furthermore, migration, invasion and EMT of SW480 cells were significantly suppressed by BTG3 overexpression. Further investigations showed the inhibition of Wnt/β-catenin signaling pathway. We then used GSK3β specific inhibitor SB-216763 to activate the Wnt/β-catenin signaling pathway. We found that Wnt/β-catenin signaling pathway activation reversed the effect of BTG3 overexpression on cell proliferation, cell cycle progression, invasion and EMT. In conclusion, BTG3 overexpression inhibited cell growth, induced cell cycle arrest and suppressed the metastasis of SW480 cells via the Wnt/β-catenin signaling pathway. BTG3 may be considered as a therapeutic target in CRC treatment.

Iro/IRX transcription factors negatively regulate Dpp/TGF-β pathway activity during intestinal tumorigenesis.

PubMed

Martorell, Òscar; Barriga, Francisco M; Merlos-Suárez, Anna; Stephan-Otto Attolini, Camille; Casanova, Jordi; Batlle, Eduard; Sancho, Elena; Casali, Andreu

2014-11-01

Activating mutations in Wnt and EGFR/Ras signaling pathways are common in colorectal cancer (CRC). Remarkably, clonal co-activation of these pathways in the adult Drosophila midgut induces "tumor-like" overgrowths. Here, we show that, in these clones and in CRC cell lines, Dpp/TGF-β acts as a tumor suppressor. Moreover, we discover that the Iroquois/IRX-family-protein Mirror downregulates the transcription of core components of the Dpp pathway, reducing its tumor suppressor activity. We also show that this genetic interaction is conserved in human CRC cells, where the Iro/IRX proteins IRX3 and IRX5 diminish the response to TGF-β. IRX3 and IRX5 are upregulated in human adenomas, and their levels correlate inversely with the gene expression signature of response to TGF-β. In addition, Irx5 expression confers a growth advantage in the presence of TGF-β, but is selected against in its absence. Together, our results identify a set of Iro/IRX proteins as conserved negative regulators of Dpp/TGF-β activity. We propose that during the characteristic adenoma-to-carcinoma transition of human CRC, the activity of IRX proteins could reduce the sensitivity to the cytostatic effect of TGF-β, conferring a growth advantage to tumor cells prior to the acquisition of mutations in TGF-β pathway components. © 2014 The Authors.

KRAS Mutation Is a Predictor of Oxaliplatin Sensitivity in Colon Cancer Cells

PubMed Central

Lin, Yu-Lin; Ou, Da-Liang; Lin, Liang-In; Tseng, Li-Hui; Chang, Yih-Leong; Yeh, Kun-Huei; Cheng, Ann-Lii

2012-01-01

Molecular biomarkers to determine the effectiveness of targeted therapies in cancer treatment have been widely adopted in colorectal cancer (CRC), but those to predict chemotherapy sensitivity remain poorly defined. We tested our hypothesis that KRAS mutation may be a predictor of oxaliplatin sensitivity in CRC. KRAS was knocked-down in KRAS-mutant CRC cells (DLD-1G13D and SW480G12V) by small interfering RNAs (siRNA) and overexpressed in KRAS-wild-type CRC cells (COLO320DM) by KRAS-mutant vectors to generate paired CRC cells. These paired CRC cells were tested by oxaliplatin, irinotecan and 5FU to determine the change in drug sensitivity by MTT assay and flow cytometry. Reasons for sensitivity alteration were further determined by western blot and real-time quantitative reverse transcriptase polymerase chain reaction (qRT -PCR). In KRAS-wild-type CRC cells (COLO320DM), KRAS overexpression by mutant vectors caused excision repair cross-complementation group 1 (ERCC1) downregulation in protein and mRNA levels, and enhanced oxaliplatin sensitivity. In contrast, in KRAS-mutant CRC cells (DLD-1G13D and SW480G12V), KRAS knocked-down by KRAS-siRNA led to ERCC1 upregulation and increased oxaliplatin resistance. The sensitivity of irinotecan and 5FU had not changed in the paired CRC cells. To validate ERCC1 as a predictor of sensitivity for oxaliplatin, ERCC1 was knocked-down by siRNA in KRAS-wild-type CRC cells, which restored oxaliplatin sensitivity. In contrast, ERCC1 was overexpressed by ERCC1-expressing vectors in KRAS-mutant CRC cells, and caused oxaliplatin resistance. Overall, our findings suggest that KRAS mutation is a predictor of oxaliplatin sensitivity in colon cancer cells by the mechanism of ERCC1 downregulation. PMID:23209813

Multiple effects of the special AT-rich binding protein 1 (SATB1) in colon carcinoma.

PubMed

Frömberg, Anja; Rabe, Michael; Aigner, Achim

2014-12-01

SATB1 (special AT-rich binding protein 1) is a global chromatin organizer regulating the expression of a large number of genes. Overexpression has been found in various solid tumors and positively correlated with prognostic and clinicopathological properties. In colorectal cancer (CRC), SATB1 overexpression and its correlation with poor differentiation, invasive depth, TNM (tumor, nodes, metastases) stage and prognosis have been demonstrated. However, more detailed studies on the SATB1 functions in CRC are warranted. In this article, we comprehensively analyze the cellular and molecular role of SATB1 in CRC cell lines with different SATB1 expression levels by using RNAi-mediated knockdown. Using siRNAs with different knockdown efficacies, we demonstrate antiproliferative, cell cycle-inhibitory and proapoptotic effects of SATB1 knockdown in a SATB1 gene dose-dependent manner. Tumor growth inhibition is confirmed in vivo in a subcutaneous tumor xenograft mouse model using stable knockdown cells. The in-depth analysis of cellular effects reveals increased activities of caspases-3, -7, -8, -9 and other mediators of apoptotic pathways. Similarly, the analysis of E- and N-cadherin, slug, twist, β-catenin and MMP7 indicates SATB1 effects on epithelial-mesenchymal transition (EMT) and matrix breakdown. Our results also establish SATB1 effects on receptor tyrosine kinases and (proto-)oncogenes such as HER receptors and Pim-1. Taken together, this suggests a more complex molecular interplay between tumor-promoting and possible inhibitory effects in CRC by affecting multiple pathways and molecules involved in proliferation, cell cycle, EMT, invasion and cell survival. © 2014 UICC.

Alternative splicing of TIA-1 in human colon cancer regulates VEGF isoform expression, angiogenesis, tumour growth and bevacizumab resistance.

PubMed

Hamdollah Zadeh, Maryam A; Amin, Elianna M; Hoareau-Aveilla, Coralie; Domingo, Enric; Symonds, Kirsty E; Ye, Xi; Heesom, Katherine J; Salmon, Andrew; D'Silva, Olivia; Betteridge, Kai B; Williams, Ann C; Kerr, David J; Salmon, Andrew H J; Oltean, Sebastian; Midgley, Rachel S; Ladomery, Michael R; Harper, Steven J; Varey, Alexander H R; Bates, David O

2015-01-01

The angiogenic capability of colorectal carcinomas (CRC), and their susceptibility to anti-angiogenic therapy, is determined by expression of vascular endothelial growth factor (VEGF) isoforms. The intracellular protein T-cell Intracellular Antigen (TIA-1) alters post-transcriptional RNA processing and binds VEGF-A mRNA. We therefore tested the hypothesis that TIA-1 could regulate VEGF-A isoform expression in colorectal cancers. TIA-1 and VEGF-A isoform expression was measured in colorectal cancers and cell lines. We discovered that an endogenous splice variant of TIA-1 encoding a truncated protein, short TIA-1 (sTIA-1) was expressed in CRC tissues and invasive K-Ras mutant colon cancer cells and tissues but not in adenoma cell lines. sTIA-1 was more highly expressed in CRC than in normal tissues and increased with tumour stage. Knockdown of sTIA-1 or over-expression of full length TIA-1 (flTIA-1) induced expression of the anti-angiogenic VEGF isoform VEGF-A165b. Whereas flTIA-1 selectively bound VEGF-A165 mRNA and increased translation of VEGF-A165b, sTIA-1 prevented this binding. In nude mice, xenografted colon cancer cells over-expressing flTIA-1 formed smaller, less vascular tumours than those expressing sTIA-1, but flTIA-1 expression inhibited the effect of anti-VEGF antibodies. These results indicate that alternative splicing of an RNA binding protein can regulate isoform specific expression of VEGF providing an added layer of complexity to the angiogenic profile of colorectal cancer and their resistance to anti-angiogenic therapy. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Long non-coding RNA PlncRNA-1 promotes cell proliferation and hepatic metastasis in colorectal cancer.

PubMed

Jia, Gui-Qing; Zhang, Ming-Ming; Wang, Kang; Zhao, Gao-Ping; Pang, Ming-Hui; Chen, Zhe-Yu

2018-05-08

Emerging evidence has identified that long non-coding RNAs (lncRNAs) may play an important role in the pathogenesis of many cancer types, including colorectal cancer (CRC). However, the role of PlncRNA-1 in CRC remains unclear. The aim of our present study was to investigate the potential functions of PlncRNA-1 in CRC and to identify the underlying mechanisms of action. We demonstrated that up-regulated PlncRNA-1 in CRC tissues and cells promoted cell proliferation by accelerating cell cycle process and inhibiting cell apoptosis in vitro, enhanced tumor growth and matastasis in vivo and was associated with cell migration and invasion, EMT process of CRC cells. In addition, PlncRNA-1 was a target of miR-204 and enhanced the expression of an endogenous miR-204 target, MMP9 in CRC cells. Furthermore, we found that PlncRNA-1 activates Wnt/β-catenin pathway through the miR-204 in CRC cells. These results suggest that the PlncRNA-1/miR-204/ Wnt/β-catenin regulatory network may shed light on tumorigenesis in CRC. © 2018 Wiley Periodicals, Inc.

B7-H1 expression is associated with expansion of regulatory T cells in colorectal carcinoma

PubMed Central

Hua, Dong; Sun, Jing; Mao, Yong; Chen, Lu-Jun; Wu, Yu-Yu; Zhang, Xue-Guang

2012-01-01

AIM: To investigate the expression of B7-H1 in human colorectal carcinoma (CRC) to define its regulating effects on T cells in tumor microenvironment. METHODS: One hundred and two paraffin blocks and 33 fresh samples of CRC tissues were subject to this study. Immunohistochemistry was performed for B7-H1 and CD3 staining in CRC tissues. Ficoll-Hypaque density gradient centrifugation was used to isolate peripheral blood mononuclear cells of fresh CRC tissues; flow cytometry and immunofluorescence staining were used for detection of regulatory T cells. Data was analyzed with statistical software. RESULTS: Costimulatory molecule B7-H1 was found strongly expressed in CRC tissues, localized in tumor cell membrane and cytoplasm, while weak or none expression of B7-H1 was detected in pared normal colorectal tissues. Meanwhile, CD3 positive T cells were found congregated in CRC tumor nest and stroma. Statistic analysis showed that B7-H1 expression level was negatively correlated to the total T cell density in tumor nest (P < 0.0001) and tumor stroma (P = 0.0200) of 102 cases of CRC tissues. Among the total T cells, a variable amount of regulatory T cells with a clear Foxp3+ (forkhead box P3) staining could be detected in CRC tissues and patients’ blood. Interestingly, in the 33 samples (15 cases of B7-H1high CRC tissues and 18 cases of B7-H1low CRC tissues) of freshly isolated mononuclear cells from CRC tissues, the percentages of CD4+Foxp3+ and CD8+Foxp3+ regulatory T cells were found remarkably higher in B7-H1high CRC tissues than in B7-H1low CRC tissues (P = 0.0024, P = 0.0182), indicating that B7-H1 expression was involved in proliferation of regulatory T cell. No significant difference was found in CRC peripheral blood (P = 0.0863, P = 0.0678). PD-1 is the specific ligand for B7-H1 pathway transferring inhibitory signal to T cell, which is expressed by activated T cell. Our further analysis of PD-1 expression on T cells in CRC tissues showed that conventional T cells (CD4+Foxp3-/CD8+Foxp3-), which was thought to contribute to the anti-tumor immune response, highly expressed PD-1; while regulatory T cells (CD4+Foxp3+/CD8+Foxp3-) almost failed to express PD-1. The average percentage of PD-1 expression on regulatory T cells was significantly higher than the percentage of PD-1 on conventional T cells (CD4+Foxp3- T cell, P < 0.0001; CD8+Foxp3- T cell, P < 0.0001). The diverse expression of PD-1 might lead to different fate of T cell subsets in B7-H1 over-expression CRC tumor microenvironment. CONCLUSION: B7-H1 expression in tumor cells can inhibit the conventional T cell proliferation in tumor microenvironment through the PD-1 expression on conventional T cells. PMID:22408358

High early growth response 1 (EGR1) expression correlates with resistance to anti-EGFR treatment in vitro and with poorer outcome in metastatic colorectal cancer patients treated with cetuximab.

PubMed

Kumar, S S; Tomita, Y; Wrin, J; Bruhn, M; Swalling, A; Mohammed, M; Price, T J; Hardingham, J E

2017-06-01

Biomarkers, such as mutant RAS, predict resistance to anti-EGFR therapy in only a proportion of patients, and hence, other predictive biomarkers are needed. The aims were to identify candidate genes upregulated in colorectal cancer cell lines resistant to anti-EGFR monoclonal antibody treatment, to knockdown (KD) these genes in the resistant cell lines to determine if sensitivity to anti-EGFR antibody was restored, and finally to perform a pilot correlative study of EGR1 expression and outcomes in a cohort of metastatic colorectal cancer (mCRC) patients given cetuximab therapy. Comparative expression array analysis of resistant cell lines (SW48, COLO-320DM, and SNU-C1) vs sensitive cell lines (LIM1215, CaCo2, and SW948) was performed. The highest up-regulated gene in each resistant cell line was knocked down (KD) using RNA interference, and effect on proliferation was assessed with and without anti-EGFR treatment. Expression of the candidate genes in patients' tumours treated with cetuximab was assessed by immunohistochemistry; survival analyses were performed comparing high vs low expression. Genes significantly upregulated in resistant cell lines were EGR1 (early growth response protein 1), HBEGF (heparin-binding epidermal growth factor-like growth factor), and AKT3 (AKT serine/threonine kinase 3). KD of each gene resulted in the respective cells being more sensitive to anti-EGFR treatment, suggesting that the resistant phenotype was reversed. In the pilot study of mCRC patients treated with cetuximab, both median PFS (1.38 months vs 6.79 months; HR 2.77 95% CI 1.2-19.4) and median OS (2.59 months vs 9.82 months; HR 3.0 95% CI 1.3-23.2) were significantly worse for those patients with high EGR1 expression. High EGR1 expression may be a candidate biomarker of resistance to anti-EGFR therapy.

MiRNA-362-3p induces cell cycle arrest through targeting of E2F1, USF2 and PTPN1 and is associated with recurrence of colorectal cancer.

PubMed

Christensen, Lise Lotte; Tobiasen, Heidi; Holm, Anja; Schepeler, Troels; Ostenfeld, Marie S; Thorsen, Kasper; Rasmussen, Mads H; Birkenkamp-Demtroeder, Karin; Sieber, Oliver M; Gibbs, Peter; Lubinski, Jan; Lamy, Philippe; Laurberg, Søren; Oster, Bodil; Hansen, Kristian Q; Hagemann-Madsen, Rikke; Byskov, Kristina; Ørntoft, Torben F; Andersen, Claus L

2013-07-01

Colorectal cancer (CRC) is one of the leading causes of cancer deaths in Western countries. A significant number of CRC patients undergoing curatively intended surgery subsequently develop recurrence and die from the disease. MicroRNAs (miRNAs) are aberrantly expressed in cancers and appear to have both diagnostic and prognostic significance. In this study, we identified novel miRNAs associated with recurrence of CRC, and their possible mechanism of action. TaqMan(®) Human MicroRNA Array Set v2.0 was used to profile the expression of 667 miRNAs in 14 normal colon mucosas and 46 microsatellite stable CRC tumors. Four miRNAs (miR-362-3p, miR-570, miR-148 a* and miR-944) were expressed at a higher level in tumors from patients with no recurrence (p<0.015), compared with tumors from patients with recurrence. A significant association with increased disease free survival was confirmed for miR-362-3p in a second independent cohort of 43 CRC patients, using single TaqMan(®) microRNA assays. In vitro functional analysis showed that over-expression of miR-362-3p in colon cancer cell lines reduced cell viability, and proliferation mainly due to cell cycle arrest. E2F1, USF2 and PTPN1 were identified as potential miR-362-3p targets by mRNA profiling of HCT116 cells over-expressing miR-362-3p. Subsequently, these genes were confirmed as direct targets by Luciferase reporter assays and their knockdown in vitro phenocopied the effects of miR-362-3p over-expression. We conclude that miR-362-3p may be a novel prognostic marker in CRC, and hypothesize that the positive effects of augmented miR-362-3p expression may in part be mediated through the targets E2F1, USF2 and PTPN1. Copyright © 2012 UICC.

Aberrant methylation of GCNT2 is tightly related to lymph node metastasis of primary CRC.

PubMed

Nakamura, Kazunori; Yamashita, Keishi; Sawaki, Hiromichi; Waraya, Mina; Katoh, Hiroshi; Nakayama, Nobukazu; Kawamata, Hiroshi; Nishimiya, Hiroshi; Ema, Akira; Narimatsu, Hisashi; Watanabe, Masahiko

2015-03-01

Glycoprotein expression profile is dramatically altered in human cancers; however, specific glycogenes have not been fully identified. A comprehensive real-time polymerase chain reaction (PCR) system for glycogenes (CRPS-G) identified several outstanding glycogenes. GCNT2 was of particular interest after GCNT2 expression and epigenetics were rigorously investigated in primary colorectal cancer (CRC). The highlights of this work can be summarized as follows: (i) Expression of GCNT2 was remarkably suppressed. (ii) Silenced expression of GCNT2 was reactivated by combined demethylating agents. (iii) Promoter DNA methylation of GCNT2 was silenced in CRC cell lines and tissues. Hypomethylation of GCNT2 variant 2 is tightly associated with lymph node metastasis in primary CRC. (iv) GCNT2 methylation level in the normal tissues also showed a close association with that in the tumor tissues and reflected lymph node metastasis. We identified aberrant expression of GCNT2, which can be explained by promoter DNA hypermethylation. Hypomethylation of the GCNT2 variant 2 reflected lymph node metastasis of CRC in the tumor and normal tissues. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Novel evidence for curcumin and boswellic acid induced chemoprevention through regulation of miR-34a and miR-27a in colorectal cancer

PubMed Central

Toden, Shusuke; Okugawa, Yoshinaga; Buhrmann, Constanze; Nattamai, Durgha; Anguiano, Esperanza; Baldwin, Nicole; Shakibaei, Mehdi; Boland, C. Richard; Goel, Ajay

2015-01-01

Colorectal cancer (CRC) is one of the most common causes of cancer-associated mortality worldwide, but it is truly a preventable disease. Both curcumin and boswellic acids are well-established dietary botanicals with potent anti-tumorigenic properties which have been shown to modulate multiple oncogenic pathways. Recent data suggest that the chemopreventive effects of these botanicals may in part be mediated through regulation of key cancer-related microRNAs (miRNAs) and their downstream gene targets. Here, we investigated the anti-tumorigenic effects of curcumin and 3 acetyl-11-keto-β-boswellic acid (AKBA) on modulation of specific cancer-related miRNAs in CRC cells and validated their protective effects in vivo using a xenograft mouse model. Both curcumin and AKBA inhibited cellular proliferation, induced apoptosis and cell cycle arrest in CRC cell lines, and these effects were significantly enhanced with combined treatment. Gene-expression arrays revealed that curcumin and AKBA regulated distinct cancer signaling pathways including key cell-cycle regulatory genes. Combined bioinformatics and in-silico analysis identified apoptosis, proliferation and cell-cycle regulatory signaling pathways as key modulators of curcumin and AKBA-induced anti-cancer effects. We discovered that curcumin and AKBA induced upregulation of tumor-suppressive miR-34a and downregulation of miR-27a in CRC cells. Furthermore, we demonstrated in a mouse xenograft model that both curcumin and AKBA treatments suppressed tumor growth, which corresponded with alterations in the expression of miR-34a and miR-27a, consistent with our in vitro findings. Herein we provide novel mechanistic evidence for the chemopreventive effects of curcumin and AKBA through regulation of specific miRNAs in colorectal cancer. PMID:25712055

Integrated genomic analysis of colorectal cancer progression reveals activation of EGFR through demethylation of the EREG promoter

PubMed Central

Qu, X; Sandmann, T; Frierson, H; Fu, L; Fuentes, E; Walter, K; Okrah, K; Rumpel, C; Moskaluk, C; Lu, S; Wang, Y; Bourgon, R; Penuel, E; Pirzkall, A; Amler, L; Lackner, M R; Tabernero, J; Hampton, G M; Kabbarah, O

2016-01-01

Key molecular drivers that underlie transformation of colonic epithelium into colorectal adenocarcinoma (CRC) are well described. However, the mechanisms through which clinically targeted pathways are activated during CRC progression have yet to be elucidated. Here, we used an integrative genomics approach to examine CRC progression. We used laser capture microdissection to isolate colonic crypt cells, differentiated surface epithelium, adenomas, carcinomas and metastases, and used gene expression profiling to identify pathways that were differentially expressed between the different cell types. We identified a number of potentially important transcriptional changes in developmental and oncogenic pathways, and noted a marked upregulation of EREG in primary and metastatic cancer cells. We confirmed this pattern of gene expression by in situ hybridization and observed staining consistent with autocrine expression in the tumor cells. Upregulation of EREG during the adenoma–carcinoma transition was associated with demethylation of two key sites within its promoter, and this was accompanied by an increase in the levels of epidermal growth factor receptor (EGFR) phosphorylation, as assessed by reverse-phase protein analysis. In CRC cell lines, we demonstrated that EREG demethylation led to its transcriptional upregulation, higher levels of EGFR phosphorylation, and sensitization to EGFR inhibitors. Low levels of EREG methylation in patients who received cetuximab as part of a phase II study were associated with high expression of the ligand and a favorable response to therapy. Conversely, high levels of promoter methylation and low levels of EREG expression were observed in tumors that progressed after treatment. We also noted an inverse correlation between EREG methylation and expression levels in several other cancers, including those of the head and neck, lung and bladder. Therefore, we propose that upregulation of EREG expression through promoter demethylation might be an important means of activating the EGFR pathway during the genesis of CRC and potentially other cancers. PMID:27270421

PDGFRα/β and VEGFR2 polymorphisms in colorectal cancer: incidence and implications in clinical outcome

PubMed Central

2012-01-01

Background Angiogenesis plays an essential role in tumor growth and metastasis, and is a major target in cancer therapy. VEGFR and PDGFR are key players involved in this process. The purpose of this study was to assess the incidence of genetic variants in these receptors and its potential clinical implications in colorectal cancer (CRC). Methods VEGFR2, PDGFRα and PDGFRβ mutations were evaluated by sequencing their tyrosine kinase domains in 8 CRC cell lines and in 92 samples of patients with CRC. Correlations with clinicopathological features and survival were analyzed. Results Four SNPs were identified, three in PDGFRα [exon 12 (A12): c.1701A>G; exon 13 (A13): c.1809G>A; and exon 17 (A17): c.2439+58C>A] and one in PDGFRβ [exon 19 (B19): c.2601A>G]. SNP B19, identified in 58% of tumor samples and in 4 cell lines (LS174T, LS180, SW48, COLO205), was associated with higher PDGFR and pPDGFR protein levels. Consistent with this observation, 5-year survival was greater for patients with PDGFR B19 wild type tumors (AA) than for those harboring the G-allele genotype (GA or GG) (51% vs 17%; p=0.073). Multivariate analysis confirmed SNP B19 (p=0.029) was a significant prognostic factor for survival, independent of age (p=0.060) or TNM stage (p<0.001). Conclusions PDGFRβ exon 19 c.2601A>G SNP is commonly encountered in CRC patients and is associated with increased pathway activation and poorer survival. Implications regarding its potential influence in response to PDGFR-targeted agents remain to be elucidated. PMID:23146028

Zoledronate Triggers Vδ2 T Cells to Destroy and Kill Spheroids of Colon Carcinoma: Quantitative Image Analysis of Three-Dimensional Cultures.

PubMed

Varesano, Serena; Zocchi, Maria Raffaella; Poggi, Alessandro

2018-01-01

New successful anti-cancer strategies are based on the stimulation of immune reaction against tumors: however, preclinical testing of such treatments is still a challenge. To improve the screening of anti-cancer drugs, three-dimensional (3D) culture systems, including spheroids, have been validated as preclinical models. We propose the spheroid 3D system to test anti-tumor drug-induced immune responses. We show that colorectal carcinoma (CRC) spheroids, generated with the epithelial growth factor (EGF), can be co-cultured with Vδ2 T cells to evaluate the anti-tumor activity of these effector lymphocytes. By computerized image analysis, the precise and unbiased measure of perimeters and areas of tumor spheroids is achievable, beside the calculation of their volume. CRC spheroid size is related to ATP content and cell number, as parameters for cell metabolism and proliferation; in turn, crystal violet staining can check the viability of cells inside the spheroids to detect tumor killing by Vδ2 T cells. In this 3D cultures, we tested (a) zoledronate that is known to activate Vδ2 T cells and (b) the therapeutic anti-EGF receptor humanized antibody cetuximab that can elicit the antibody-dependent cytotoxicity of tumor cells by effector lymphocytes. Zoledronate triggers Vδ2 T cells to kill and degrade CRC spheroids; we detected the T-cell receptor dependency of zoledronate effect, conceivably due to the recognition of phosphoantigens produced as a drug effect on target cell metabolism. In addition, cetuximab triggered Vδ2 T lymphocytes to exert the antibody-dependent cellular cytotoxicity of CRC spheroids. Finally, the system reveals differences in the sensitivity of CRC cell lines to the action of Vδ2 T lymphocytes and in the efficiency of anti-tumor effectors from distinct donors. A limitation of this model is the absence of cells, including fibroblasts, that compose tumor microenvironment and influence drug response. Nevertheless, the system can be improved by setting mixed spheroids, made of stromal and cancer cells. We conclude that this type of spheroid 3D culture is a feasible and reliable system to evaluate and measure anti-tumor drug-induced immune responses beside direct anti-cancer drug effect.

Aberrant methylation of PSD disturbs Rac1-mediated immune responses governing neutrophil chemotaxis and apoptosis in ulcerative colitis-associated carcinogenesis.

PubMed

Kato, Takaharu; Suzuki, Koichi; Okada, Shinichiro; Kamiyama, Hidenori; Maeda, Takafumi; Saito, Masaaki; Koizumi, Kei; Miyaki, Yuichiro; Konishi, Fumio

2012-04-01

We previously reported that the Pleckstrin and Sec7 domain-containing (PSD) gene is preferentially methylated in patients with ulcerative colitis (UC) who developed colorectal cancer (CRC), and is implicated in UC-associated carcinogenesis through its inhibition of apoptosis. This study aimed to determine the potential effect of PSD methylation on its downstream molecule, Ras-related C3 botulinum toxin substrate 1 (Rac1), which governs neutrophil chemotaxis and apoptosis signaling. PSD was knocked down in a normal human fibroblast cell line (HNDF) and a neutrophil-like cell line (HL-60). Both NHDF and HL-60 cells exhibited numerous filamentous-actin (F-actin) rich membrane extensions, resulting in the activation of Rac1; this activation was hampered by PSD silencing. Lipopolysaccharide, a reactive oxygen species (ROS) inducer, stimulated NHDF cells to release ROS and activated caspase‑3/7 in the presence of neutrophils, which was inhibited by PSD knockdown. Migration assays demonstrated that chemotaxis of HL-60 cells was affected by PSD silencing in NHDF cells. Tissue sections from 6 UC patients with CRC and 15 UC patients without CRC were examined. To verify Rac1-mediated chemotaxis in tissue sections, we evaluated the grade of neutrophil infiltration by histological assessment and assessed F-actin and PSD expression by immunohistochemistry. Neutrophil infiltration, F-actin and PSD expression were significantly decreased in specimens from UC patients with PSD methylation compared with those without. Decreased levels of F-actin expression were observed in colorectal mucosa, as well as in infiltrating cells with PSD methylation. PSD expression was preferentially inhibited in colorectal mucosa by PSD methylation, whereas PSD expression was rarely observed in infiltrating cells, regardless of PSD methylation status. These data indicate that aberrant methylation of PSD occurs in UC-associated colorectal mucosa, enabling circumvention of Rac1-mediated immune responses governing neutrophil chemotaxis and apoptosis, and thus plays a pivotal role in the mechanisms underlying UC-associated carcinogenesis.

Differential RNA-seq analysis comparing APC-defective and APC-restored SW480 colorectal cancer cells.

PubMed

King, Lauren E; Love, Christopher G; Sieber, Oliver M; Faux, Maree C; Burgess, Antony W

2016-03-01

The adenomatous polyposis coli (APC) tumour suppressor gene is mutated in about 80% of colorectal cancers (CRC) Brannon et al. (2014) [1]. APC is a large multifunctional protein that regulates many biological functions including Wnt signalling (through the regulation of beta-catenin stability) Reya and Clevers (2005) [2], cell migration Kroboth et al. (2007), Sansom et al. (2004) [3], [4], mitosis Kaplan et al. (2001) [5], cell adhesion Faux et al. (2004), Carothers et al. (2001) [6], [7] and differentiation Sansom et al. (2004) [4]. Although the role of APC in CRC is often described as the deregulation of Wnt signalling, its other biological functions suggest that there are other factors at play that contribute to the onset of adenomas and the progression of CRC upon the truncation of APC. To identify genes and pathways that are dysregulated as a consequence of loss of function of APC, we compared the gene expression profiles of the APC mutated human CRC cell line SW480 following reintroduction of wild-type APC (SW480 + APC) or empty control vector (SW480 + vector control) Faux et al. (2004) . Here we describe the RNA-seq data derived for three biological replicates of parental SW480, SW480 + vector control and SW480 + APC cells, and present the bioinformatics pipeline used to test for differential gene expression and pathway enrichment analysis. A total of 1735 genes showed significant differential expression when APC was restored and were enriched for genes associated with cell polarity, Wnt signalling and the epithelial to mesenchymal transition. There was additional enrichment for genes involved in cell-cell adhesion, cell-matrix junctions, angiogenesis, axon morphogenesis and cell movement. The raw and analysed RNA-seq data have been deposited in the Gene Expression Omnibus (GEO) database under accession number GSE76307. This dataset is useful for further investigations of the impact of APC mutation on the properties of colorectal cancer cells.

B7-H1 expression is associated with expansion of regulatory T cells in colorectal carcinoma.

PubMed

Hua, Dong; Sun, Jing; Mao, Yong; Chen, Lu-Jun; Wu, Yu-Yu; Zhang, Xue-Guang

2012-03-07

To investigate the expression of B7-H1 in human colorectal carcinoma (CRC) to define its regulating effects on T cells in tumor microenvironment. One hundred and two paraffin blocks and 33 fresh samples of CRC tissues were subject to this study. Immunohistochemistry was performed for B7-H1 and CD3 staining in CRC tissues. Ficoll-Hypaque density gradient centrifugation was used to isolate peripheral blood mononuclear cells of fresh CRC tissues; flow cytometry and immunofluorescence staining were used for detection of regulatory T cells. Data was analyzed with statistical software. Costimulatory molecule B7-H1 was found strongly expressed in CRC tissues, localized in tumor cell membrane and cytoplasm, while weak or none expression of B7-H1 was detected in pared normal colorectal tissues. Meanwhile, CD3 positive T cells were found congregated in CRC tumor nest and stroma. Statistic analysis showed that B7-H1 expression level was negatively correlated to the total T cell density in tumor nest (P < 0.0001) and tumor stroma (P = 0.0200) of 102 cases of CRC tissues. Among the total T cells, a variable amount of regulatory T cells with a clear Foxp3⁺ (forkhead box P3) staining could be detected in CRC tissues and patients' blood. Interestingly, in the 33 samples (15 cases of B7-H1(high) CRC tissues and 18 cases of B7-H1(low) CRC tissues) of freshly isolated mononuclear cells from CRC tissues, the percentages of CD4⁺Foxp3⁺ and CD8⁺Foxp3⁺ regulatory T cells were found remarkably higher in B7-H1(high) CRC tissues than in B7-H1(low) CRC tissues (P = 0.0024, P = 0.0182), indicating that B7-H1 expression was involved in proliferation of regulatory T cell. No significant difference was found in CRC peripheral blood (P = 0.0863, P = 0.0678). PD-1 is the specific ligand for B7-H1 pathway transferring inhibitory signal to T cell, which is expressed by activated T cell. Our further analysis of PD-1 expression on T cells in CRC tissues showed that conventional T cells (CD4⁺Foxp3⁻/CD8⁺Foxp3⁻), which was thought to contribute to the anti-tumor immune response, highly expressed PD-1; while regulatory T cells (CD4⁺Foxp3⁺/CD8⁺Foxp3⁻) almost failed to express PD-1. The average percentage of PD-1 expression on regulatory T cells was significantly higher than the percentage of PD-1 on conventional T cells (CD4⁺Foxp3⁻ T cell, P < 0.0001; CD8⁺Foxp3⁻ T cell, P < 0.0001). The diverse expression of PD-1 might lead to different fate of T cell subsets in B7-H1 over-expression CRC tumor microenvironment. B7-H1 expression in tumor cells can inhibit the conventional T cell proliferation in tumor microenvironment through the PD-1 expression on conventional T cells.

Overexpression of paxillin induced by miR-137 suppression promotes tumor progression and metastasis in colorectal cancer

PubMed Central

Xu, Rui-Hua

2013-01-01

The deregulation of paxillin (PXN) has been involved in the progression and metastasis of different malignancies including colorectal cancer (CRC). miR-137 is frequently suppressed in CRC. PXN is predicted to be a direct target of miR-137 in CRC cells. On this basis, we hypothesized that overexpression of PXN induced by suppression of miR-137 may promote tumor progression and metastasis and predicts poor prognosis. We detected the expression of PXN and miR-137 in clinical tumor tissues by immunohistochemical analysis and real-time PCR, positive PXN staining was observed in 198 of the 247 (80.1%) cases, whereas no or weak PXN staining was observed in the adjacent non-cancerous area. Higher level of PXN messenger RNA (mRNA) and lower level of miR-137 was observed in cancer tissues than adjacent non-cancerous tissues. High expression of PXN and low expression of miR-137 was associated with aggressive tumor phenotype and adverse prognosis. Moreover, the expression of PXN was negatively correlated with miR-137 expression. A dual-luciferase reporter gene assay validated that PXN was a direct target of miR-137. The use of miR-137 mimics or inhibitor could decrease or increase PXN mRNA and protein levels in CRC cell lines. Knockdown of PXN or ectopic expression of miR-137 could markedly inhibit cell proliferation, migration and invasion in vitro and repress tumor growth and metastasis in vivo. Taken together, these results demonstrated that overexpression of PXN induced by suppression of miR-137 promotes tumor progression and metastasis and could serve as an independent prognostic indicator in CRC patients. PMID:23275153

Competing endogenous RNA network crosstalk reveals novel molecular markers in colorectal cancer.

PubMed

Samir, Nehal; Matboli, Marwa; El-Tayeb, Hanaa; El-Tawdi, Ahmed; Hassan, Mohmed K; Waly, Amr; El-Akkad, Hesham A E; Ramadan, Mohamed G; Al-Belkini, Tarek N; El-Khamisy, Sherif; El-Asmar, Farid

2018-05-08

The competing endogenous RNA networks play a pivotal role in cancer diagnosis and progression. Novel properstrategies for early detection of colorectal cancer (CRC) are strongly needed. We investigated a novel CRC-specific RNA-based integrated competing endogenous network composed of lethal3 malignant brain tumor like1 (L3MBTL1) gene, long non-coding intergenic RNA- (lncRNA RP11-909B2.1) and homo sapiens microRNA-595 (hsa-miRNA-595) using in silico data analysis. RT-qPCR-based validation of the network was achieved in serum of 70 patients with CRC, 40 patients with benign colorectal neoplasm, and 20 healthy controls. Moreover, in cancer tissues of 20 of the 70 CRC cases were involved in the study. The expression of RNA-based biomarker network in both CRC and adjacent non-tumor tissues and their correlation with the serum levels of this network members was investigated. Lastly, the expression levels of the chosen ceRNA was verified in CRC cell line. Our results revealed that the three RNAs-based biomarker network (long non-coding intergenic RNA-[lncRNA RP11-909B2.1], Homo sapiens microRNA-595 [hsa-miRNA-595], and L3MBTL1 mRNA), had high sensitivity and specificity for discriminating CRC from healthy controls and also from benign colorectal neoplasm. The data suggest that among these three RNAs, serum lncRNA RP11-909B2.1 could be a promising independent prognostic factors in CRC. The circulatory RNA based biomarker panel can act as potential biomarker for CRC diagnosis and prognosis. © 2018 Wiley Periodicals, Inc.

Upregulated STAT3 and RhoA signaling in colorectal cancer (CRC) regulate the invasion and migration of CRC cells.

PubMed

Zhang, G-Y; Yang, W-H; Chen, Z

2016-05-01

We aimed to reveal the expression and activation of signal transducers and activators of transcription 3 (STAT3) and RhoA/Rho-associated coiled-coil forming kinase 1 (ROCK1) signaling in CRC tissues, and to investigate the regulatory role of STAT3 and RhoA signaling in the invasion and migration of colorectal cancer cells. We examined the expression of STAT3, RhoA and ROCK1 in CRC tissues with real-time PCR and Western blotting methods. And then we examined the interaction between STAT3 and RhoA/ROCK1 signaling in CRC HT-29 cells with gain-of-function and loss-of-function strategies. In addition, we determined the regulation by STAT3 and RhoA/ROCK1 on the invasion and migration of CRC HT-29 cells. Our study demonstrated a significant upregulation of RhoA and ROCK1 expression and STAT3-Y705 phosphorylation in 32 CRC specimens, compared to the 17 normal CRC tissues. Further study demonstrated there was a coordination between STAT3 and RhoA/Rock signaling in the HT-29 cells. Moreover, STAT3 knockdown or RhoA knockdown significantly repressed the migration and invasion in HT-29 cells and vice versa. STAT3 and RhoA signaling regulate the invasion and migration of CRC cells, implying the orchestrated and oncogenic roles of STAT3 and RhoA/ROCK1 signaling in CRC.

Ellagitannins from pomegranate ameliorates 5-fluorouracil-induced intestinal mucositis in rats while enhancing its chemotoxicity against HT-29 colorectal cancer cells through intrinsic apoptosis induction.

PubMed

Chen, Xiao-Xin; Lam, Kar Ho; Feng, Yibin; Xu, Kai; Sze, Stephen C W; Tang, Chi Wai; Leung, George P H; Lee, Calvin Kai-Fai; Shi, Jun; Yang, Zhijun; Li, Sheng-Tao; Zhang, Zhang-Jin; Zhang, Yanbo

2018-06-19

Worldwide, colorectal cancer (CRC) is a deleterious disease causing millions of death annually. 5-Fluorouracil (5-FU) is a first-line chemotherapy for CRC, but chemoresistance and gastrointestinal mucositis limit its efficacy. Polyphenol-rich foods are increasingly popular due to their potential beneficial role in cancer. Ellagitannins is a group of phenolic compounds commonly found in pomegranate, strawberries, raspberries, etc. The objective of this study was to explore whether ellagitannins from pomegranate (PETs) could ameliorate 5-FU-induced intestinal mucositis and enhance its efficacy against CRC. The results showed that PETs (100 mg/kg) counteracted 5-FU-induced intestinal mucositis in rats. The number of apoptotic cells per crypt was reduced from 1.50±0.21 to 0.85±0.18 (P<0.05). Moreover, PETs induced HT-29 CRC cell death through intrinsic apoptosis as demonstrated by dissipation of mitochondrial membrane potential, increased Bax to Bcl-2 ratio, and cleavage of caspase 9 and caspase 3. PETs and 5-FU combination treatments exhibited synergistic cytotoxicity against HT-29 cells with a weighted combination index of 0.3494. PETs (80 µg/mL) and 5-FU (40 µg/mL) treatments for 48 h induced 14.03±0.76% and 16.42±1.15% of HT-29 cells to undergo apoptosis while the combination treatment further increased apoptosis cells to 34.00±1.54% (P<0.05). Combination treatment of the cells also enhanced S phase cell cycle arrest as compared with PETs or 5-FU monotherapy (P<0.05). These results suggest that dietary ellagitannins from pomegranate could alleviate intestinal mucositis in rats induced by 5-FU while enhancing its toxicity against HT-29 cells through potentiation of apoptosis and cell cycle arrest.

Tumor-associated down-regulation of 15-lipoxygenase-1 is reversed by celecoxib in colorectal cancer.

PubMed

Heslin, Martin J; Hawkins, Ashley; Boedefeld, William; Arnoletti, J Pablo; Frolov, Andrey; Soong, Richie; Urist, Marshall M; Bland, Kirby I

2005-06-01

To evaluate the role of celecoxib on 15-lipoxygenase-1 (15-LOX-1) expression, protein levels, and rates of apoptosis in colorectal cancer cell lines. Also, to evaluate the expression of 15-LOX-1 in human normal mucosa, adenoma, and carcinoma with correlation to overall survival. The function of 15-LOX-1 is to maintain normal rates of apoptosis (programmed cell death). Decreased apoptosis is one mechanism of cancer growth and dissemination. It is our hypothesis that expression of 15-LOX-1 is reduced in human colorectal cancer (CRC) and the administration of celecoxib can reverse this process and induce apoptosis. Effect of celecoxib in cell culture: The effect of 40 micromol/L celecoxib was compared with untreated controls in tissue culture utilizing HT-29 and DLD-1 CRC cell lines. Expression of 15-LOX-1 protein was measured by immunoblot. Induction of apoptosis was evaluated by annexin V staining. All data are presented as mean +/- SEM, with significance defined as P < 0.05. 15-LOX-1 in human CRC: From February 1998 to January 2002, 126 patients underwent surgical resection of either colorectal adenomas (n = 24) or carcinomas (n = 102), or both (n = 25). Tissue was macrodissected, snap frozen, and stored at -80 degrees C. After tissue processing, RNA was extracted and gene expression of 15-LOX-1 was quantified utilizing ABI prism real-time quantitative RT-PCR. Significance evaluated by the Wilcoxon signed rank test. Effect of celecoxib in cell culture: After 72 hours of treatment with celecoxib, immunoblot demonstrated a 1.5- to 2-fold increase in 15-LOX-1 protein expression in HT-29 and DLD-1 cells, respectively. Celecoxib produced greater than a 2-fold increase in the rate of apoptosis compared with control cells in both cell lines (P < 0.05). 15-LOX-1 in human CRC: The mean age of the patients was 62 +/- 1 years; 78% were white and 48% were female. The mean size of the polyps and cancers were 3.0 +/- 0.4 and 5.0 +/- 0.1 cm, respectively. Expression of 15-LOX-1 relative to S9 was 30 in normal mucosa and significantly down-regulated to 11 in adenomas and 16 in carcinomas (P < 0.05). 15-LOX-1 gene expression is significantly reduced in both human colorectal adenomas and carcinomas and associated with decreased survival. Administration of celecoxib restores 15-LOX-1 protein expression and induces apoptosis. Down-regulation of 15-LOX-1 is an early event in the adenoma to carcinoma sequence, and reversal with celecoxib may represent one mechanism for chemoprevention of polyps or treatment of carcinomas.

CYP24A1 inhibition facilitates the anti-tumor effect of vitamin D3 on colorectal cancer cells

PubMed Central

Kósa, János P; Horváth, Péter; Wölfling, János; Kovács, Dóra; Balla, Bernadett; Mátyus, Péter; Horváth, Evelin; Speer, Gábor; Takács, István; Nagy, Zsolt; Horváth, Henrik; Lakatos, Péter

2013-01-01

AIM: The effects of vitamin D3 have been investigated on various tumors, including colorectal cancer (CRC). 25-hydroxyvitamin-D3-24-hydroxylase (CYP24A1), the enzyme that inactivates the active vitamin D3 metabolite 1,25-dihydroxyvitamin D3 (1,25-D3), is considered to be the main enzyme determining the biological half-life of 1,25-D3. During colorectal carcinogenesis, the expression and concentration of CYP24A1 increases significantly, suggesting that this phenomenon could be responsible for the proposed efficacy of 1,25-D3 in the treatment of CRC. The aim of this study was to investigate the anti-tumor effects of vitamin D3 on the human CRC cell line Caco-2 after inhibition of the cytochrome P450 component of CYP24A1 activity. METHODS: We examined the expression of CYP24A1 mRNA and the effects of 1,25-D3 on the cell line Caco-2 after inhibition of CYP24A1. Cell viability and proliferation were determined by means of sulforhodamine-B staining and bromodeoxyuridine incorporation, respectively, while cytotoxicity was estimated via the lactate dehydrogenase content of the cell culture supernatant. CYP24A1 expression was measured by real-time reverse transcription polymerase chain reaction. A number of tetralone compounds were synthesized to investigate their CP24A1 inhibitory activity. RESULTS: In response to 1,25-D3, CYP24A1 mRNA expression was enhanced significantly, in a time- and dose-dependent manner. Caco-2 cell viability and proliferation were not influenced by the administration of 1,25-D3 alone, but were markedly reduced by co-administration of 1,25-D3 and KD-35, a CYP24A1-inhibiting tetralone. Our data suggest that the mechanism of action of co-administered KD-35 and 1,25-D3 does not involve a direct cytotoxic effect, but rather the inhibition of cell proliferation. CONCLUSION: These findings demonstrate that the selective inhibition of CYP24A1 by compounds such as KD-35 may be a new approach for enhancement of the anti-tumor effect of 1,25-D3 on CRC. PMID:23674869

Heterogeneity of neuroblastoma cell identity defined by transcriptional circuitries.

PubMed

Boeva, Valentina; Louis-Brennetot, Caroline; Peltier, Agathe; Durand, Simon; Pierre-Eugène, Cécile; Raynal, Virginie; Etchevers, Heather C; Thomas, Sophie; Lermine, Alban; Daudigeos-Dubus, Estelle; Geoerger, Birgit; Orth, Martin F; Grünewald, Thomas G P; Diaz, Elise; Ducos, Bertrand; Surdez, Didier; Carcaboso, Angel M; Medvedeva, Irina; Deller, Thomas; Combaret, Valérie; Lapouble, Eve; Pierron, Gaelle; Grossetête-Lalami, Sandrine; Baulande, Sylvain; Schleiermacher, Gudrun; Barillot, Emmanuel; Rohrer, Hermann; Delattre, Olivier; Janoueix-Lerosey, Isabelle

2017-09-01

Neuroblastoma is a tumor of the peripheral sympathetic nervous system, derived from multipotent neural crest cells (NCCs). To define core regulatory circuitries (CRCs) controlling the gene expression program of neuroblastoma, we established and analyzed the neuroblastoma super-enhancer landscape. We discovered three types of identity in neuroblastoma cell lines: a sympathetic noradrenergic identity, defined by a CRC module including the PHOX2B, HAND2 and GATA3 transcription factors (TFs); an NCC-like identity, driven by a CRC module containing AP-1 TFs; and a mixed type, further deconvoluted at the single-cell level. Treatment of the mixed type with chemotherapeutic agents resulted in enrichment of NCC-like cells. The noradrenergic module was validated by ChIP-seq. Functional studies demonstrated dependency of neuroblastoma with noradrenergic identity on PHOX2B, evocative of lineage addiction. Most neuroblastoma primary tumors express TFs from the noradrenergic and NCC-like modules. Our data demonstrate a previously unknown aspect of tumor heterogeneity relevant for neuroblastoma treatment strategies.

Toll-like receptor 3 as an immunotherapeutic target for KRAS mutated colorectal cancer

PubMed Central

Maitra, Radhashree; Augustine, Titto; Dayan, Yitzchak; Chandy, Carol; Coffey, Matthew; Goel, Sanjay

2017-01-01

New therapeutic interventions are essential for improved management of patients with metastatic colorectal cancer (mCRC). This is especially critical for those patients whose tumors harbor a mutation in the KRAS oncogene (40-45% of all patients). This patient cohort is excluded from receiving anti-EGFR monoclonal antibodies that have added a significant therapeutic benefit for KRAS wild type CRC patients. Reovirus, a double stranded (ds) RNA virus is in clinical development for patients with chemotherapy refractory KRAS mutated tumors. Toll Like Receptor (TLR) 3, a member of the toll like receptor family of the host innate immune system is the pattern recognition motif for dsRNA pathogens. Using TLR3 expressing commercial HEK-Blue™-hTLR3 cells we confirm that TLR3 is the host pattern recognition motif responsible for the detection of reovirus. Further, our investigation with KRAS mutated HCT116 cell line showed that effective expression of host TLR3 dampens the infection potential of reovirus by mounting a robust innate immune response. Down regulation of TLR3 expression with siRNA improves the anticancer activity of reovirus. In vivo experiments using human CRC cells derived xenografts in athymic mice further demonstrate the beneficial effects of TLR3 knock down by improving tumor response rates to reovirus. Strategies to mitigate the TLR3 response pathway can be utilized as a tool towards improved reovirus efficacy to specifically target the dissemination of KRAS mutated CRC. PMID:28422714

STIM1 Overexpression Promotes Colorectal Cancer Progression, Cell Motility and COX-2 Expression

PubMed Central

Wang, Jaw-Yuan; Sun, Jianwei; Huang, Ming-Yii; Wang, Yu-Shiuan; Hou, Ming-Feng; Sun, Yan; He, Huifang; Krishna, Niveditha; Chiu, Siou-Jin; Lin, Shengchen; Yang, Shengyu; Chang, Wei-Chiao

2014-01-01

Tumor metastasis is the major cause of death among cancer patients, with more than 90% of cancer-related death attributable to the spreading of metastatic cells to secondary organs. Store-operated Ca2+ entry (SOCE) is the predominant Ca2+ entry mechanism in most cancer cells, and STIM1 is the endoplasmic reticulum (ER) Ca2+ sensor for store-operated channels (SOC). Here we reported that the STIM1 was overexpressed in colorectal cancer (CRC) patients. STIM1 overexpression in CRC was significantly associated with tumor size, depth of invasion, lymphnode metastasis status and serum levels of carcinoembryonic antigen. Furthermore, ectopic expression of STIM1 promoted CRC cell motility, while depletion of STIM1 with shRNA inhibited CRC cell migration. Our data further suggested that STIM1 promoted CRC cell migration through increasing the expression of cyclooxygenase-2 (COX-2) and production of prostaglandin E2 (PGE2). Importantly, ectopically expressed COX-2 or exogenous PGE2 were able to rescue migration defect in STIM1 knockdown CRC cells, and inhibition of COX-2 with ibuprofen and indomethacin abrogated STIM1-mediated CRC cell motility. In short, our data provided clinicopathological significance for STIM1 and store-operated Ca2+ entry in CRC progression, and implicated a role for COX-2 in STIM1-mediated CRC metastasis. Our studies also suggested a new approach to inhibit STIM1-mediated metastasis with COX-2 inhibitors. PMID:25381814

The Effect of Inflammatory Status on Butyrate and Folate Uptake by Tumoral (Caco-2) and Non-Tumoral (IEC-6) Intestinal Epithelial Cells

PubMed Central

Couto, Mafalda R.; Gonçalves, Pedro; Catarino, Telmo A.; Martel, Fátima

2017-01-01

Objective Colorectal cancer (CRC) is the second leading cause of cancer death in occidental countries. Chronic inflammatory bowel disease (crohn’s disease and ulcerative colitis) is associated with an increased risk for CRC development. The aim of this work was to investigate the relationship between inflammatory status and absorption of nutrients with a role in CRC pathogenesis. Materials and Methods In this experimental study, we evaluated the in vitro effect of tumour necrosis factor-alpha (TNF-α), interferon-γ (IF-γ), and acetylsalicylic acid on 14C-butyrate (14C- BT), 3H-folic acid (3H-FA) uptake, and on proliferation, viability and differentiation of Caco-2 and IEC-6 cells in culture. Results The proinflammatory cytokines TNF-α and INF-γ were found to decrease uptake of a low concentration of 14C-BT (10 µM) by Caco-2 (tumoral) and IEC-6 (normal) intestinal epithelial cell lines. However, the effect of TNF-α and INF-γ in IEC-6 cells is most probably related to a cytotoxic and antiproliferative impact. In contrast, INF-γ increases uptake of a high concentration (10 mM) of 14C-BT in Caco-2 cells. The anticarcinogenic effect of BT (10 mM) in these cells is not affected by the presence of this cytokine. On the other hand, acetylsalicylic acid stimulates 14C-BT uptake by Caco-2 cells and potentiates its antiproliferative effect. Finally, both TNF-α and INF-γ cause a significant decrease in 3H-FA uptake by Caco-2 cells. Conclusion The inflammatory status has an impact upon cellular uptake of BT and FA, two nutrients with a role in CRC pathogenesis. Moreover, the anti-inflammatory acetylsalicylic acid potentiates the anticarcinogenic effect of BT in Caco-2 cells by increasing its cellular uptake. PMID:28580313

IL1B-CGTC haplotype is associated with colorectal cancer in admixed individuals with increased African ancestry

PubMed Central

Sanabria-Salas, María Carolina; Hernández-Suárez, Gustavo; Umaña-Pérez, Adriana; Rawlik, Konrad; Tenesa, Albert; Serrano-López, Martha Lucía; Sánchez de Gómez, Myriam; Rojas, Martha Patricia; Bravo, Luis Eduardo; Albis, Rosario; Plata, José Luis; Green, Heather; Borgovan, Theodor; Li, Li; Majumdar, Sumana; Garai, Jone; Lee, Edward; Ashktorab, Hassan; Brim, Hassan; Li, Li; Margolin, David; Fejerman, Laura; Zabaleta, Jovanny

2017-01-01

Single-nucleotide polymorphisms (SNPs) in cytokine genes can affect gene expression and thereby modulate inflammation and carcinogenesis. However, the data on the association between SNPs in the interleukin 1 beta gene (IL1B) and colorectal cancer (CRC) are conflicting. We found an association between a 4-SNP haplotype block of the IL1B (-3737C/-1464G/-511T/-31C) and CRC risk, and this association was exclusively observed in individuals with a higher proportion of African ancestry, such as individuals from the Coastal Colombian region (odds ratio, OR 2.06; 95% CI 1.31–3.25; p < 0.01). Moreover, a significant interaction between this CRC risk haplotype and local African ancestry dosage was identified in locus 2q14 (p = 0.03). We conclude that Colombian individuals with high African ancestry proportions at locus 2q14 harbour more IL1B-CGTC copies and are consequently at an increased risk of CRC. This haplotype has been previously found to increase the IL1B promoter activity and is the most frequent haplotype in African Americans. Despite of limitations in the number of samples and the lack of functional analysis to examine the effect of these haplotypes on CRC cell lines, our results suggest that inflammation and ethnicity play a major role in the modulation of CRC risk. PMID:28157220

Activation of IRE1α-XBP1 pathway induces cell proliferation and invasion in colorectal carcinoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin, Chun; Jin, Zhao; Chen, Nian-zhao

2016-01-29

Cell proliferation and tumor metastasis are considered as the main reasons for death in colorectal carcinoma (CRC). IRE1α-XBP1 pathway is the most conserved UPR pathways, which are activated during ER stress caused by the accumulation of unfolded or misfolded protein in the lumen of ER. Here, we demonstrated the critical role of IRE1α-XBP1 pathway and underlying molecular mechanism in cell proliferation and tumor metastasis in CRC. By the use of tissue microarray analysis of samples from 119 patients with CRC, IRE1α was determined to be an independent predictor of overall survival as higher expression of IRE1α in CRC patients showedmore » lower survival rates (p = 0.0041). RNA interference and ectopic expression of IRE1α were applied to determine the molecular effects of IRE1α in CRC cells. The silencing of IRE1α inhibited the proliferation and blocked the invasion of CRC cells in vitro, while ectopic expression of IRE1α in turn promoted cell proliferation and invasion. IRE1α-XBP1 pathway regulated the mitosis of CRC cells through the directly binding of XBP1s to Cyclin D1 promoter to activate Cyclin D1 expression. Our results reveal that IRE1α-XBP1 pathway plays an important role in tumor progression and epithelial-to-mesenchymal transition (EMT), and IRE1α could be employed as a novel prognostic marker and a promising therapeutic target for CRC. - Highlights: • IRE1 was determined to be an independent predictor of overall survival in CRC patient. • IRE1-XBP1 pathway promoted CRC cell proliferation through regulating Cyclin D1 expression. • IRE1-XBP1 pathway played important role in EMT of CRC cells.« less

CENPI is overexpressed in colorectal cancer and regulates cell migration and invasion.

PubMed

Ding, Na; Li, Rongxin; Shi, Wenhao; He, Cui

2018-06-21

Centromere protein I (CENPI),an important member of centromere protein family, has been suggest to serve as a oncogene in breast cancer, but the clinical significance and biological function of CENPI in colorectal cancer (CRC) is still unclear. In our results, we found CENPI was overexpressed in CRC tissues and cells, and associated with clinical stage, tumor depth, lymph node metastasis, distant metastasis and differentiation in CRC patients. However, there was no significant association between CENPI protein expression and overall survival time in colon cancer patients and rectal cancer patients through analyzing TCGA survival data. Moreover, CENPI mRNA and protein were increased in metastatic lymph nodes compared with primary CRC tissues. Down-regulation of CENPI expression suppresses CRC cell migration, invasion and epithelial mesenchymal transition process. In conclusion, CENPI is overexpressed in CRC and functions as oncogene in modulating CRC cell migration, invasion and EMT process. Copyright © 2018. Published by Elsevier B.V.

Targeting colorectal cancer cells by a novel sphingosine kinase 1 inhibitor PF-543

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ju, TongFa; Gao, DaQuan; Fang, Zheng-yu, E-mail: fangzhengyu158@sina.com

In this study, we showed that PF-543, a novel sphingosine kinase 1 (SphK1) inhibitor, exerted potent anti-proliferative and cytotoxic effects against a panel of established (HCT-116, HT-29 and DLD-1) and primary human colorectal cancer (CRC) cells. Its sensitivity was negatively associated with SphK1 expression level in the CRC cells. Surprisingly, PF-543 mainly induced programmed necrosis, but not apoptosis, in the CRC cells. CRC cell necrotic death was detected by lactate dehydrogenase (LDH) release, mitochondrial membrane potential (MMP) collapse and mitochondrial P53-cyclophilin-D (Cyp-D) complexation. Correspondingly, the necrosis inhibitor necrostatin-1 largely attenuated PF-543-induced cytotoxicity against CRC cells. Meanwhile, the Cyp-D inhibitors (sanglifehrinmore » A and cyclosporin A), or shRNA-mediated knockdown of Cyp-D, remarkably alleviated PF-543-induced CRC cell necrotic death. Reversely, over-expression of wild-type Cyp-D in HCT-116 cells significantly increased PF-543's sensitivity. In vivo, PF-543 intravenous injection significantly suppressed HCT-116 xenograft growth in severe combined immunodeficient (SCID) mice, whiling remarkably improving the mice survival. The in vivo activity by PF-543 was largely attenuated when combined with the Cyp-D inhibitor cyclosporin A. Collectively, our results demonstrate that PF-543 exerts potent anti-CRC activity in vitro and in vivo. Mitochondrial programmed necrosis pathway is likely the key mechanism responsible for PF-543's actions in CRC cells. - Highlights: • PF-543 is anti-proliferative and cytotoxic to established and primary CRC cells. • PF-543 induces programmed necrosis, but not apoptosis, in CRC cells. • Modulation of mitochondrial protein cyclophilin-D alters PF-543's sensitivity. • PF-543 inhibits HCT-116 xenograft growth in SCID mice, improving mice survival. • Co-administration of cyclophilin-D inhibitor CsA inhibits PF-543's activity in vivo.« less

Generation, Characterization and Application of Antibodies Directed against HERV-H Gag Protein in Colorectal Samples.

PubMed

Mullins, Christina S; Hühns, Maja; Krohn, Mathias; Peters, Sven; Cheynet, Valérie; Oriol, Guy; Guillotte, Michèle; Ducrot, Sandrine; Mallet, François; Linnebacher, Michael

2016-01-01

A substantial part of the human genome originates from transposable elements, remnants of ancient retroviral infections. Roughly 8% of the human genome consists of about 400,000 LTR elements including human endogenous retrovirus (HERV) sequences. Mainly, the interplay between epigenetic and post-transcriptional mechanisms is thought to silence HERV expression in most physiological contexts. Interestingly, aberrant reactivation of several HERV-H loci appears specific to colorectal carcinoma (CRC). The expression of HERV-H Gag proteins (Gag-H) was assessed using novel monoclonal mouse anti Gag-H antibodies. In a flow cytometry screen four antibody clones were tested on a panel of primary CRC cell lines and the most well performing ones were subsequently validated in western blot analysis. Finally, Gag-H protein expression was analyzed by immune histology on cell line cytospins and on clinical samples. There, we found a heterogeneous staining pattern with no background staining of endothelial, stromal and infiltrating immune cells but diffuse staining of the cytoplasm for positive tumor and normal crypt cells of the colonic epithelium. Taken together, the Gag-H antibody clone(s) present a valuable tool for staining of cells with colonic origin and thus form the basis for future more detailed investigations. The observed Gag-H protein staining in colonic epithelium crypt cells demands profound analyses of a potential role for Gag-H in the normal physiology of the human gut.

Cancer stem cells in colorectal cancer: a review.

PubMed

Munro, Matthew J; Wickremesekera, Susrutha K; Peng, Lifeng; Tan, Swee T; Itinteang, Tinte

2018-02-01

Colorectal cancer (CRC) is the second most common cancer in women and the third most common in men. Adenocarcinoma accounts for 90% of CRC cases. There has been accumulating evidence in support of the cancer stem cell (CSC) concept of cancer which proposes that CSCs are central in the initiation of cancer. CSCs have been the focus of study in a range of cancers, including CRC. This has led to the identification and understanding of genes involved in the induction and maintenance of pluripotency of stem cells, and markers for CSCs, including those investigated specifically in CRC. Knowledge of the expression pattern of CSCs in CRC has been increasing in recent years, revealing a heterogeneous population of cells within CRC ranging from pluripotent to differentiated cells, with overlapping and sometimes unique combinations of markers. This review summarises current literature on the understanding of CSCs in CRC, including evidence of the presence of CSC subpopulations, and the stem cell markers currently used to identify and localise these CSC subpopulations. Future research into this field may lead to improved methods for early detection of CRC, novel therapy and monitoring of treatment for CRC and other cancer types. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Characterisation of a Cell Culture System for Investigating Nerve Agent Neurotoxicology. Part 1

DTIC Science & Technology

2012-03-01

eds. CRC Press), pp. 1-24. Sawyer,T.W., Weiss,M.T., and Unger,R.J. (1992). Anticholinesterase activity of organophosphate nerve agents in neuronal...were confirmed. The presence of muscarinic receptors and acetylcholinesterase activity was determined. Importantly, differential acetylcholinesterase... activity assays that will provide the basis for an ongoing research programme. The neuroblastoma cell lines chosen can potentially be used as a

Clinical value of circulating endothelial cell levels in metastatic colorectal cancer patients treated with first-line chemotherapy and bevacizumab.

PubMed

Malka, D; Boige, V; Jacques, N; Vimond, N; Adenis, A; Boucher, E; Pierga, J Y; Conroy, T; Chauffert, B; François, E; Guichard, P; Galais, M P; Cvitkovic, F; Ducreux, M; Farace, F

2012-04-01

We investigated whether circulating endothelial cells (CECs) predict clinical outcome of first-line chemotherapy and bevacizumab in metastatic colorectal cancer (mCRC) patients. In a substudy of the randomized phase II FNCLCC ACCORD 13/0503 trial, CECs (CD45- CD31+ CD146+ 7-amino-actinomycin- cells) were enumerated in 99 patients by four-color flow cytometry at baseline and after one cycle of treatment. We correlated CEC levels with objective response rate (ORR), 6-month progression-free survival (PFS) rate (primary end point of the trial), PFS, and overall survival (OS). Multivariate analyses of potential prognostic factors, including CEC counts and Köhne score, were carried out. By multivariate analysis, high baseline CEC levels were the only independent prognostic factor for 6-month PFS rate (P < 0.01) and were independently associated with worse PFS (P = 0.02). High CEC levels after one cycle were the only independent prognostic factor for ORR (P = 0.03). High CEC levels at both time points independently predicted worse ORR (P = 0.025), 6-month PFS rate (P = 0.007), and PFS (P = 0.02). Köhne score was the only variable associated with OS. CEC levels at baseline and after one treatment cycle may independently predict ORR and PFS in mCRC patients starting first-line bevacizumab and chemotherapy.

Introduction of hsa-miR-103a and hsa-miR-1827 and hsa-miR-137 as new regulators of Wnt signaling pathway and their relation to colorectal carcinoma.

PubMed

Fasihi, Ali; M Soltani, Bahram; Atashi, Amir; Nasiri, Shirzad

2018-07-01

Wnt signaling is hyper-activated in most of human cancers including colorectal carcinoma (CRC). Therefore, the introduction of new regulators for Wnt pathway possesses promising diagnostic and therapeutic applications in cancer medicine. Bioinformatics analysis introduced hsa-miR-103a, hsa-miR-1827, and hsa-miR-137 as potential regulators of Wnt signaling pathway. Here, we intended to examine the effect of these human miRNAs on Wnt signaling pathway components, on the cell cycle progression in CRC originated cell lines and their expression in CRC tissues. RT-qPCR results indicated upregulation of hsa-miR-103a, hsa-miR-1827, and downregulation of hsa-miR-137 in CRC tissues. Overexpression of hsa-miR-103a and hsa-miR-1827 in SW480 cells resulted in elevated Wnt activity, detected by both Top/Flash assay and RT-qPCR analysis. Inhibition of Wnt signaling by using PNU-74654 or IWP-2 small molecules suggested that these miRNAs exerts their effect at the β-catenin degradation complex level. Then, RT-qPCR, dual luciferase assay, and western blotting analysis indicated that APC and APC2 transcripts were targeted by hsa-miR-103a, hsa-miR-1827 while, Wnt3a and β-catenin genes were upregulated. However, hsa-miR-137 downregulated Wnt3a and β-catenin genes. Further, hsa-miR-103a and hsa-miR-1827 overexpression resulted in cell cycle progression and reduced apoptotic rate in SW480 cells, unlike hsa-miR-137 overexpression which resulted in cell cycle suppression, detected by flowcytometry and Anexin analysis. Overall, our data introduced hsa-miR-103a, hsa-miR-1827 as onco-miRNAs and hsa-miR-137 as tumor suppressor which exert their effect through regulation of Wnt signaling pathway in CRC and introduced them as potential target for therapy. © 2017 Wiley Periodicals, Inc.

NR2F2 inhibits Smad7 expression and promotes TGF-β-dependent epithelial-mesenchymal transition of CRC via transactivation of miR-21.

PubMed

Wang, Hao; Nie, Lei; Wu, Lei; Liu, Qiufang; Guo, Xueyan

2017-03-25

Metastasis is one of the most decisive factors influencing CRC patient prognosis and current studies suggest that a molecular mechanism known as EMT broadly regulates cancer metastasis. NR2F2 is a key molecule in the development of CRC, but the roles and underlying mechanisms of NR2F2 in TGF-β induced EMT in CRC remain largely unknown. In the current study, we were interested to examine the role of NR2F2 in the TGF-β-induced EMT in CRC. Here, we found NR2F2 was upregulated in CRC cells and promotes TGF-β-induced EMT in CRC. Using comparative miRNA profiling TGF-β pre-treated CRC cells in which NR2F2 had been knocked down with that of control cells, we identified miR-21 as a commonly downregulated miRNA in HT29 cells treated with TGF-β and NR2F2 siRNA, and its downregulation inhibiting migration and invasion of CRC cells. Moreover, we found NR2F2 could transcriptional activated miR-21 expression by binding to miR-21 promoter in HT29 by ChIP and luciferase assay. In the last, our data demonstrated that Smad7 was the direct target of miR-21 in CRC cells. Thus, NR2F2 could promote TGF-β-induced EMT and inhibit Smad7 expression via transactivation of miR-21, and NR2F2 may be a new common therapeutic target for CRC. Copyright © 2017 Elsevier Inc. All rights reserved.

UCN-01 enhances cytotoxicity of irinotecan in colorectal cancer stem-like cells by impairing DNA damage response

PubMed Central

Pilozzi, Emanuela; De Luca, Gabriele; Cappellari, Marianna; Fanciulli, Maurizio; Goeman, Frauke; Melucci, Elisa; Biffoni, Mauro; Ricci-Vitiani, Lucia

2016-01-01

Colorectal cancer (CRC) is one of the most common and lethal cancers worldwide. Despite recent progress, the prognosis of advanced stage CRC remains poor, mainly because of cancer recurrence and metastasis. The high morbidity and mortality of CRC has been recently ascribed to a small population of tumor cells that hold the potential of tumor initiation, i.e. cancer stem cells (CSCs), which play a pivotal role in cancer recurrence and metastasis and are not eradicated by current therapy. We screened CRC-SCs in vitro with a library of protein kinase inhibitors and showed that CRC-SCs are resistant to specific inhibition of the major signaling pathways involved in cell survival and proliferation. Nonetheless, broad-spectrum inhibition by the staurosporin derivative UCN-01 blocks CRC-SC growth and potentiates the activity of irinotecan in vitro and in vivo CRC-SC-derived models. Reverse-Phase Protein Microarrays (RPPA) revealed that, albeit CRC-SCs display individual phospho-proteomic profiles, sensitivity of CRC-SCs to UCN-01 relies on the interference with the DNA damage response mediated by Chk1. Combination of LY2603618, a specific Chk1/2 inhibitor, with irinotecan resulted in a significant reduction of CRC-SC growth in vivo, confirming that irinotecan treatment coupled to inhibition of Chk1 represents a potentially effective therapeutic approach for CRC treatment. PMID:27286453

DcR3 induces epithelial-mesenchymal transition through activation of the TGF-β3/SMAD signaling pathway in CRC.

PubMed

Liu, Yan-Ping; Zhu, Hui-Fang; Liu, Ding-Li; Hu, Zhi-Yan; Li, Sheng-Nan; Kan, He-Ping; Wang, Xiao-Yan; Li, Zu-Guo

2016-11-22

Decoy receptor 3 (DcR3), a novel member of the tumor necrosis factor receptor (TNFR) family, was recently reported to be associated with tumorigenesis and metastasis. However, the role of DcR3 in human colorectal cancer (CRC) has not been fully elucidated. In this study, we found that DcR3 expression was significantly higher in human colorectal cancer tissues than in paired normal tissues, and that DcR3 expression was strongly correlated with tumor invasion, lymph node metastases and poor prognoses. Moreover, DcR3 overexpression significantly enhanced CRC cell proliferation and migration in vitro and tumorigenesis in vivo. Conversely, DcR3 knockdown significantly repressed CRC cell proliferation and migration in vitro, and DcR3 deficiency also attenuated CRC tumorigenesis and metastasis in vivo. Functionally, DcR3 was essential for TGF-β3/SMAD-mediated epithelial-mesenchymal transition (EMT) of CRC cells. Importantly, cooperation between DcR3 and TGF-β3/SMAD-EMT signaling-related protein expression was correlated with survival and survival time in CRC patients. In conclusion, our results demonstrate that DcR3 may be a prognostic biomarker for CRC and that this receptor facilitates CRC development and metastasis by participating in TGF-β3/SMAD-mediated EMT of CRC cells.

Optimising the use of cetuximab in the continuum of care for patients with metastatic colorectal cancer.

PubMed

Goldberg, Richard M; Montagut, Clara; Wainberg, Zev A; Ronga, Philippe; Audhuy, François; Taieb, Julien; Stintzing, Sebastian; Siena, Salvatore; Santini, Daniele

2018-01-01

The anti-epidermal growth factor receptor (EGFR) monoclonal antibody cetuximab in combination with chemotherapy is a standard of care in the first-line treatment of RAS wild-type (wt) metastatic colorectal cancer (mCRC) and has demonstrated efficacy in later lines. Progressive disease (PD) occurs when tumours develop resistance to a therapy, although controversy remains about whether PD on a combination of chemotherapy and targeted agents implies resistance to both components. Here, we propose that some patients may gain additional clinical benefit from the reuse of cetuximab after having PD on regimens including cetuximab in an earlier treatment line. We conducted a non-systematic literature search in PubMed and reviewed published and ongoing clinical trials, focusing on later-line cetuximab reuse in patients with mCRC. Evidence from multiple studies suggests that cetuximab can be an efficacious and tolerable treatment when continued or when fit patients with mCRC are retreated with it after a break from anti-EGFR therapy. Furthermore, on the basis of available preclinical and clinical evidence, we propose that longitudinal monitoring of RAS status may identify patients suitable for such a strategy. Patients who experience progression on cetuximab plus chemotherapy but have maintained RAS wt tumour status may benefit from continuation of cetuximab with a chemotherapy backbone switch because they have probably developed resistance to the chemotherapeutic agents rather than the biologic component of the regimen. Conversely, patients whose disease progresses on cetuximab-based therapy due to drug-selected clonal expansion of RAS- mutant tumour cells may regain sensitivity to cetuximab following a defined break from anti-EGFR therapy. Looking to the future, we propose that RAS status determination at disease progression by liquid, needle or excisional biopsy may identify patients eligible for cetuximab continuation and rechallenge. With this approach, treatment benefit can be extended, adding to established continuum-of-care strategies in patients with mCRC.

Antibody validation and scoring guidelines for ABCG2 immunohistochemical staining in formalin-fixed paraffin-embedded colon cancer tissue

PubMed Central

Cederbye, Camilla Natasha; Palshof, Jesper Andreas; Hansen, Tine Plato; Duun-Henriksen, Anne Katrine; Linnemann, Dorte; Stenvang, Jan; Nielsen, Dorte Lisbet; Brünner, Nils; Viuff, Birgitte Martine

2016-01-01

Overexpression of the ATP-dependent drug efflux pump ABCG2 is a major molecular mechanism of multidrug resistance in cancer and might be a predictive biomarker for drug response. Contradictory results have been reported for immunohistochemical studies of ABCG2 protein expression in colorectal cancer (CRC), probably because of the use of different antibodies and scoring approaches. In this study, we systematically studied six commercially available anti-ABCG2 antibodies, using cell lines with up-regulation of ABCG2, and selected one antibody for validation in CRC tissue. Furthermore, we established scoring guidelines for ABCG2 expression based on the clinically used guidelines for HER2 immunohistochemistry assessment in gastric cancer. The guidelines provide a semi-quantitative measure of the basolateral membrane staining of ABCG2 and disregard the apical membrane staining and the cytoplasmic signal. Intra-tumor heterogeneity in ABCG2 immunoreactivity was observed; however, statistical analyses of tissue microarrays (TMAs) and the corresponding whole sections from primary tumors of 57 metastatic CRC patients revealed a strong positive correlation between maximum TMA scores and whole sections, especially when more than one core was used. In conclusion, here, we provide validated results to guide future studies on the associations between ABCG2 immunoreactivity in tumor cells and the benefits of chemotherapeutic treatment in patients with CRC. PMID:27257141

Maintenance of cancer stemness by miR-196b-5p contributes to chemoresistance of colorectal cancer cells via activating STAT3 signaling pathway

PubMed Central

Zhang, Xin; Peng, Yao; Ye, Ziyu; Ma, Yan; Liang, Yangfang; Cao, Longbin; Li, Xiangyong; Li, Ronggang; Sun, Lixia; Liu, Qiongru; Wu, Jinhua; Zhou, Keyuan; Zeng, Jincheng

2017-01-01

Emerging studies indicated that cancer stem cells represent a subpopulation of cells within the tumor that is responsible for chemotherapeutic resistance. However, the underlying mechanism is still not clarified yet. Here we report that miR-196b-5p is dramatically upregulated in CRC tissues and high expression of miR-196b-5p correlates with poor survival in CRC patients. Moreover, recurrent gains (amplification) contribute to the miR-196b-5p overexpression in CRC tissues. Silencing miR-196b-5p suppresses spheroids formation ability, the fraction of SP cells, expression of stem cell factors and the mitochondrial potential, and enhances the apoptosis induced by 5-fluorouracil in CRC cells; while ectopic expression of miR-196b-5p yields an opposite effect. In addition, downregulation of miR-196b-5p resensitizes CRC cells to 5-fluorouracil in vivo. Our results further demonstrate that miR-196b-5p promotes stemness and chemoresistance of CRC cells to 5-fluorouracil via targeting negative regulators SOCS1 and SOCS3 of STAT3 signaling pathway, giving rise to activation of STAT3 signaling. Interestingly, miR-196b-5p is highly enriched in the serum exosomes of patients with CRC compared to the healthy control subjects. Thus, our results unravel a novel mechanism of miR-196b-5p implicating in the maintenance of stem cell property and chemotherapeutic resistance in CRC, offering a potential rational registry of anti-miR-196b-5p combining with conventional chemotherapy against CRC. PMID:28591704

Prohibitin, relocated to the front ends, can control the migration directionality of colorectal cancer cells

PubMed Central

Guo, Li-Li; Hu, Chun-Ting; Huang, Ying-Xin; Huang, Guan; Jing, Fang-Yan; Liu, Chao; Li, Zhuo-Yi; Zhou, Na; Yan, Qian-Wen; Lei, Yan; Zhu, Shi-Jie; Cheng, Zhi-Qiang; Cao, Guang-Wen; Deng, Yong-Jian; Ding, Yan-Qing

2017-01-01

Directional migration is a cost-effective movement allowing invasion and metastatic spread of cancer cells. Although migration related to cytoskeletal assembly and microenvironmental chemotaxis has been elucidated, little is known about interaction between extracellular and intracellular molecules for controlling the migrational directionality. A polarized expression of prohibitin (PHB) in the front ends of CRC cells favors metastasis and is correlated with poor prognosis for 545 CRC patients. A high level of vascular endothelial growth factor (VEGF) in the interstitial tissue of CRC patients is associated with metastasis. VEGF bound to its receptor, neuropilin-1, can stimulate the activation of cell division cycle 42, which recruits intra-mitochondrial PHB to the front end of a CRC cell. This intracellular relocation of PHB results in the polymerization and reorganization of filament actin extending to the front end of the cell. As a result, the migration directionality of CRC cells is targeted towards VEGF. Together, these findings identify PHB as a key modulator of directional migration of CRC cells and a target for metastasis. PMID:29100316

Long non-coding RNA MIAT promotes growth and metastasis of colorectal cancer cells through regulation of miR-132/Derlin-1 pathway.

PubMed

Liu, Zhaoxia; Wang, Hai; Cai, Hongwei; Hong, Ye; Li, Yan; Su, Dongming; Fan, Zhining

2018-01-01

Recently, long non-coding RNA (lncRNA) MIAT has been demonstrated as an oncogenic gene in several types of cancer. However, the role and mechanism of MIAT in colorectal cancer (CRC) have not been investigated. Real-time PCR was used to measure MIAT expression in CRC tissues and cells. Small interfering RNA specific for MIAT (si-MIAT) was used to down-regulate MIAT expression in CRC cells. The interaction of MIAT and miR-132 was measured by RNA pull-down assay. The effect of si-MIAT on CRC cells apoptosis and metastasis were measured by flow cytometry assay, invasion and migration assay, respectively. In present study, we found that MIAT was highly expressed in CRC tissues and cells. MIAT knockdown inhibited proliferation, migration and invasion and enhanced apoptosis of CRC cells. Further, we demonstrated that MIAT acted as a competing endogenous RNA for miR-132, antagonized its functions, and resulted in the de-repression of its target gene Derlin-1, which acted as an oncogene in promoting growth and metastasis of CRC cells. In LOVO and SW480 cells with si-MIAT, miR-132 inhibitor resulted in an increase of cell proliferation, migration and invasion and a decrease of cell apoptosis, which was partially abolished by transfection of Derlin-1 shRNA. Our data indicated that highly expressed MIAT was an oncogenic lncRNA that promoted the growth and metastasis of CRC through miR-132/Derlin-1 axis.

The β-catenin E3 ubiquitin ligase SIAH-1 is regulated by CSN5/JAB1 in CRC cells.

PubMed

Jumpertz, Sandra; Hennes, Thomas; Asare, Yaw; Vervoorts, Jörg; Bernhagen, Jürgen; Schütz, Anke K

2014-09-01

COP9 signalosome subunit 5 (CSN5) plays a decisive role in cellular processes such as cell cycle regulation and apoptosis via promoting protein degradation, gene transcription, and nuclear export. CSN5 regulates cullin-RING-E3 ligase (CRL) activity through its deNEDDylase function. It is overexpressed in several tumor entities, but its role in colorectal cancer (CRC) is poorly understood. Wnt/β-catenin signaling is aberrant in most CRC cells, resulting in increased levels of oncogenic β-catenin and thus tumor progression. Under physiological conditions, β-catenin levels are tightly regulated by continuous proteasomal degradation. We recently showed that knockdown of CSN5 in model and CRC cells results in decreased (phospho)-β-catenin levels. Reduced β-catenin levels were associated with an attenuated proliferation rate of different CRC cell types after CSN5 knockdown. The canonical Wnt pathway involves degradation of β-catenin by a β-TrCP1-containing E3 ligase, but is mostly non-functional in CRC cells. We thus hypothesized that alternative β-catenin degradation mediated by SIAH-1 (seven in absentia homolog-1), is responsible for the effect of CSN5 on β-catenin signaling in CRC cells. We found that SIAH-1 plays an essential role in β-catenin degradation in HCT116 CRC cells and that CSN5 affects β-catenin target gene expression in these cells. Of note, CSN5 affected SIAH-1 mRNA and SIAH-1 protein levels. Moreover, β-catenin and SIAH-1 form protein complexes with CSN5 in HCT116 cells. Lastly, we demonstrate that CSN5 promotes SIAH-1 degradation in HCT116 and SW480 cells and that this is associated with its deNEDDylase activity. In conclusion, we have identified a CSN5/β-catenin/SIAH-1 interaction network that might control β-catenin degradation in CRC cells. Copyright © 2014 Elsevier Inc. All rights reserved.

RGC32 induces epithelial-mesenchymal transition by activating the Smad/Sip1 signaling pathway in CRC.

PubMed

Wang, Xiao-Yan; Li, Sheng-Nan; Zhu, Hui-Fang; Hu, Zhi-Yan; Zhong, Yan; Gu, Chuan-Sha; Chen, Shi-You; Liu, Teng-Fei; Li, Zu-Guo

2017-05-04

Response gene to complement 32 (RGC32) is a transcription factor that regulates the expression of multiple genes involved in cell growth, viability and tissue-specific differentiation. However, the role of RGC32 in tumorigenesis and tumor progression in colorectal cancer (CRC) has not been fully elucidated. Here, we showed that the expression of RGC32 was significantly up-regulated in human CRC tissues versus adjacent normal tissues. RGC32 expression was significantly correlated with invasive and aggressive characteristics of tumor cells, as well as poor survival of CRC patients. We also demonstrated that RGC32 overexpression promoted proliferation, migration and tumorigenic growth of human CRC cells in vitro and in vivo. Functionally, RGC32 facilitated epithelial-mesenchymal transition (EMT) in CRC via the Smad/Sip1 signaling pathway, as shown by decreasing E-cadherin expression and increasing vimentin expression. In conclusion, our findings suggested that overexpression of RGC32 facilitates EMT of CRC cells by activating Smad/Sip1 signaling.

Vascular endothelial growth factor receptor-1 mediates migration of human colorectal carcinoma cells by activation of Src family kinases

PubMed Central

Lesslie, D P; Summy, J M; Parikh, N U; Fan, F; Trevino, J G; Sawyer, T K; Metcalf, C A; Shakespeare, W C; Hicklin, D J; Ellis, L M; Gallick, G E

2006-01-01

Vascular endothelial growth factor (VEGF) is the predominant pro-angiogenic cytokine in human malignancy, and its expression correlates with disease recurrence and poor outcomes in patients with colorectal cancer. Recently, expression of vascular endothelial growth factor receptors (VEGFRs) has been observed on tumours of epithelial origin, including those arising in the colon, but the molecular mechanisms governing potential VEGF-driven biologic functioning in these tumours are not well characterised. In this report, we investigated the role of Src family kinases (SFKs) in VEGF-mediated signalling in human colorectal carcinoma (CRC) cell lines. Vascular endothelial growth factor specifically activated SFKs in HT29 and KM12L4 CRC cell lines. Further, VEGF stimulation resulted in enhanced cellular migration, which was effectively blocked by pharmacologic inhibition of VEGFR-1 or Src kinase. Correspondingly, migration studies using siRNA clones with reduced Src expression confirmed the requirement for Src in VEGF-induced migration in these cells. Furthermore, VEGF treatment enhanced VEGFR-1/SFK complex formation and increased tyrosine phosphorylation of focal adhesion kinase, p130 cas and paxillin. Finally, we demonstrate that VEGF-induced migration is not due, at least in part, to VEGF acting as a mitogen. These results suggest that VEGFR-1 promotes migration of tumour cells through a Src-dependent pathway linked to activation of focal adhesion components that regulate this process. PMID:16685275

SUMOylated MAFB promotes colorectal cancer tumorigenesis

PubMed Central

Xie, Yin-Yin; Sun, Xiao-Jian; Zhao, Ren; Huang, Qiu-Hua

2016-01-01

The transcription factor, v-maf avian musculoaponeurotic fibrosarcoma oncogene homolog B (MAFB), promotes tumorigenesis in some cancers. In this study, we found that MAFB levels were increased in clinical colorectal cancer (CRC) samples, and higher expression correlated with more advanced TNM stage. We identified MAFB amplifications in a majority of tumor types in an assessment of The Cancer Genome Atlas database. Altered MAFB levels due to gene amplification, deletion, mutation, or transcription upregulation occurred in 9% of CRC cases within the database. shRNA knockdown experiments demonstrated that MAFB deficiency blocked CRC cell proliferation by arresting the cell cycle at G0/G1 phase in vitro. We found that MAFB could be SUMOylated by SUMO1 at lysine 32, and this modification was critical for cell cycle regulation by MAFB in CRC cells. SUMOylated MAFB directly regulated cyclin-dependent kinase 6 transcription by binding to its promoter. MAFB knockdown CRC cell xenograft tumors in mice grew more slowly than controls, and wild-type MAFB-overexpressing tumors grew more quickly than tumors overexpressing MAFB mutated at lysine 32. These data suggest that SUMOylated MAFB promotes CRC tumorigenesis through cell cycle regulation. MAFB and its SUMOylation process may serve as novel therapeutic targets for CRC treatment. PMID:27829226

ITGBL1 promotes migration, invasion and predicts a poor prognosis in colorectal cancer.

PubMed

Qiu, Xiao; Feng, Jue-Rong; Qiu, Jun; Liu, Lan; Xie, Yang; Zhang, Yu-Peng; Liu, Jing; Zhao, Qiu

2018-05-14

Colorectal cancer (CRC) is one of the most common malignancies worldwide; its progression and prognosis are associated with oncogenes. The present study aimed to identify differentially expressed genes (DEGs) and explore the role and potential mechanism of integrin subunit β like 1 (ITGBL1) in CRC. The microarray dataset GSE41258 was used to screen DEGs involved in CRC. Survival analysis was performed to predict the prognosis of CRC patients. To validate ITGBL1 expression, immunohistochemistry, quantitative real-time PCR and western blotting were performed in CRC tissues and cells. Subsequently, the effects of ITGBL1 were evaluated through colony formation, cell proliferation, migration and invasion assays. Finally, we took advantage of Gene Ontology (GO) analysis and Gene Set Enrichment Analysis (GSEA) to explore potential function and mechanism of ITGBL1 in CRC. In our study, 182 primary CRC tissues and 54 normal colon tissues were contained in GSE41258 dataset. A total of 318 DEGs were screened, among which ITGBL1 was found to be significantly up-regulated in CRC, and its high expression was associated with shortened survival of CRC patients. Moreover, knockdown of ITGBL1 promoted CRC cell proliferation, migration and invasion. Finally, GO analysis revealed that ITGBL1 was associated with cell adhesion. GSEA indicated that ITGBL1 was enriched in ECM receptor interaction and focal adhesion. In conclusion, a novel oncogene ITGBL1 was identified and demonstrated to be associated with the progression and prognosis of CRC, which might be a potential therapeutic target and prognostic biomarker for CRC patients. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Periostin expression in intra-tumoral stromal cells is prognostic and predictive for colorectal carcinoma via creating a cancer-supportive niche

PubMed Central

Tan, Xiaojie; Ding, Yibo; Luo, Yanxin; Cai, Hui; Liu, Yan; Gao, Xianhua; Liu, Qizhi; Yu, Yongwei; Du, Yan; Wang, Hao; Ma, Liye; Wang, Jianping; Chen, Kun; Ding, Yanqing; Fu, Chuangang; Cao, Guangwen

2016-01-01

Periostin (POSTN) expression in cancer cells and circulation has been related to poor prognosis of colorectal carcinoma (CRC). However, the role of POSTN expressed in intra-tumoral stroma on CRC progression remains largely unknown. This study enrolled 1098 CRC patients who received surgical treatment in Shanghai and Guangzhou, Mainland China. In Shanghai cohort, immunohistochemistry score of stromal POSTN expression increased consecutively from adjacent mucosa, primary CRC tissues, to metastatic CRC tissues (P < 0.001), while medium- and high-stromal POSTN expression, rather than epithelial POSTN expression, independently predicted unfavorable prognoses of CRC, adjusted for covariates including TNM stage and postoperative chemotherapy in multivariate Cox models. The results in Shanghai cohort were faithfully replicated in Guangzhou cohort. Stromal POSTN expression dose-dependently predicted an unfavorable prognosis of stage III CRC patients with postoperative chemotherapy in both cohorts. POSTN derived from colonic fibroblasts or recombinant POSTN significantly promoted proliferation, anchorage independent growth, invasion, and chemo-resistance of CRC cells; whereas these effects were counteracted via targeting to PI3K/Akt or Wnt/β-catenin signaling pathway. CRC cell RKO-derived factor(s) significantly induced POSTN production in colonic fibroblasts and autocrine POSTN promoted proliferation, migration, and anchorage independent growth of fibroblasts. Conclusively, stromal POSTN is prognostic and predictive for CRC via creating a niche to facilitate cancer progression. Targeting POSTN-induced signaling pathways may be therapeutic options for metastatic or chemoresistant CRC. PMID:26556874

DcR3 induces epithelial-mesenchymal transition through activation of the TGF-β3/SMAD signaling pathway in CRC

PubMed Central

Hu, Zhi-Yan; Li, Sheng-Nan; Kan, He-Ping; Wang, Xiao-Yan; Li, Zu-Guo

2016-01-01

Decoy receptor 3 (DcR3), a novel member of the tumor necrosis factor receptor (TNFR) family, was recently reported to be associated with tumorigenesis and metastasis. However, the role of DcR3 in human colorectal cancer (CRC) has not been fully elucidated. In this study, we found that DcR3 expression was significantly higher in human colorectal cancer tissues than in paired normal tissues, and that DcR3 expression was strongly correlated with tumor invasion, lymph node metastases and poor prognoses. Moreover, DcR3 overexpression significantly enhanced CRC cell proliferation and migration in vitro and tumorigenesis in vivo. Conversely, DcR3 knockdown significantly repressed CRC cell proliferation and migration in vitro, and DcR3 deficiency also attenuated CRC tumorigenesis and metastasis in vivo. Functionally, DcR3 was essential for TGF-β3/SMAD-mediated epithelial-mesenchymal transition (EMT) of CRC cells. Importantly, cooperation between DcR3 and TGF-β3/SMAD-EMT signaling-related protein expression was correlated with survival and survival time in CRC patients. In conclusion, our results demonstrate that DcR3 may be a prognostic biomarker for CRC and that this receptor facilitates CRC development and metastasis by participating in TGF-β3/SMAD-mediated EMT of CRC cells. PMID:27764793

miR-133b down-regulates ABCC1 and enhances the sensitivity of CRC to anti-tumor drugs.

PubMed

Chen, Miao; Li, Daojiang; Gong, Ni; Wu, Hao; Su, Chen; Xie, Canbin; Xiang, Hong; Lin, Changwei; Li, Xiaorong

2017-08-08

Multidrug resistance (MDR) is the main cause of failed chemotherapy treatments. Therefore, preventing MDR is pivotal in treating colorectal cancer (CRC). In a previous study miR-133b was shown to be a tumor suppressor. Additionally, in CRC cells transfected with miR-133b, ATP-binding cassette (ABC) subfamily C member 1(ABCC1) was shown to be significantly down regulated. Whether miR-133b also enhances the chemosensitivity of drugs used to treat CRC by targeting ABCC1 is still unclear. Here, we utilized flow cytometry and high-performance liquid chromatography (HPLC) analysis to identify the ability of miR-133b to reserve MDR in CRC. We then used a dual-luciferase reporter assay to validate that miR-133b targets ABCC1. Further in vivo experiments were designed to validate the method in which miR-133b reversed MDR in CRC cells. The results demonstrated that the level of miR-133b was down-regulated and the expression of ABCC1 was up-regulated in drug-resistant CRC cells compared to non-drug-resistant CRC cells. The restoration of miR-133b expression in CRC drug-resistant cells in vitro resulted in reduced IC50s to chemotherapeutic drugs, significantly induced G1 accumulation, inhibited growth and promoted necrosis in combination with either 5-fluorouracil (5-FU) or vincristine (VCR), and decreased the expression of ABCC1. The dual-luciferase assay demonstrated that miR-133b directly targets ABCC1. The combination of agomiRNA-133b with chemotherapeutic drugs in vivo inhibited tumor growth induced by CRC drug-resistant cells. A xenograft from the in vivo model resulted in up-regulated levels of miR-133b and down-regulated levels of ABCC1. Therefore, miR-133b enhances the chemosensitivity of CRC cells to anti-tumor drugs by directly down-regulating ABCC1. This discovery provides a therapeutic strategy in which miR-133b is used as a potential sensitizer for drug-resistant CRC.

Loss of p53-inducible long non-coding RNA LINC01021 increases chemosensitivity

PubMed Central

Kaller, Markus; Götz, Ursula; Hermeking, Heiko

2017-01-01

We have previously identified the long non-coding RNA LINC01021 as a direct p53 target (Hünten et al. Mol Cell Proteomics. 2015; 14:2609-2629). Here, we show that LINC01021 is up-regulated in colorectal cancer (CRC) cell lines upon various p53-activating treatments. The LINC01021 promoter and the p53 binding site lie within a MER61C LTR, which originated from insertion of endogenous retrovirus 1 (ERV1) sequences. Deletion of this MER61C element by a CRISPR/Cas9 approach, as well as siRNA-mediated knockdown of LINC01021 RNA significantly enhanced the sensitivity of the CRC cell line HCT116 towards the chemotherapeutic drugs doxorubicin and 5-FU, suggesting that LINC01021 is an integral part of the p53-mediated response to DNA damage. Inactivation of LINC01021 and also its ectopic expression did not affect p53 protein expression and transcriptional activity, implying that LINC01021 does not feedback to p53. Furthermore, in CRC patient samples LINC01021 expression positively correlated with a wild-type p53-associated gene expression signature. LINC01021 expression was increased in primary colorectal tumors and displayed a bimodal distribution that was particularly pronounced in the mesenchymal CMS4 consensus molecular subtype of CRCs. CMS4 tumors with low LINC01021 expression were associated with poor patient survival. Our results suggest that the genomic redistribution of ERV1-derived p53 response elements and generation of novel p53-inducible lncRNA-encoding genes was selected for during primate evolution as integral part of the cellular response to various forms of genotoxic stress. PMID:29262524

MicroRNA-146a promote cell migration and invasion in human colorectal cancer via carboxypeptidase M/src-FAK pathway

PubMed Central

Zhan, Cheng; Le-Meng, Zhang; Liu, Hongchun; Cai, Yu; Tu, Chuantao; Li, Xi; Zou, Yanting; Zhang, Shuncai

2017-01-01

Colorectal cancer (CRC) is one of the most common cancers worldwide, and microRNAs play important roles in CRC progression. This study aimed to investigate the roles of miR-146a-5p in human CRC and their molecular mechanisms. First, we found that miR-146a-5p was significantly upregulated in CRC tissues and promoted the migration of CRC cells. Then, we identified carboxypeptidase M (CPM) as a direct target of miR-146a-5p, and found that it inhibited the migration and invasion of CRC cells. Our results also showed that CPM expression was positively correlated with overall survival and negatively correlated with recurrence, lymph node invasion, and N stage. Furthermore, we demonstrated that both miR-146a-5p and CPM regulated Src and FAK expression, while the Src-FAK signaling pathway is widely known to be associated with the migration and invasion of multiple tumor cells. This study is the first to demonstrate the functional and mechanistic relationship of the miR-146a-5p/CPM/Src-FAK axis and its effect on the migration and invasion of CRC cells. Thus, miR-146a-5p represents potential targets for CRC diagnosis and therapy. PMID:28186967

Selective targeting of KRAS-Mutant cells by miR-126 through repression of multiple genes essential for the survival of KRAS-Mutant cells

PubMed Central

Hara, Toshifumi; Jones, Matthew F.; Subramanian, Murugan; Li, Xiao Ling; Ou, Oliver; Zhu, Yuelin; Yang, Yuan; Wakefield, Lalage M.; Hussain, S. Perwez; Gaedcke, Jochen; Ried, Thomas; Luo, Ji; Caplen, Natasha J.; Lal, Ashish

2014-01-01

MicroRNAs (miRNAs) regulate the expression of hundreds of genes. However, identifying the critical targets within a miRNA-regulated gene network is challenging. One approach is to identify miRNAs that exert a context-dependent effect, followed by expression profiling to determine how specific targets contribute to this selective effect. In this study, we performed miRNA mimic screens in isogenic KRAS-Wild-type (WT) and KRAS-Mutant colorectal cancer (CRC) cell lines to identify miRNAs selectively targeting KRAS-Mutant cells. One of the miRNAs we identified as a selective inhibitor of the survival of multiple KRAS-Mutant CRC lines was miR-126. In KRAS-Mutant cells, miR-126 over-expression increased the G1 compartment, inhibited clonogenicity and tumorigenicity, while exerting no effect on KRAS-WT cells. Unexpectedly, the miR-126-regulated transcriptome of KRAS-WT and KRAS-Mutant cells showed no significant differences. However, by analyzing the overlap between miR-126 targets with the synthetic lethal genes identified by RNAi in KRAS-Mutant cells, we identified and validated a subset of miR-126-regulated genes selectively required for the survival and clonogenicity of KRAS-Mutant cells. Our strategy therefore identified critical target genes within the miR-126-regulated gene network. We propose that the selective effect of miR-126 on KRAS-Mutant cells could be utilized for the development of targeted therapy for KRAS mutant tumors. PMID:25245095

Interplay between long noncoding RNA ZEB1-AS1 and miR-101/ZEB1 axis regulates proliferation and migration of colorectal cancer cells.

PubMed

Xiong, Wan-Cheng; Han, Na; Wu, Nan; Zhao, Ke-Lei; Han, Chen; Wang, Hui-Xin; Ping, Guan-Fang; Zheng, Peng-Fei; Feng, Hailong; Qin, Lei; He, Peng

2018-01-01

Long noncoding RNAs (lncRNAs) are dysregulated in many diseases. MicroRNA-101 (miR-101) functions as a tumor suppressor by directly targeting ZEB1 in various cancers. However, the potential mechanism of lncRNA ZEB1-AS1 and miR-101/ZEB1 axis in CRC remains unknown. In this study, we further investigated the potential interplay between miR-101/ZEB1 axis and lncRNA ZEB1-AS1 in colorectal cancer (CRC). Results showed that ZEB1-AS1 was upregulated in CRC tissues and cells. MiR-101 was downregulated in CRC tissues and negatively correlated with ZEB1-AS1 and ZEB1 expression levels in CRC. Functional experiments showed that, consistent with ZEB1-AS1 depletion, miR-101 overexpression and ZEB1 depletion inhibited the proliferation and migration of CRC cells. Overexpression of miR-101 partially abolished the effects of ZEB1-AS1 on the proliferation and migration of these cells. Moreover, combined ZEB1-AS1 depletion and miR-101 overexpression significantly inhibited cell proliferation and migration of the CRC cells. Hence, ZEB1-AS1 functioned as a molecular sponge for miR-101 and relieved the inhibition of ZEB1 caused by miR-101. This study revealed a novel regulatory mechanism between ZEB1-AS1 and miR-101/ZEB1 axis. The interplay between ZEB1-AS1 and miR-101/ZEB1 axis contributed to the proliferation and migration of CRC cells, and targeting this interplay could be a promising strategy for CRC treatment.

R-spondin1/Wnt-enhanced Ascl2 autoregulation controls the self-renewal of colorectal cancer progenitor cells.

PubMed

Ye, Jun; Liu, Shanxi; Shang, Yangyang; Chen, Haoyuan; Wang, Rongquan

2018-06-25

The Wnt signaling pathway controls stem cell identity in the intestinal epithelium and cancer stem cells (CSCs). The transcription factor Ascl2 (Wnt target gene) is fate decider of intestinal cryptic stem cells and colon cancer stem cells. It is unclear how Wnt signaling is translated into Ascl2 expression and keeping the self-renewal of CRC progenitor cells. We showed that the exogenous Ascl2 in colorectal cancer (CRC) cells activated the endogenous Ascl2 expression via a direct autoactivatory loop, including Ascl2 binding to its own promoter and further transcriptional activation. Higher Ascl2 expression in human CRC cancerous tissues led to greater enrichment in Ascl2 immunoprecipitated DNA within the Ascl2 promoter in the CRC cancerous sample than the peri-cancerous mucosa. Ascl2 binding to its own promoter and inducing further transcriptional activation of the Ascl2 gene was predominant in the CD133 + CD44 + CRC population. R-spondin1/Wnt activated Ascl2 expression dose-dependently in the CD133 + CD44 + CRC population, but not in the CD133 - CD44 - CRC population, which was caused by differences in Ascl2 autoregulation under R-spondin1/Wnt activation. R-spondin1/Wnt treatment in the CD133 + CD44 + or CRC CD133 - CD44 - populations exerted a different pattern of stemness maintenance, which was defined by alterations of the mRNA levels of stemness-associated genes, the protein expression levels (Bmi1, C-myc, Oct-4 and Nanog) and tumorsphere formation. The results indicated that Ascl2 autoregulation formed a transcriptional switch that was enhanced by Wnt signaling in the CD133 + CD44 + CRC population, thus conferring their self-renewal.

Promoter hypermethylation and downregulation of the FAS gene may be involved in colorectal carcinogenesis

PubMed Central

MANOOCHEHRI, MEHDI; BORHANI, NASIM; KARBASI, ASHRAF; KOOCHAKI, AMENEH; KAZEMI, BAHRAM

2016-01-01

Aberrant DNA methylation has been investigated in carcinogenesis and as biomarker for the early detection of colorectal cancer (CRC). The present study aimed to define the methylation status in the regulatory elements of two proapoptotic genes, Fas cell surface death receptor (FAS) and BCL2-associated X protein (BAX). DNA methylation analysis was performed in tumor and adjacent normal tissue using HpaII/MspI restriction digestion and methylation-specific polymerase chain reaction (PCR). The results observed downregulation of the FAS and BAX genes in the CRC tissues compared with the adjacent normal samples. Furthermore, demethylation using 5-aza-2′-deoxycytidine treatment followed by reverse-transcription quantitative PCR were performed on the HT-29 cell line to measure BAX and FAS mRNA expression following demethylation. The 5-aza-2′-deoxycytidine treatment resulted in significant FAS gene upregulation in the HT-29 cell line, but no significant difference in BAX expression. Furthermore, analysis of CpG islands in the FAS gene promoter revealed that the FAS promoter was significantly hypermethylated in 53.3% of tumor tissues compared with adjacent normal samples. Taken together, the results indicate that decreased expression of the FAS gene due to hypermethylation of its promoter may lead to apoptotic resistance, and acts as an important step during colorectal carcinogenesis. PMID:27347139

Epigenetic-Mediated Downregulation of μ-Protocadherin in Colorectal Tumours

PubMed Central

Mateusz, Bujko; Paulina, Kober; Małgorzata, Statkiewicz; Michal, Mikula; Marcin, Ligaj; Lech, Zwierzchowski; Jerzy, Ostrowski; Aleksander, Siedlecki Janusz

2015-01-01

Carcinogenesis involves altered cellular interaction and tissue morphology that partly arise from aberrant expression of cadherins. Mucin-like protocadherin is implicated in intercellular adhesion and its expression was found decreased in colorectal cancer (CRC). This study has compared MUPCDH (CDHR5) expression in three key types of colorectal tissue samples, for normal mucosa, adenoma, and carcinoma. A gradual decrease of mRNA levels and protein expression was observed in progressive stages of colorectal carcinogenesis which are consistent with reports of increasing MUPCDH 5′ promoter region DNA methylation. High MUPCDH methylation was also observed in HCT116 and SW480 CRC cell lines that revealed low gene expression levels compared to COLO205 and HT29 cell lines which lack DNA methylation at the MUPCDH locus. Furthermore, HCT116 and SW480 showed lower levels of RNA polymerase II and histone H3 lysine 4 trimethylation (H3K4me3) as well as higher levels of H3K27 trimethylation at the MUPCDH promoter. MUPCDH expression was however restored in HCT116 and SW480 cells in the presence of 5-Aza-2′-deoxycytidine (DNA methyltransferase inhibitor). Results indicate that μ-protocadherin downregulation occurs during early stages of tumourigenesis and progression into the adenoma-carcinoma sequence. Epigenetic mechanisms are involved in this silencing. PMID:25972897

Promoter hypermethylation and downregulation of the FAS gene may be involved in colorectal carcinogenesis.

PubMed

Manoochehri, Mehdi; Borhani, Nasim; Karbasi, Ashraf; Koochaki, Ameneh; Kazemi, Bahram

2016-07-01

Aberrant DNA methylation has been investigated in carcinogenesis and as biomarker for the early detection of colorectal cancer (CRC). The present study aimed to define the methylation status in the regulatory elements of two proapoptotic genes, Fas cell surface death receptor (FAS) and BCL2-associated X protein (BAX). DNA methylation analysis was performed in tumor and adjacent normal tissue using Hpa II/ Msp I restriction digestion and methylation-specific polymerase chain reaction (PCR). The results observed downregulation of the FAS and BAX genes in the CRC tissues compared with the adjacent normal samples. Furthermore, demethylation using 5-aza-2'-deoxycytidine treatment followed by reverse-transcription quantitative PCR were performed on the HT-29 cell line to measure BAX and FAS mRNA expression following demethylation. The 5-aza-2'-deoxycytidine treatment resulted in significant FAS gene upregulation in the HT-29 cell line, but no significant difference in BAX expression. Furthermore, analysis of CpG islands in the FAS gene promoter revealed that the FAS promoter was significantly hypermethylated in 53.3% of tumor tissues compared with adjacent normal samples. Taken together, the results indicate that decreased expression of the FAS gene due to hypermethylation of its promoter may lead to apoptotic resistance, and acts as an important step during colorectal carcinogenesis.

Hsa_circ_0001649: A circular RNA and potential novel biomarker for colorectal cancer.

PubMed

Ji, Wenxin; Qiu, Chunli; Wang, Mao; Mao, Ning; Wu, Shaofeng; Dai, Yinhai

2018-02-26

The circRNAs are differentially expressed in a wide range of cancers in regulating their initiation and progression, and could be used to make a diagnosis for some diseases like tumor as a new biomarker. However, the correlation and the mechanism of action between circRNAs and colorectal cancer (CRC) are still unclear. In this study, by using qRT-PCRs, we detected the expression level of hsa_circ_0001649 in tissue and serum samples from CRC patients, and the cultured cell has been detected. We found that the hsa_circ_0001649 in CRC is significantly lower than the expression level of correspondent nontumorous tissues (n = 64, P < 0.01). We also tested the HCT116 cell lines, and the similar result is observed (n = 15, P < 0.01). Moreover, we detected the serum samples obtained before and after surgery, showing significantly the expression level of hsa_circ_0001649 in the same patient is up-regulated after surgery (n = 18, P < 0.01). Besides, we analyzed the correlation between clinicopathological date and the expression level of hsa_circ_0001649, we found that hsa_circ_0001649 expression level is closely associated with pathological differentiation (P = 0.037), and the result also illustrated that the expression level of hsa_circ_0001649 is no direct correlation with age, gender, TMN stage, lymphatic metastasis, CEA, CA19-9, and CA-724 levels. The area under the receiver operating characteristic (ROC) curve was 0.857. In conclusion, this study showed that the expression level of hsa_circ_0001649 was down-regulated in CRC and could use it as a new biomarker for specific and sensitive inspection of CRC. Copyright © 2018 Elsevier Inc. All rights reserved.

Generation, Characterization and Application of Antibodies Directed against HERV-H Gag Protein in Colorectal Samples

PubMed Central

Mullins, Christina S.; Hühns, Maja; Krohn, Mathias; Peters, Sven; Cheynet, Valérie; Oriol, Guy; Guillotte, Michèle; Ducrot, Sandrine; Mallet, François; Linnebacher, Michael

2016-01-01

Introduction A substantial part of the human genome originates from transposable elements, remnants of ancient retroviral infections. Roughly 8% of the human genome consists of about 400,000 LTR elements including human endogenous retrovirus (HERV) sequences. Mainly, the interplay between epigenetic and post-transcriptional mechanisms is thought to silence HERV expression in most physiological contexts. Interestingly, aberrant reactivation of several HERV-H loci appears specific to colorectal carcinoma (CRC). Results The expression of HERV-H Gag proteins (Gag-H) was assessed using novel monoclonal mouse anti Gag-H antibodies. In a flow cytometry screen four antibody clones were tested on a panel of primary CRC cell lines and the most well performing ones were subsequently validated in western blot analysis. Finally, Gag-H protein expression was analyzed by immune histology on cell line cytospins and on clinical samples. There, we found a heterogeneous staining pattern with no background staining of endothelial, stromal and infiltrating immune cells but diffuse staining of the cytoplasm for positive tumor and normal crypt cells of the colonic epithelium. Conclusion Taken together, the Gag-H antibody clone(s) present a valuable tool for staining of cells with colonic origin and thus form the basis for future more detailed investigations. The observed Gag-H protein staining in colonic epithelium crypt cells demands profound analyses of a potential role for Gag-H in the normal physiology of the human gut. PMID:27119520

Ras inhibitors display an anti-metastatic effect by downregulation of lysyl oxidase through inhibition of the Ras-PI3K-Akt-HIF-1α pathway.

PubMed

Yoshikawa, Yoko; Takano, Osamu; Kato, Ichiro; Takahashi, Yoshihisa; Shima, Fumi; Kataoka, Tohru

2017-12-01

Metastasis stands as the major obstacle for the survival from cancers. Nonetheless most existing anti-cancer drugs inhibit only cell proliferation, and discovery of agents having both anti-proliferative and anti-metastatic properties would be more beneficial. We previously reported the discovery of small-molecule Ras inhibitors, represented by Kobe0065, that displayed anti-proliferative activity on xenografts of human colorectal cancer (CRC) cell line SW480 carrying the K-ras G12V gene. Here we show that treatment of cancer cells carrying the activated ras genes with Kobe0065 or a siRNA targeting Ras downregulates the expression of lysyl oxidase (LOX), which has been implicated in metastasis. LOX expression is enhanced by co-expression of Ras G12V through activation of phosphatidylinositol 3-kinase (PI3K)/Akt and concomitant accumulation of hypoxia-inducible factor (HIF)-1α. Furthermore, Kobe0065 effectively inhibits not only migration and invasion of cancer cells carrying the activated ras genes but also lung metastasis of human CRC cell line SW620 carrying the K-ras G12V gene. Collectively, these results indicate that Kobe0065 prevents metastasis through inhibition of the Ras-PI3K-Akt-HIF-1α-LOX signaling and suggest that Ras inhibitors in general might exhibit both anti-proliferative and anti-metastatic properties toward cancer cells carrying the activated ras genes. Copyright © 2017 Elsevier B.V. All rights reserved.

Tricaproin Isolated From Simarouba glauca Inhibits the Growth of Human Colorectal Carcinoma Cell Lines by Targeting Class-1 Histone Deacetylases

PubMed Central

Jose, Asha; Chaitanya, Motamari V. N. L.; Kannan, Elango; Madhunapantula, SubbaRao V.

2018-01-01

While anticancer properties of Simarouba glauca (SG, commonly known as Paradise tree) are well documented in ancient literature, the underlying mechanisms leading to cancer cell death begin to emerge very recently. The leaves of SG have been used as potential source of anticancer agents in traditional medicine. Recently attempts have been made to isolate anticancer agents from the leaves of SG using solvent extraction, which identified quassinoids as the molecules with tumoricidal activity. However, it is not known whether the anti-cancer potential of SG leaves is just because of quassinoids alone or any other phytochemicals also contribute for the potency of SG leaf extracts. Therefore, SG leaves were first extracted with hexane, chloroform, ethyl acetate, 70% ethanol, water and anti-cancer potential (for inhibiting colorectal cancer (CRC) cells HCT-116 and HCT-15 proliferation) determined using Sulforhodamine-B (SRB) assay. The chloroform fraction with maximal anticancer activity was further fractionated by activity-guided isolation procedure and structure of the most potent compound determined using spectral analysis. Analysis of the structural characterization data showed the presence of tricaproin (TCN). TCN inhibited CRC cells growth in a time- and dose dependent manner but not the normal cell line BEAS-2B. Mechanistically, TCN reduced oncogenic Class-I Histone deacetylases (HDACs) activity, followed by inducing apoptosis in cells. In conclusion, the anti-cancer potential of SG is in part due to the presence of TCN in the leaves. PMID:29593526

Cysteine Dioxygenase 1 Is a Tumor Suppressor Gene Silenced by Promoter Methylation in Multiple Human Cancers

PubMed Central

Brait, Mariana; Ling, Shizhang; Nagpal, Jatin K.; Chang, Xiaofei; Park, Hannah Lui; Lee, Juna; Okamura, Jun; Yamashita, Keishi; Sidransky, David; Kim, Myoung Sook

2012-01-01

The human cysteine dioxygenase 1 (CDO1) gene is a non-heme structured, iron-containing metalloenzyme involved in the conversion of cysteine to cysteine sulfinate, and plays a key role in taurine biosynthesis. In our search for novel methylated gene promoters, we have analyzed differential RNA expression profiles of colorectal cancer (CRC) cell lines with or without treatment of 5-aza-2′-deoxycytidine. Among the genes identified, the CDO1 promoter was found to be differentially methylated in primary CRC tissues with high frequency compared to normal colon tissues. In addition, a statistically significant difference in the frequency of CDO1 promoter methylation was observed between primary normal and tumor tissues derived from breast, esophagus, lung, bladder and stomach. Downregulation of CDO1 mRNA and protein levels were observed in cancer cell lines and tumors derived from these tissue types. Expression of CDO1 was tightly controlled by promoter methylation, suggesting that promoter methylation and silencing of CDO1 may be a common event in human carcinogenesis. Moreover, forced expression of full-length CDO1 in human cancer cells markedly decreased the tumor cell growth in an in vitro cell culture and/or an in vivo mouse model, whereas knockdown of CDO1 increased cell growth in culture. Our data implicate CDO1 as a novel tumor suppressor gene and a potentially valuable molecular marker for human cancer. PMID:23028699

The epithelial polarity regulator LGALS9/galectin-9 induces fatal frustrated autophagy in KRAS mutant colon carcinoma that depends on elevated basal autophagic flux

PubMed Central

Wiersma, Valerie R; de Bruyn, Marco; Wei, Yunwei; van Ginkel, Robert J; Hirashima, Mitsuomi; Niki, Toshiro; Nishi, Nozomu; Zhou, Jin; Pouwels, Simon D; Samplonius, Douwe F; Nijman, Hans W; Eggleton, Paul; Helfrich, Wijnand; Bremer, Edwin

2015-01-01

Oncogenic mutation of KRAS (Kirsten rat sarcoma viral oncogene homolog) in colorectal cancer (CRC) confers resistance to both chemotherapy and EGFR (epidermal growth factor receptor)-targeted therapy. We uncovered that KRAS mutant (KRASmut) CRC is uniquely sensitive to treatment with recombinant LGALS9/Galectin-9 (rLGALS9), a recently established regulator of epithelial polarity. Upon treatment of CRC cells, rLGALS9 rapidly internalizes via early- and late-endosomes and accumulates in the lysosomal compartment. Treatment with rLGALS9 is accompanied by induction of frustrated autophagy in KRASmut CRC, but not in CRC with BRAF (B-Raf proto-oncogene, serine/threonine kinase) mutations (BRAFmut). In KRASmut CRC, rLGALS9 acts as a lysosomal inhibitor that inhibits autophagosome-lysosome fusion, leading to autophagosome accumulation, excessive lysosomal swelling and cell death. This antitumor activity of rLGALS9 directly correlates with elevated basal autophagic flux in KRASmut cancer cells. Thus, rLGALS9 has potent antitumor activity toward refractory KRASmut CRC cells that may be exploitable for therapeutic use. PMID:26086204

Estrogen enhances mismatch repair by induction of MLH1 expression via estrogen receptor-β

PubMed Central

Lu, Jun-Yu; Jin, Peng; Gao, Wei; Wang, De-Zhi; Sheng, Jian-Qiu

2017-01-01

Epidemiological data demonstrated that hormone replace treatment has protective effect against colorectal cancer (CRC). Our previous studies showed that this effect may be associated with DNA mismatch repair. This study aims to investigate the mechanism of estrogen induction of MLH1, and whether colorectal tumor proliferation can be inhibited through induction of MLH1 by estrogen signal pathway. Human CRC cell lines were used to examine the regulation of MLH1 expression by over-expression and depletion of estrogen receptor-α (ERα) and estrogen receptor-β (ERβ), under the treatment with 17β-estradiol or β-Estradiol 6-(O-carboxy-methyl)oxime:BSA, followed by a real-time Q-PCR and Western blotting analysis. Luciferase reporter and chromatin immunoprecipitation assays were used to identify the estrogen response elements in the proximal promoter of MLH1 gene. Then, the influence of estrogen-induced MLH1 on CRC tumor growth were determined in vitro and in vivo. We found that mismatch repair ability and microsatellite stability of cells were enhanced by estrogen via induction of MLH1 expression, which was mediated by ERβ, through a transcriptional activation process. Furthermore, we identified that ERβ exerted an inhibitory effect on CRC tumor proliferation in vitro and in vivo, combined with 5-FU, through up-regulation of MLH1 expression. Finally, we concluded that estrogen enhances mismatch repair ability and tumor inhibition effect in vitro and in vivo, via induction of MLH1 expression mediated by ERβ. PMID:28404976

Estrogen enhances mismatch repair by induction of MLH1 expression via estrogen receptor-β.

PubMed

Lu, Jun-Yu; Jin, Peng; Gao, Wei; Wang, De-Zhi; Sheng, Jian-Qiu

2017-06-13

Epidemiological data demonstrated that hormone replace treatment has protective effect against colorectal cancer (CRC). Our previous studies showed that this effect may be associated with DNA mismatch repair. This study aims to investigate the mechanism of estrogen induction of MLH1, and whether colorectal tumor proliferation can be inhibited through induction of MLH1 by estrogen signal pathway. Human CRC cell lines were used to examine the regulation of MLH1 expression by over-expression and depletion of estrogen receptor-α (ERα) and estrogen receptor-β (ERβ), under the treatment with 17β-estradiol or β-Estradiol 6-(O-carboxy-methyl)oxime:BSA, followed by a real-time Q-PCR and Western blotting analysis. Luciferase reporter and chromatin immunoprecipitation assays were used to identify the estrogen response elements in the proximal promoter of MLH1 gene. Then, the influence of estrogen-induced MLH1 on CRC tumor growth were determined in vitro and in vivo. We found that mismatch repair ability and microsatellite stability of cells were enhanced by estrogen via induction of MLH1 expression, which was mediated by ERβ, through a transcriptional activation process. Furthermore, we identified that ERβ exerted an inhibitory effect on CRC tumor proliferation in vitro and in vivo, combined with 5-FU, through up-regulation of MLH1 expression. Finally, we concluded that estrogen enhances mismatch repair ability and tumor inhibition effect in vitro and in vivo, via induction of MLH1 expression mediated by ERβ.

Unregulated smooth-muscle myosin in human intestinal neoplasia.

PubMed

Alhopuro, Pia; Phichith, Denis; Tuupanen, Sari; Sammalkorpi, Heli; Nybondas, Miranda; Saharinen, Juha; Robinson, James P; Yang, Zhaohui; Chen, Li-Qiong; Orntoft, Torben; Mecklin, Jukka-Pekka; Järvinen, Heikki; Eng, Charis; Moeslein, Gabriela; Shibata, Darryl; Houlston, Richard S; Lucassen, Anneke; Tomlinson, Ian P M; Launonen, Virpi; Ristimäki, Ari; Arango, Diego; Karhu, Auli; Sweeney, H Lee; Aaltonen, Lauri A

2008-04-08

A recent study described a recessive ATPase activating germ-line mutation in smooth-muscle myosin (smmhc/myh11) underlying the zebrafish meltdown (mlt) phenotype. The mlt zebrafish develops intestinal abnormalities reminiscent of human Peutz-Jeghers syndrome (PJS) and juvenile polyposis (JP). To examine the role of MYH11 in human intestinal neoplasia, we searched for MYH11 mutations in patients with colorectal cancer (CRC), PJS and JP. We found somatic protein-elongating frameshift mutations in 55% of CRCs displaying microsatellite instability and in the germ-line of one individual with PJS. Additionally, two somatic missense mutations were found in one microsatellite stable CRC. These two missense mutations, R501L and K1044N, and the frameshift mutations were functionally evaluated. All mutations resulted in unregulated molecules displaying constitutive motor activity, similar to the mutant myosin underlying mlt. Thus, MYH11 mutations appear to contribute also to human intestinal neoplasia. Unregulated MYH11 may affect the cellular energy balance or disturb cell lineage decisions in tumor progenitor cells. These data challenge our view on MYH11 as a passive differentiation marker functioning in muscle contraction and add to our understanding of intestinal neoplasia.

NCOA5 promotes proliferation, migration and invasion of colorectal cancer cells via activation of PI3K/AKT pathway

PubMed Central

Xie, Yufeng; He, Yang; Wang, Zhenxin; Qin, Lei

2017-01-01

The nuclear receptor coactivator 5 (NCOA5) displays both coactivator and corepressor functions. Previous studies showed that alteration of NCOA5 participates in carcinogenesis and progression. However, its roles in colorectal cancer (CRC) remain unknown. Herein, we demonstrated that expression of NCOA5 in human CRC tissues was notably higher than that in adjacent tissues, which significantly correlated with clinicopathological features such as length of tumor, regional lymph node staging and cancer staging. Knockdown of NCOA5 markedly suppressed proliferation, migration and invasion of SW620 high malignant CRC cells. Silencing of NCOA5 also inhibited in vivo growth of SW620 CRC subcutaneously xenografted tumors in athymic BALB/c nude mice. Meanwhile, Overexpression of NCOA5 facilitated these processes in SW480 low malignant CRC cells. Furthermore, knockdown of NCOA5 induced cell cycle G1 phase arrest in SW620 cells, whereas overexpression of NCOA5 promoted G1 to S phase transition in SW480 cells. Mechanistic studies revealed that NCOA5 upregulated phospho-protein kinase B (p-PKB/AKT), Cyclin D1 and matrix metalloproteinase 9 (MMP9) as well as downregulated P27 in CRC cells. Notably, PI3K inhibitor LY294002 obviously attenuated the effects of NCOA5 on p-AKT, Cyclin D1, P27 and MMP9. Moreover, LY294002 and knockdown of Cyclin D1 or MMP9 remarkably blocked the tumor-promoting activity of NCOA5. Collectively, NCOA5 promoted CRC cell proliferation, migration and invasion by upregulating Cyclin D1 and MMP9 while downregulating P27 to a great extent via activating PI3K/AKT signaling pathway. These findings suggested that NCOA5 exhibits an oncogenic effect in human CRC and represents a novel therapeutic target for CRC. PMID:29296214

Long noncoding RNA lnc-sox5 modulates CRC tumorigenesis by unbalancing tumor microenvironment.

PubMed

Wu, Kaiming; Zhao, Zhenxian; Liu, Kuanzhi; Zhang, Jian; Li, Guanghua; Wang, Liang

2017-07-03

Long non-coding RNAs (LncRNAs) have been recently regarded as systemic regulators in multiple biologic processes including tumorigenesis. In this study, we observed the expression of lncRNA lnc-sox5 was significantly increased in colorectal cancer (CRC). Despite the CRC cell growth, cell cycle and cell apoptosis was not affected by lnc-sox5 knock-down, lnc-sox5 knock-down suppressed CRC cell migration and invasion. In addition, xenograft animal model suggested that lnc-sox5 knock-down significantly suppressed the CRC tumorigenesis. Our results also showed that the expression of indoleamine 2,3-dioxygenase 1 (IDO1) was significantly reduced by lnc-sox5 knock-down and therefore modulated the infiltration and cytotoxicity of CD3 + CD8 + T cells. Taken together, these results suggested that lnc-sox5 unbalances tumor microenvironment to regulate colorectal cancer progression.

Selective targeting of mutant adenomatous polyposis coli (APC) in colorectal cancer.

PubMed

Zhang, Lu; Theodoropoulos, Panayotis C; Eskiocak, Ugur; Wang, Wentian; Moon, Young-Ah; Posner, Bruce; Williams, Noelle S; Wright, Woodring E; Kim, Sang Bum; Nijhawan, Deepak; De Brabander, Jef K; Shay, Jerry W

2016-10-19

Mutations in the adenomatous polyposis coli (APC) gene are common in colorectal cancer (CRC), and more than 90% of those mutations generate stable truncated gene products. We describe a chemical screen using normal human colonic epithelial cells (HCECs) and a series of oncogenically progressed HCECs containing a truncated APC protein. With this screen, we identified a small molecule, TASIN-1 (truncated APC selective inhibitor-1), that specifically kills cells with APC truncations but spares normal and cancer cells with wild-type APC. TASIN-1 exerts its cytotoxic effects through inhibition of cholesterol biosynthesis. In vivo administration of TASIN-1 inhibits tumor growth of CRC cells with truncated APC but not APC wild-type CRC cells in xenograft models and in a genetically engineered CRC mouse model with minimal toxicity. TASIN-1 represents a potential therapeutic strategy for prevention and intervention in CRC with mutant APC. Copyright © 2016, American Association for the Advancement of Science.

Circular BANP, an upregulated circular RNA that modulates cell proliferation in colorectal cancer.

PubMed

Zhu, Mingchen; Xu, Yijun; Chen, Yun; Yan, Feng

2017-04-01

Circular RNAs (circRNAs) are recently identified as widespread and diverse endogenous noncoding RNAs that may harbor vital functions in human and animals. However, the role of circRNAs in the process of tumorigenesis and development of colorectal cancer (CRC) remains vague. Here we characterized the circRNA expression profile from three paired CRC cancerous and adjacent normal tissues by human circRNA array, and identified 136 significantly overexpressed circRNAs and 243 downregulated circRNAs in CRC cancerous tissues (>2-fold changes). We further validated one circRNA generated from Exon 5-11 of BANP gene, termed circ-BANP. In addition, RT-PCR result showed that circ-BANP was overexpressed in 35 CRC cancerous tissues. Knockdown of circ-BANP with siRNA significantly attenuate the proliferation of CRC cells. In summary, our findings demonstrated that dysregulated circ-BANP appears to have an important role in CRC cells and could serve as a prognostic and therapeutic marker for CRC. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Smad4-Mediated Signaling Inhibits Intestinal Neoplasia by Inhibiting Expression of β-Catenin

PubMed Central

Freeman, Tanner J.; Smith, J. Joshua; Chen, Xi; Washington, M. Kay; Roland, Joseph T.; Means, Anna L.; Eschrich, Steven A.; Yeatman, Timothy J.; Deane, Natasha G.; Beauchamp, R. Daniel

2012-01-01

Background & Aims Mutational inactivation of APC is an early event in colorectal cancer (CRC) progression that affects the stability and increases the activity of β-catenin, a mediator of Wnt signaling. CRC progression also involves inactivation of signaling via transforming growth factor (TGF)β and bone morphenogenic protein (BMP), which are tumor suppressors. However, the interactions between these pathways are not clear. We investigated the effects of loss of the transcription factor Smad4 loss on levels of β-catenin mRNA and Wnt signaling. Methods We used microarray analysis to associate levels of Smad4 and β-catenin mRNA in colorectal tumor samples from 250 patients. We performed oligonucleotide-mediated knockdown of Smad4 in human embryonic kidney (HEK293T) and in HCT116 colon cancer cells and transgenically expressed Smad4 in SW480 colon cancer cells. We analyzed adenomas from (APCΔ1638/+) and (APCΔ1638/+)x(K19CreERT2Smad4lox/lox) mice using laser-capture microdissection. Results In human CRC samples, reduced levels of Smad4 correlated with increased levels of β-catenin mRNA. In Smad4-depleted cell lines, levels of β-catenin mRNA and Wnt signaling increased. Inhibition of BMP or depletion of Smad4 in HEK293T cells increased binding of RNA polymerase II to the β-catenin gene. Expression of Smad4 in SW480 cells reduced Wnt signaling and levels of β-catenin mRNA. In mice with heterozygous disruption of Apc(APCΔ1638/+), Smad4-deficient intestinal adenomas had increased levels of β-catenin mRNA and expression of Wnt target genes, compared with adenomas from APCΔ1638/+mice that expressed Smad4. Conclusions Transcription of β-catenin is inhibited by BMP signaling to Smad4. These findings provide important information about the interaction among TGF-β, BMP, and Wnt signaling pathways in CRC progression. PMID:22115830

Murine Endogenous Retroviruses Are Detectable in Patient-Derived Xenografts but Not in Patient-Individual Cell Lines of Human Colorectal Cancer.

PubMed

Bock, Stephanie; Mullins, Christina S; Klar, Ernst; Pérot, Philippe; Maletzki, Claudia; Linnebacher, Michael

2018-01-01

Endogenous retroviruses are remnants of retroviral infections. In contrast to their human counterparts, murine endogenous retroviruses (mERV) still can synthesize infectious particles and retrotranspose. Xenotransplanted human cells have occasionally been described to be mERV infected. With genetic engineered mice and patient-derived xenografts (PDXs) on the rise as eminent research tools, we here systematically investigated, if different tumor models harbor mERV infections. Relevant mERV candidates were first preselected by next generation sequencing (NGS) analysis of spontaneous lymphomas triggered by colorectal cancer (CRC) PDX tissue. Two primer systems were designed for each of these candidates (AblMLV, EcoMLV, EndoPP, MLV, and preXMRV) and implemented in an quantitative real-time (RT-qPCR) screen using murine tissues ( n = 11), PDX-tissues ( n = 22), PDX-derived cell lines ( n = 13), and patient-derived tumor cell lines ( n = 14). The expression levels of mERV varied largely both in the PDX samples and in the mouse tissues. No mERV signal was, however, obtained from cDNA or genomic DNA of CRC cell lines. Expression of EcoMLV was higher in PDX than in murine tissues; for EndoPP it was the opposite. These two were thus further investigated in 40 additional PDX. In addition, four patient-derived cell lines free of any mERV expression were subcutaneously injected into immunodeficient mice. Outgrowing cell-derived xenografts barely expressed EndoPP. In contrast, the expression of EcoMLV was even higher than in surrounding mouse tissues. This expression gradually vanished within few passages of re-cultivated cells. In summary, these results strongly imply that: (i) PDX and murine tissues in general are likely to be contaminated by mERV, (ii) mERV are expressed transiently and at low level in fresh PDX-derived cell cultures, and (iii) mERV integration into the genome of human cells is unlikely or at least a very rare event. Thus, mERVs are stowaways present in murine cells, in PDX tissues and early thereof-derived cell cultures. We conclude that further analysis is needed concerning their impact on results obtained from studies performed with PDX but also with murine tumor models.

Circulating and tumor-infiltrating Tim-3 in patients with colorectal cancer

PubMed Central

Gao, Quanli; Yuan, Peng; Zhao, Peng; Yuan, Huijuan; Fan, Huijie; Li, Tiepeng; Qin, Peng; Han, Lu; Fang, Weijia; Suo, Zhenhe

2015-01-01

T-cell exhaustion represents a progressive loss of T-cell function. The inhibitory receptor PD-1 is known to negatively regulate CD8+ T cell responses directed against tumor antigen, but the blockades of PD-1 pathway didn't show the objective responses in patients with colorectal cancer (CRC). Thus, further exploring the molecular mechanism responsible for inducing T-cell dysfunction in CRC patients may reveal effective strategies for immune therapy. This study aims to characterize co-inhibitory receptors on T cells in CRC patients to identify novel targets for immunotherapy. In this study, peripheral blood samples from 20 healthy controls and 54 consented CRC patients, and tumor and matched paraneoplastic tissues from 7 patients with advanced CRC, subjected to multicolor flow cytometric analysis of the expression of PD-1 and Tim-3 receptors on CD8+ T cells. It was found that CRC patients presented with significantly higher levels of circulating Tim-3+PD-1+CD8+ T cells compared to the healthy controls (medians of 3.12% and 1.99%, respectively, p = 0.0403). A similar increase of Tim-3+PD-1+CD8+ T cells was also observed in the tumor tissues compared to paraneoplastic tussues. Tim-3+PD-1+CD8+ T cells in tumor tissues produced even less cytokine than that in paraneoplastic tissues. Functional ex vivo experiments showed that Tim-3+PD-1+CD8+ T cells produced significantly less IFN-γ than Tim-3−PD-1−CD8+ T cells, followed by Tim-3+PD-1−CD8+ T cells, and Tim-3−PD-1+CD8+ T cells, indicating a stronger inhibition of IFN-γ production of Tim-3+CD8+ T cells. It is also found in this study that Tim-3+PD-1+CD8+ T cell increase in circulation was correlated with clinical cancer stage but not histologic grade and serum concentrations of cancer biomarker CEA. Our results indicate that upregulation of the inhibitory receptor Tim-3 may restrict T cell responses in CRC patients, and therefore blockage of Tim-3 and thus restoring T cell responses may be a potential therapeutic approach for CRC patients. PMID:26008981

Bone morphogenetic protein 4 induces differentiation of colorectal cancer stem cells and increases their response to chemotherapy in mice.

PubMed

Lombardo, Ylenia; Scopelliti, Alessandro; Cammareri, Patrizia; Todaro, Matilde; Iovino, Flora; Ricci-Vitiani, Lucia; Gulotta, Gaspare; Dieli, Francesco; de Maria, Ruggero; Stassi, Giorgio

2011-01-01

The limited clinical response observed in many patients with colorectal cancer may be related to the presence of chemoresistant colorectal cancer stem cells (CRC-SCs). Bone morphogenetic protein 4 (BMP4) promotes the differentiation of normal colonic stem cells. We investigated whether BMP4 might be used to induce differentiation of CRC-SCs and for therapeutic purposes. CRC-SCs were isolated from 25 tumor samples based on expression of CD133 or using a selection culture medium. BMP4 expression and activity on CRC-SCs were evaluated in vitro; progeny of the stem cells were evaluated by immunofluorescence, immunoblot, and flow cytometry analyses. The potential therapeutic effect of BMP4 was assessed in immunocompromised mice after injection of CRC-SCs that responded to chemotherapy (n = 4) or that did not (n = 2). CRC-SCs did not express BMP4 whereas differentiated cells did. Recombinant BMP4 promoted differentiation and apoptosis of CRC-SCs in 12 of 15 independent experiments; this effect did not depend on Small Mothers against decapentaplegic (Smad)4 expression level or microsatellite stability. BMP4 activated the canonical and noncanonical BMP signaling pathways, including phosphoInositide 3-kinase (PI3K) and PKB (protein kinase B)/AKT. Mutations in PI3K or loss of Phosphatase and Tensin homolog (PTEN) in Smad4-defective tumors made CRC-SCs unresponsive to BMP4. Administration of BMP4 to immunocompromised mice with tumors that arose from CRC-SCs increased the antitumor effects of 5-fluorouracil and oxaliplatin. BMP4 promotes terminal differentiation, apoptosis, and chemosensitization of CRC-SCs in tumors that do not have simultaneous mutations in Smad4 and constitutive activation of PI3K. BMP4 might be developed as a therapeutic agent against cancer stem cells in advanced colorectal tumors. Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.

Somatic mutations in cancer: Stochastic versus predictable.

PubMed

Gold, Barry

2017-02-01

The origins of human cancers remain unclear except for a limited number of potent environmental mutagens, such as tobacco and UV light, and in rare cases, familial germ line mutations that affect tumor suppressor genes or oncogenes. A significant component of cancer etiology has been deemed stochastic and correlated with the number of stem cells in a tissue, the number of times the stem cells divide and a low incidence of random DNA polymerase errors that occur during each cell division. While somatic mutations occur during each round of DNA replication, mutations in cancer driver genes are not stochastic. Out of a total of 2843 codons, 1031 can be changed to stop codons by a single base substitution in the tumor suppressor APC gene, which is mutated in 76% of colorectal cancers (CRC). However, the nonsense mutations, which comprise 65% of all the APC driver mutations in CRC, are not random: 43% occur at Arg CGA codons, although they represent <3% of the codons. In TP53, CGA codons comprise <3% of the total 393 codons but they account for 72% and 39% of the mutations in CRC and ovarian cancer OVC, respectively. This mutation pattern is consistent with the kinetically slow, but not stochastic, hydrolytic deamination of 5-methylcytosine residues at specific methylated CpG sites to afford T·G mismatches that lead to C→T transitions and stop codons at CGA. Analysis of nonsense mutations in CRC, OVC and a number of other cancers indicates the need to expand the predictable risk factors for cancer to include, in addition to random polymerase errors, the methylation status of gene body CGA codons in tumor suppressor genes. Copyright © 2017. Published by Elsevier B.V.

Increased PTP1B expression and phosphatase activity in colorectal cancer results in a more invasive phenotype and worse patient outcome.

PubMed

Hoekstra, Elmer; Das, Asha M; Swets, Marloes; Cao, Wanlu; van der Woude, C Janneke; Bruno, Marco J; Peppelenbosch, Maikel P; Kuppen, Peter J K; Ten Hagen, Timo L M; Fuhler, Gwenny M

2016-04-19

Cell signaling is dependent on the balance between phosphorylation of proteins by kinases and dephosphorylation by phosphatases. This balance if often disrupted in colorectal cancer (CRC), leading to increased cell proliferation and invasion. For many years research has focused on the role of kinases as potential oncogenes in cancer, while phosphatases were commonly assumed to be tumor suppressive. However, this dogma is currently changing as phosphatases have also been shown to induce cancer growth. One of these phosphatases is protein tyrosine phosphatase 1B (PTP1B). Here we report that the expression of PTP1B is increased in colorectal cancer as compared to normal tissue, and that the intrinsic enzymatic activity of the protein is also enhanced. This suggests a role for PTP1B phosphatase activity in CRC formation and progression. Furthermore, we found that increased PTP1B expression is correlated to a worse patient survival and is an independent prognostic marker for overall survival and disease free survival. Knocking down PTP1B in CRC cell lines results in a less invasive phenotype with lower adhesion, migration and proliferation capabilities. Together, these results suggest that inhibition of PTP1B activity is a promising new target in the treatment of colorectal cancer and the prevention of metastasis.

An apple oligogalactan suppresses endotoxin-induced cyclooxygenase-2 expression by inhibition of LPS pathways.

PubMed

Li, Yuhua; Fan, Lei; Sun, Yang; Zhang, Dian; Yue, Zhenggang; Niu, Yinbo; Meng, Jin; Yang, Tiehong; Liu, Wenchao; Mei, Qibing

2013-10-01

Colorectal cancer (CRC) is one of the most common cancers and a leading cause of cancer-related mortality in developed countries. Many ingredients of apples have been proven to have anti-inflammatory and anti-carcinogenic characteristics, and show benefits for CRC prevention. The aim of this study, therefore, was to evaluate inhibitory effect of an apple oligogalactan (AOG) on pro-inflammatory endotoxin lipopolysaccharide (LPS)-activated human colon carcinoma cells HT-29 and SW-620 and investigate the possible mechanisms. The two cell lines were pretreated with AOG (0.1-1 g/L) for 30 min and then treated with 10 μg/mL LPS. Real time PCR, Western blot, electrophoretic mobility shift assay (EMSA), and ELISA were used to detect the expression and activity of cyclooxygenase-2 (COX-2), NF-κB and MAPKs pathways. AOG significantly inhibited the expression and activity of COX-2 in LPS-activated human colon carcinoma cells HT-29 and SW-620. The mechanisms of AOG-suppressed COX-2 expression may be through inhibiting the phosphorylation of MAPKs and the activation of NF-κB and AP-1. These data may provide another molecular basis for understanding how apples act to prevent CRC and indicate that AOG may be useful for treatment of colitis and prevention of carcinogenesis. Copyright © 2013 Elsevier B.V. All rights reserved.

Molecular profiling of ALDH1+ colorectal cancer stem cells reveals preferential activation of MAPK, FAK, and oxidative stress pro-survival signalling pathways.

PubMed

Vishnubalaji, Radhakrishnan; Manikandan, Muthurangan; Fahad, Mohamed; Hamam, Rimi; Alfayez, Musaad; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M

2018-03-02

Tumour heterogeneity leads to variable clinical response and inaccurate diagnostic and prognostic assessment. Cancer stem cells (CSCs) represent a subpopulation responsible for invasion, metastasis, therapeutic resistance, and recurrence in many human cancer types. However, the true identity of colorectal cancer (CRC) SCs remains elusive. Here, we aimed to characterize and define the gene expression portrait of CSCs in CRC-model SW403 cells. We found that ALDH + positive cells are clonogenic and highly proliferative; their global gene expression profiling-based molecular signature revealed gene enrichment related to DNA damage, MAPK, FAK, oxidative stress response, and Wnt signalling. ALDH + cells showed enhanced ROS stress resistance, whereas MAPK/FAK pathway pharmacologic inhibition limited their survival. Conversely, 5-fluorouracil increased the ALDH + cell fraction among the SW403, HCT116 and SW620 CRC models. Notably, analysis of ALDH1A1 and POU5F1 expression levels in cohorts of 462 or 420 patients for overall (OS) or disease-free (DFS) survival, respectively, obtained from the Cancer Genome Atlas CRC dataset, revealed strong association between elevated expression and poor OS ( p = 0.006) and poor DFS ( p = 0.05), thus implicating ALDH1A1 and POU5F1 in CRC prognosis. Our data reveal distinct molecular signature of ALDH + CSCs in CRC and suggest pathways relevant for successful targeted therapies and management of CRC.

Cancer Stem Cells and Molecular Biology Test in Colorectal Cancer: Therapeutic Implications.

PubMed

Effendi-Ys, Rustam

2017-10-01

Colorectal cancer (CRC) is the third most frequent cancer in males, the second in females, and is the second leading cause of cancer related death worldwide. Within Indonesia's 250 million population, the incidence rates for CRC per 100,000 population were 15.2 for males and 10.2 for females, and estimated 63,500 cases per year.  More than 50% of colorectal cancer patients will develop metastasis. CRC is still the main cause of tumor-related death, and although most CRC patients are treated with surgery to remove the tumor tissue, some of the CRC patients recurred. Chemotherapy used as adjuvant or neoadjuvant therapy also has several problems, in which these treatments are useless in tumor cells with chemo-resistance. Molecular testing of CRC from tumor tissues has important implications for the selection of treatment. Biomarkers can be used as prognostic value, molecular predictive factors, and targeted therapy. Recent research reported that, cancer stem cells (CSCs) are considered as the origin of tumorigenesis, development, metastasis and recurrence. At present, it has been shown that CSCs existed in many tumors including CRC. This review aims to summarize the issue on CSCs, and the future development of drugs that target colorectal cancer stem cells.

FAP positive fibroblasts induce immune checkpoint blockade resistance in colorectal cancer via promoting immunosuppression.

PubMed

Chen, Lingling; Qiu, Xiangting; Wang, Xinhua; He, Jian

2017-05-20

Immune checkpoint blockades that significantly prolonged survival of melanoma patients have been less effective on colorectal cancer (CRC) patients. Growing evidence suggested that fibroblast activation protein-alpha (FAP) on cancer associate fibroblasts (CAFs) has critical roles in regulating antitumor immune response by inducing tumor-promoting inflammation. In this study, we explored the roles of FAP in regulating the tumor immunity and immune checkpoint blockades resistance in CRC experimental systems. We found that CAFs with high FAP expression could induce immune checkpoint blockade resistance in CRC mouse model. Mechanistically, CAFs with high FAP expression promoted immunosuppression in the CRC tumor immune microenvironment by up-regulating CCL2 secretion, recruiting myeloid cells, and decreasing T-cell activity. In human CRC samples, FAP expression was proportional to myeloid cells number, but inversely related to T-cell number. High FAP expression also predicted poor survival of CRC patients. Taken together, our study suggested that high FAP expression in CAFs is one reason leading to immune checkpoint blockades resistance in CRC patients and FAP is an optional target for reversing immune checkpoint blockades resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

TAM receptors Tyro3 and Mer as novel targets in colorectal cancer.

PubMed

Schmitz, Robin; Valls, Aida Freire; Yerbes, Rosario; von Richter, Sophie; Kahlert, Christoph; Loges, Sonja; Weitz, Jürgen; Schneider, Martin; Ruiz de Almodovar, Carmen; Ulrich, Alexis; Schmidt, Thomas

2016-08-30

CRC remains the third most common cancer worldwide with a high 5-year mortality rate in advanced cases. Combined with chemotherapy, targeted therapy is an additional treatment option. However as CRC still escapes targeted therapy the vigorous search for new targets is warranted to increase patients´ overall survival. In this study we describe a new role for Gas6/protein S-TAM receptor interaction in CRC. Gas6, expressed by tumor-infiltrating M2-like macrophages, enhances malignant properties of tumor cells including proliferation, invasion and colony formation. Upon chemotherapy macrophages increase Gas6 synthesis, which significantly attenuates the cytotoxic effect of 5-FU chemotherapy on tumor cells. The anti-coagulant protein S has similar effects as Gas6.In CRC patient samples Tyro3 was overexpressed within the tumor. In-vitro inhibition of Tyro3 and Mer reduces tumor cell proliferation and sensitizes tumor cells to chemotherapy. Moreover high expression of Tyro3 and Mer in tumor tissue significantly shortens CRC patients´ survival. Various in vitro models were used to investigate the role of Gas6 and its TAM receptors in human CRC cells, by stimulation (rhGas6) and knockdown (siRNA) of Axl, Tyro3 and Mer. In terms of a translational research, we additionally performed an expression analysis in human CRC tissue and analyzed the medical record of these patients. Tyro3 and Mer represent novel therapeutic targets in CRC and warrant further preclinical and clinical investigation in the future.

Individual patient data analysis of progression-free survival versus overall survival as a first-line end point for metastatic colorectal cancer in modern randomized trials: findings from the analysis and research in cancers of the digestive system database.

PubMed

Shi, Qian; de Gramont, Aimery; Grothey, Axel; Zalcberg, John; Chibaudel, Benoist; Schmoll, Hans-Joachim; Seymour, Matthew T; Adams, Richard; Saltz, Leonard; Goldberg, Richard M; Punt, Cornelis J A; Douillard, Jean-Yves; Hoff, Paulo M; Hecht, Joel Randolph; Hurwitz, Herbert; Díaz-Rubio, Eduardo; Porschen, Rainer; Tebbutt, Niall C; Fuchs, Charles; Souglakos, John; Falcone, Alfredo; Tournigand, Christophe; Kabbinavar, Fairooz F; Heinemann, Volker; Van Cutsem, Eric; Bokemeyer, Carsten; Buyse, Marc; Sargent, Daniel J

2015-01-01

Progression-free survival (PFS) has previously been established as a surrogate for overall survival (OS) for first-line metastatic colorectal cancer (mCRC). Because mCRC treatment has advanced in the last decade with extended OS, this surrogacy requires re-examination. Individual patient data from 16,762 patients were available from 22 first-line mCRC studies conducted from 1997 to 2006; 12 of those studies tested antiangiogenic and/or anti-epidermal growth factor receptor agents. The relationship between PFS (first event of progression or death) and OS was evaluated by using R(2) statistics (the closer the value is to 1, the stronger the correlation) from weighted least squares regression of trial-specific hazard ratios estimated by using Cox and Copula models. Forty-four percent of patients received a regimen that included biologic agents. Median first-line PFS was 8.3 months, and median OS was 18.2 months. The correlation between PFS and OS was modest (R(2), 0.45 to 0.69). Analyses limited to trials that tested treatments with biologic agents, nonstrategy trials, or superiority trials did not improve surrogacy. In modern mCRC trials, in which survival after the first progression exceeds time to first progression, a positive but modest correlation was observed between OS and PFS at both the patient and trial levels. This finding demonstrates the substantial variability in OS introduced by the number of lines of therapy and types of effective subsequent treatments and the associated challenge to the use of OS as an end point to assess the benefit attributable to a single line of therapy. PFS remains an appropriate primary end point for first-line mCRC trials to detect the direct treatment effect of new agents. © 2014 by American Society of Clinical Oncology.

Individual Patient Data Analysis of Progression-Free Survival Versus Overall Survival As a First-Line End Point for Metastatic Colorectal Cancer in Modern Randomized Trials: Findings From the Analysis and Research in Cancers of the Digestive System Database

PubMed Central

Shi, Qian; de Gramont, Aimery; Grothey, Axel; Zalcberg, John; Chibaudel, Benoist; Schmoll, Hans-Joachim; Seymour, Matthew T.; Adams, Richard; Saltz, Leonard; Goldberg, Richard M.; Punt, Cornelis J.A.; Douillard, Jean-Yves; Hoff, Paulo M.; Hecht, Joel Randolph; Hurwitz, Herbert; Díaz-Rubio, Eduardo; Porschen, Rainer; Tebbutt, Niall C.; Fuchs, Charles; Souglakos, John; Falcone, Alfredo; Tournigand, Christophe; Kabbinavar, Fairooz F.; Heinemann, Volker; Van Cutsem, Eric; Bokemeyer, Carsten; Buyse, Marc; Sargent, Daniel J.

2015-01-01

Purpose Progression-free survival (PFS) has previously been established as a surrogate for overall survival (OS) for first-line metastatic colorectal cancer (mCRC). Because mCRC treatment has advanced in the last decade with extended OS, this surrogacy requires re-examination. Methods Individual patient data from 16,762 patients were available from 22 first-line mCRC studies conducted from 1997 to 2006; 12 of those studies tested antiangiogenic and/or anti–epidermal growth factor receptor agents. The relationship between PFS (first event of progression or death) and OS was evaluated by using R2 statistics (the closer the value is to 1, the stronger the correlation) from weighted least squares regression of trial-specific hazard ratios estimated by using Cox and Copula models. Results Forty-four percent of patients received a regimen that included biologic agents. Median first-line PFS was 8.3 months, and median OS was 18.2 months. The correlation between PFS and OS was modest (R2, 0.45 to 0.69). Analyses limited to trials that tested treatments with biologic agents, nonstrategy trials, or superiority trials did not improve surrogacy. Conclusion In modern mCRC trials, in which survival after the first progression exceeds time to first progression, a positive but modest correlation was observed between OS and PFS at both the patient and trial levels. This finding demonstrates the substantial variability in OS introduced by the number of lines of therapy and types of effective subsequent treatments and the associated challenge to the use of OS as an end point to assess the benefit attributable to a single line of therapy. PFS remains an appropriate primary end point for first-line mCRC trials to detect the direct treatment effect of new agents. PMID:25385741

The cytotoxic, apoptotic and oxidative effects of carbonic anhydrase IX inhibitor on colorectal cancer cells.

PubMed

Tülüce, Yasin; Ahmed, Bewar Ali; Koyuncu, İsmail; Durgun, Mustafa

2018-04-01

Colorectal cancer (CRC) is the third most common tumor, malignant and has developed one of the main reasons of cancer mortality. According to studies conducted recently; carbonic anhydrase 9 (CAIX) is an especially attractive target for cancer therapy, in part since it is limited way expressed in normal tissues on the other hand in a wide variety of solid neoplasia are overexpressed. The aim of this study was to appreciate the effects of CAIX inhibitor, namely novel synthesized sulfonamide derivative (H-4i) with high affinity for CAIX, in CAIX-positive human colorectal cancer cell (HT-29) and CAIX-negative human normal embryonic kidney cell line (HEK-293). For this reason, we planned to investigate apoptotic, cytotoxic and oxidative stress activity of H-4i on HT-29 and HEK-293 cell lines. Cell viability determined by WST-1 assay afterwards IC 50 values, apoptosis and cell cycle induction measured by flow cytometric analysis, intracellular free radical induction performed by reactive oxygen species (ROS) analyses. The IC 50 value of the sulfonamide derivative compound was found to be very low, especially in HT-29 cells, when compared to human normal cells. This research found that H-4i significantly increased cytotoxicity and ROS production, caused significant signs of apoptosis level. High level of ROS and apoptosis lead to arrest the cell cycle and reduce cell survival. The most obvious finding to emerge from the analysis that novel synthesized sulfonamide derivative H-4i is effective on HT-29 more than HEK-293. Therefore, novel derivative H-4i might be used as an anti-cancer potential compound on CRC.

CRISPR Knockout of the HuR Gene Causes a Xenograft Lethal Phenotype

PubMed Central

Lal, Shruti; Cheung, Edwin C.; Zarei, Mahsa; Preet, Ranjan; Chand, Saswati N.; Mambelli-Lisboa, Nicole C.; Romeo, Carmella; Stout, Matthew C.; Londin, Eric; Goetz, Austin; Lowder, Cinthya Y.; Nevler, Avinoam; Yeo, Charles J.; Campbell, Paul M.; Winter, Jordan M.; Dixon, Dan A.; Brody, Jonathan R.

2017-01-01

Pancreatic ductal adenocarcinoma (PDA) is the third leading cause of cancer related deaths in the U.S., while colorectal cancer (CRC) is the third most common cancer. The RNA binding protein HuR (ELAVL1), supports a pro-oncogenic network in gastrointestinal (GI) cancer cells through enhanced HuR expression. Using a publically available database, HuR expression levels were determined to be increased in primary PDA and CRC tumor cohorts as compared to normal pancreas and colon tissues, respectively. CRISPR/Cas9 technology was successfully used to delete the HuR gene in both PDA (MIA PaCa-2 and Hs 766T) and CRC (HCT116) cell lines. HuR deficiency has a mild phenotype, in vitro, as HuR-deficient MIA PaCa-2 (MIA.HuR-KO(−/−)) cells had increased apoptosis when compared to isogenic wild-type (MIA.HuR-WT(+/+)) cells. Using this isogenic system, mRNAs were identified that specifically bound to HuR and were required for transforming a 2D culture into 3D (i.e., organoids). Importantly, HuR-deficient MIA PaCa-2 and Hs 766T cells were unable to engraft tumors in vivo compared to control HuR-proficient cells, demonstrating a unique xenograft lethal phenotype. While not as a dramatic phenotype, CRISPR knockout HuR HCT116 colon cancer cells (HCT.HuR-KO(−/−)) showed significantly reduced in vivo tumor growth compared to controls (HCT.HuR-WT(+/+)). Finally, HuR deletion affects KRAS activity and controls a subset of pro-oncogenic genes. Implications The work reported here supports the notion that targeting HuR is a promising therapeutic strategy to treat GI malignancies. PMID:28242812

Curcumin modulates DNA methylation in colorectal cancer cells.

PubMed

Link, Alexander; Balaguer, Francesc; Shen, Yan; Lozano, Juan Jose; Leung, Hon-Chiu E; Boland, C Richard; Goel, Ajay

2013-01-01

Recent evidence suggests that several dietary polyphenols may exert their chemopreventive effect through epigenetic modifications. Curcumin is one of the most widely studied dietary chemopreventive agents for colon cancer prevention, however, its effects on epigenetic alterations, particularly DNA methylation, remain unclear. Using systematic genome-wide approaches, we aimed to elucidate the effect of curcumin on DNA methylation alterations in colorectal cancer cells. To evaluate the effect of curcumin on DNA methylation, three CRC cell lines, HCT116, HT29 and RKO, were treated with curcumin. 5-aza-2'-deoxycytidine (5-aza-CdR) and trichostatin A treated cells were used as positive and negative controls for DNA methylation changes, respectively. Methylation status of LINE-1 repeat elements, DNA promoter methylation microarrays and gene expression arrays were used to assess global methylation and gene expression changes. Validation was performed using independent microarrays, quantitative bisulfite pyrosequencing, and qPCR. As expected, genome-wide methylation microarrays revealed significant DNA hypomethylation in 5-aza-CdR-treated cells (mean β-values of 0.12), however, non-significant changes in mean β-values were observed in curcumin-treated cells. In comparison to mock-treated cells, curcumin-induced DNA methylation alterations occurred in a time-dependent manner. In contrast to the generalized, non-specific global hypomethylation observed with 5-aza-CdR, curcumin treatment resulted in methylation changes at selected, partially-methylated loci, instead of fully-methylated CpG sites. DNA methylation alterations were supported by corresponding changes in gene expression at both up- and down-regulated genes in various CRC cell lines. Our data provide previously unrecognized evidence for curcumin-mediated DNA methylation alterations as a potential mechanism of colon cancer chemoprevention. In contrast to non-specific global hypomethylation induced by 5-aza-CdR, curcumin-induced methylation changes occurred only in a subset of partially-methylated genes, which provides additional mechanistic insights into the potent chemopreventive effect of this dietary nutraceutical.

Curcumin Modulates DNA Methylation in Colorectal Cancer Cells

PubMed Central

Link, Alexander; Balaguer, Francesc; Shen, Yan; Lozano, Juan Jose; Leung, Hon-Chiu E.; Boland, C. Richard; Goel, Ajay

2013-01-01

Aim Recent evidence suggests that several dietary polyphenols may exert their chemopreventive effect through epigenetic modifications. Curcumin is one of the most widely studied dietary chemopreventive agents for colon cancer prevention, however, its effects on epigenetic alterations, particularly DNA methylation, remain unclear. Using systematic genome-wide approaches, we aimed to elucidate the effect of curcumin on DNA methylation alterations in colorectal cancer cells. Materials and Methods To evaluate the effect of curcumin on DNA methylation, three CRC cell lines, HCT116, HT29 and RKO, were treated with curcumin. 5-aza-2′-deoxycytidine (5-aza-CdR) and trichostatin A treated cells were used as positive and negative controls for DNA methylation changes, respectively. Methylation status of LINE-1 repeat elements, DNA promoter methylation microarrays and gene expression arrays were used to assess global methylation and gene expression changes. Validation was performed using independent microarrays, quantitative bisulfite pyrosequencing, and qPCR. Results As expected, genome-wide methylation microarrays revealed significant DNA hypomethylation in 5-aza-CdR-treated cells (mean β-values of 0.12), however, non-significant changes in mean β-values were observed in curcumin-treated cells. In comparison to mock-treated cells, curcumin-induced DNA methylation alterations occurred in a time-dependent manner. In contrast to the generalized, non-specific global hypomethylation observed with 5-aza-CdR, curcumin treatment resulted in methylation changes at selected, partially-methylated loci, instead of fully-methylated CpG sites. DNA methylation alterations were supported by corresponding changes in gene expression at both up- and down-regulated genes in various CRC cell lines. Conclusions Our data provide previously unrecognized evidence for curcumin-mediated DNA methylation alterations as a potential mechanism of colon cancer chemoprevention. In contrast to non-specific global hypomethylation induced by 5-aza-CdR, curcumin-induced methylation changes occurred only in a subset of partially-methylated genes, which provides additional mechanistic insights into the potent chemopreventive effect of this dietary nutraceutical. PMID:23460897

Keratin 23 promotes telomerase reverse transcriptase expression and human colorectal cancer growth.

PubMed

Zhang, Ningning; Zhang, Rui; Zou, Kun; Yu, Wendan; Guo, Wei; Gao, Yingying; Li, Jia; Li, Mei; Tai, Yidi; Huang, Wenlin; Song, Chun; Deng, Wuguo; Cui, Xiaonan

2017-07-27

The overexpression of human telomerase reverse transcriptase (hTERT) has been associated with the proliferation and migration of colorectal cancer (CRC) cells. We investigated the roles of KRT23 and hTERT in promoting CRC cell proliferation and migration. We verified the relationship between KRT23 and hTERT in CRC using streptavidin-agarose pulldown and chromatin immunoprecipitation (ChIP) assays. One hundred and fifty-four human CRC specimens were analyzed using immunohistochemistry. The roles of KRT23 and hTERT in cell growth and migration were studied using siRNA and lentiviruses in vivo and in vitro. Western blot and wound scratch analyses were used to determine the signaling pathway for KRT23-mediated activation of CRC growth and migration. Telomerase activity was measured by using the TeloTAGGG Telomerase PCR ELISA PLUS Kit. We identified KRT23 as a new hTERT promoter-binding protein. Patients with high KRT23 and hTERT expression had markedly shorter overall survival. Overexpression of KRT23 upregulated the expression of hTERT protein, hTERT promoter-driven luciferase and telomerase activity in CRC. Conversely, inhibition of KRT23 by a KRT23-specific siRNA repressed the endogenous hTERT protein, the expression of hTERT promoter-driven luciferase and telomerase activity. Overexpression of KRT23 also promoted CRC proliferation and migration. By contrast, KRT23 inhibition significantly inhibited tumor cell growth in vitro and in vivo. KRT23 promoted cancer stem cell properties and increased the expression of CD133 and CD44. These results demonstrate that KRT23 is an important cellular factor that promotes CRC growth by activating hTERT expression and that KRT23 is a potential novel therapeutic target for CRC.

Bergamot juice extract inhibits proliferation by inducing apoptosis in human colon cancer cells.

PubMed

Visalli, Giuseppa; Ferlazzo, Nadia; Cirmi, Santa; Campiglia, Pietro; Gangemi, Sebastiano; Di Pietro, Angela; Calapai, Gioacchino; Navarra, Michele

2014-01-01

Colorectal cancer (CRC) is a leading cause of cancer mortality in the industrialized world, second to lung cancer. A lot of evidences highlight that a diet rich in fruits and vegetables may reduce the risk of some types of cancer including CRC. In this study we demonstrate that Citrus bergamia juice extracts (BJe) reduces CRC cell growth by multiple mechanisms. Low BJe concentrations inhibit MAPKs pathway and alter apoptosis-related proteins, that in turn induce cell cycle arrest and apoptosis in HT-29 cells. Instead, high concentrations of BJe induce oxidative stress causing DNA damage. Our study highlights the role of BJe as modulator of cell apoptosis in CRC cells and strengthens our previous hypothesis that the flavonoid fraction of bergamot juice may play a role as anti-cancer drug.

Physcion induces mitochondria-driven apoptosis in colorectal cancer cells via downregulating EMMPRIN.

PubMed

Chen, Xuehong; Gao, Hui; Han, Yantao; Ye, Junli; Xie, Jing; Wang, Chunbo

2015-10-05

Physcion, an anthraquinone derivative widely isolated and characterized from both terrestrial and marine sources, has anti-tumor effects on a variety of carcinoma cells, mainly through inhibition of cell proliferation, apoptosis induction and cell cycle arrest. However, little is known about the mechanisms underlying its role in tumor progression. In the present study, we investigated the molecular mechanisms involved in physcion-induced apoptosis in human colorectal cancer (CRC) lines HCT116. Our results showed that physcion inhibited tumor cell viability in a dose- and time-dependent manner, and induced cell apoptosis via intrinsic mitochondrial pathway. Our results also revealed that physcion treatment significantly inhibited extracelluar matrix metalloproteinase inducer (EMMPRIN) expression in HCT116 cells in a dose-dependent manner and overexpression of EMMPRIN protein markedly reduced physcion-induced cell apoptosis. Furthermore, our results strongly indicated the modulating effect of physcion on EMMPRIN is correlated with AMP-activated protein kinase (AMPK)/Hypoxia-inducible factor 1α (HIF-1α) signaling pathway. Our data provide the first experimental evidence that physcion induces mitochondrial apoptosis in CRC cells by downregulating of EMMPRIN via AMPK/HIF-1α signaling pathway and suggest a new mechanism to explain its anti-tumor effects. Copyright © 2015 Elsevier B.V. All rights reserved.

PD-L2 expression in colorectal cancer: Independent prognostic effect and targetability by deglycosylation.

PubMed

Wang, Huanbin; Yao, Han; Li, Chushu; Liang, Lunxi; Zhang, Yao; Shi, Hubing; Zhou, Chongzhi; Chen, Yingxuan; Fang, Jing-Yuan; Xu, Jie

2017-01-01

Colorectal cancer (CRC) is the second leading cause of cancer death worldwide, and immune checkpoint blockade therapy provides an opportunity for improving the outcome of CRC patients. Recent studies suggest that programmed death ligand-1 (PD-L1) is only expressed in 12% of CRCs. Here, we demonstrate that PD-L2 is expressed in approximately 40% CRCs, and its expression independently associates with poor survival of CRC patients. By detection of PD-L2 expression by immunofluorescence in 124 CRC cases with 10-y survival data, we found significant association between PD-L2 overexpression in cancer cells and worse overall survival (46.3 vs 69.1 mo; p = 0.0004). The association remained significant in multivariate COX regression analysis (hazard ratio = 2.778, 95% confidence interval [CI] = 1.668-4.627; p < 0.0001). In the validation CRC data set, significant association between PD-L2 overexpression and poor survival was supported by the univariate analysis (27.1 vs. 88.9 mo; p = 0.0002) and multivariate model (hazard ratio = 7.09, 95%CI 1.78-28.16; p = 0.005). Western Blot revealed strong induction of PD-L2 expression by interferon-γ (IFNγ) in CRC cells, and the mRNA levels of both genes were significantly correlated in CRC tissue samples. Suppression of glycosylation with tunicamycin caused a shift in molecular weight and significant decrease in the expression of PD-L2 protein. In conclusion, PD-L2 overexpression in CRC cells, under the regulation by IFNγ and glycosylation, associates with poor survival of patients with colorectal cancer. These findings highlight PD-L2 as a promising therapeutic target in CRC and suggest potential routes to control PD-L2 expression in CRC cells.

PD-L2 expression in colorectal cancer: Independent prognostic effect and targetability by deglycosylation

PubMed Central

Wang, Huanbin; Yao, Han; Li, Chushu; Liang, Lunxi; Zhang, Yao; Shi, Hubing; Zhou, Chongzhi; Chen, Yingxuan; Fang, Jing-Yuan

2017-01-01

ABSTRACT Colorectal cancer (CRC) is the second leading cause of cancer death worldwide, and immune checkpoint blockade therapy provides an opportunity for improving the outcome of CRC patients. Recent studies suggest that programmed death ligand-1 (PD-L1) is only expressed in 12% of CRCs. Here, we demonstrate that PD-L2 is expressed in approximately 40% CRCs, and its expression independently associates with poor survival of CRC patients. By detection of PD-L2 expression by immunofluorescence in 124 CRC cases with 10-y survival data, we found significant association between PD-L2 overexpression in cancer cells and worse overall survival (46.3 vs 69.1 mo; p = 0.0004). The association remained significant in multivariate COX regression analysis (hazard ratio = 2.778, 95% confidence interval [CI] = 1.668–4.627; p < 0.0001). In the validation CRC data set, significant association between PD-L2 overexpression and poor survival was supported by the univariate analysis (27.1 vs. 88.9 mo; p = 0.0002) and multivariate model (hazard ratio = 7.09, 95%CI 1.78–28.16; p = 0.005). Western Blot revealed strong induction of PD-L2 expression by interferon-γ (IFNγ) in CRC cells, and the mRNA levels of both genes were significantly correlated in CRC tissue samples. Suppression of glycosylation with tunicamycin caused a shift in molecular weight and significant decrease in the expression of PD-L2 protein. In conclusion, PD-L2 overexpression in CRC cells, under the regulation by IFNγ and glycosylation, associates with poor survival of patients with colorectal cancer. These findings highlight PD-L2 as a promising therapeutic target in CRC and suggest potential routes to control PD-L2 expression in CRC cells. PMID:28811964

Temporal and spatial changes of cells positive for stem-like markers in different compartments and stages of human colorectal adenoma-carcinoma sequence

PubMed Central

Cui, Guanglin; Xu, Gang; Zhu, Li; Pang, Zhigang; Zheng, Wei; Li, Zhenfeng; Yuan, Aping

2017-01-01

Considerable evidence supports the idea that stem-like cells may play an essential role during the development of colorectal cancer (CRC). To accomplish this aim, we use immunohistochemistry (IHC) and double IHC with different potential stem-like markers, anti-musashi (Msi), anti-CD133, anti- LGR5 and anti-ALDH1 to examine the presentation of stem-like cells in different compartments including adenoma/CRC epithelium, transitional crypts and tumor stroma in colorectal adenoma and CRC. The results showed that cells positive for stem-like markers were remarkably increased in number and frequently observed in the adenoma/CRC epithelium, transitional crypts and tumor stroma. Notably, the population of cells positive for stem-liker markers was expanded from the base to the middle part of the transitional crypt in both adenoma and CRC tissues, reflecting that stem-like cells are likely involved in the process of colorectal tumorigenesis. Counting results showed that the grading scores of cells positive for LGR5 and ALDH1 in the adenoma/CRC epithelium were significantly increased relative with the control epithelium, and associated with the degree of dysplasia in the adenoma and node involvement in the CRC (all P < 0.05). In addition, the density of cells positive for stem–like markers in the adenomatous/cancerous stroma was also increased and paralleled an increase in the density of proliferative stromal cells labeled by PCNA, which were primarily identified as vimentin positive fibroblasts. Our results have revealed a changed temporal and spatial presentation of stem-like markers in different stages of human colorectal adenoma-carcinoma sequence, which might be a hallmark of the adenoma-carcinoma transition. PMID:28484082

Upregulation of miR-598 promotes cell proliferation and cell cycle progression in human colorectal carcinoma by suppressing INPP5E expression

PubMed Central

Li, Kun-Ping; Fang, Yong-Ping; Liao, Jin-Qi; Duan, Jin-Dong; Feng, Li-Guang; Luo, Xiao-Zai; Liang, Zhi-Jian

2018-01-01

Colorectal cancer (CRC) is one of the most common types of cancer worldwide. Recently, microRNAs (miRs) have been considered as novel therapeutic targets for the treatment of cancer. miR-598 is a poorly investigated miR. The underlying mechanism of miR-598 in CRC cells remains to be elucidated. In the present study, miR-598 was demonstrated to be significantly upregulated in CRC tissue by analyzing data from The Cancer Genome Atlas and the Gene Expression Omnibus. The results of a polymerase chain reaction demonstrated that miR-598 expression was significantly upregulated in CRC tissues and cells. Gain of function and loss of function assays demonstrated that miR-598 significantly promoted cell proliferation and cell cycle progression. miR-598 was demonstrated to modulate cell functions by regulating 72 kDa inositol polyphosphate-5-phosphatase (INPP5E). In addition, knockdown of INPP5E counteracted the growth arrest caused by an miR-598-inhibitor. In conclusion, the present study demonstrated that miR-598 contributed to cell proliferation and cell cycle progression in CRC by targeting INPP5E. PMID:29257251

Deletions at SLC18A1 increased the risk of CRC and lower SLC18A1 expression associated with poor CRC outcome.

PubMed

Zhang, Dandan; Li, Zhenli; Xu, Xiaohong; Zhou, Dan; Tang, Shunli; Yin, Xiaoyang; Xu, Fangying; Li, Hui; Zhou, Yuan; Zhu, Tao; Deng, Hong; Zhang, Shuai; Huang, Qiong; Wang, Jing; Yin, Wei; Zhu, Yimin; Lai, Maode

2017-10-26

Copy number variations (CNVs) contribute to the development of colorectal cancer (CRC). We conducted a two-stage association study to identify CNV risk loci for CRC. We performed a gene-based rare CNV study on 694 sporadic CRC and 1641 controls using Illumina Human-OmniExpress-12v1.0 BeadChips, and further replicated in 934 CRC cases and 2680 controls for risk CNVs by using TaqMan Copy Number Assay. Tumor buddings, cancer cells in the center of primary tumor and normal intestinal epithelial cells were captured using laser capture microdissection (LCM) and were assayed using AffymetrixGeneChip® Human Genome U133 Plus 2.0 Array. In addition, The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus data were assessed for the effects of risk CNVs. We found that germline deletions affecting the last six exons of SLC18A1 significantly associated with CRC with a combined P value of 6.4 × 10-5 by a two-stage analysis. Both in TCGA CRC RNA seq dataset and GDS4382, SLC18A1 was significantly down regulated in CRC tissues than in paired normal tissues (N = 32 and 17 pairs, P = 0.004 and 0.009, respectively). In LCM samples, similar observations were obtained that the expression levels of SLC18A1 in the tumor buddings, cancer cells in the center of primary tumor, and stroma of both tumor budding and cancer cells were lower than normal intestinal epithelial and stromal cells (fold change = 0.17-0.62, 0.12-0.57 and 0.37-0.68, respectively). In summary, the germline deletions at SLC18A1 contributed to the development of CRC. The role of SLC18A1 required further exploration. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Molecular profiling of ALDH1+ colorectal cancer stem cells reveals preferential activation of MAPK, FAK, and oxidative stress pro-survival signalling pathways

PubMed Central

Vishnubalaji, Radhakrishnan; Manikandan, Muthurangan; Fahad, Mohamed; Hamam, Rimi; Alfayez, Musaad; Kassem, Moustapha; Aldahmash, Abdullah; Alajez, Nehad M.

2018-01-01

Tumour heterogeneity leads to variable clinical response and inaccurate diagnostic and prognostic assessment. Cancer stem cells (CSCs) represent a subpopulation responsible for invasion, metastasis, therapeutic resistance, and recurrence in many human cancer types. However, the true identity of colorectal cancer (CRC) SCs remains elusive. Here, we aimed to characterize and define the gene expression portrait of CSCs in CRC-model SW403 cells. We found that ALDH+ positive cells are clonogenic and highly proliferative; their global gene expression profiling-based molecular signature revealed gene enrichment related to DNA damage, MAPK, FAK, oxidative stress response, and Wnt signalling. ALDH+ cells showed enhanced ROS stress resistance, whereas MAPK/FAK pathway pharmacologic inhibition limited their survival. Conversely, 5-fluorouracil increased the ALDH+ cell fraction among the SW403, HCT116 and SW620 CRC models. Notably, analysis of ALDH1A1 and POU5F1 expression levels in cohorts of 462 or 420 patients for overall (OS) or disease-free (DFS) survival, respectively, obtained from the Cancer Genome Atlas CRC dataset, revealed strong association between elevated expression and poor OS (p = 0.006) and poor DFS (p = 0.05), thus implicating ALDH1A1 and POU5F1 in CRC prognosis. Our data reveal distinct molecular signature of ALDH+ CSCs in CRC and suggest pathways relevant for successful targeted therapies and management of CRC. PMID:29568377

Polo-like kinase 1 expression is suppressed by CCAAT/enhancer-binding protein α to mediate colon carcinoma cell differentiation and apoptosis.

PubMed

Dasgupta, Nirmalya; Thakur, Bhupesh Kumar; Ta, Atri; Das, Sayan; Banik, George; Das, Santasabuj

2017-07-01

Human polo-like kinase 1 (PLK1), a highly conserved serine/threonine kinase is a key player in several essential cell-cycle events. PLK1 is considered an oncogene and its overexpression often correlates with poor prognosis of cancers, including colorectal cancer (CRC). However, regulation of PLK1 expression in colorectal cells was never studied earlier and it is currently unknown if PLK1 regulates differentiation and apoptosis of CRC. PLK1 expression was analyzed by real-time PCR and western blotting. Transcriptional regulation was studied by reporter assay, gene knock-down, EMSA and ChIP. PLK1 expression was down-regulated during butyrate-induced differentiation of HT-29 and other CRC cells. Also, PLK1 down-regulation mediated the role of butyrate in CRC differentiation and apoptosis. We report here a novel transcriptional regulation of PLK1 by butyrate. Transcription factors CCAAT/enhancer-binding protein α (C/EBPα) and Oct-1 share an overlapping binding site over the PLK1 promoter. Elevated levels of C/EBPα by butyrate treatment of CRC cells competed out the activator protein Oct-1 from binding to the PLK1 promoter and sequestered it. Binding of C/EBPα was associated with increased deacetylation near the transcription start site (TSS) of the PLK1 promoter, which abrogated transcription through reduced recruitment of RNA polymerase II. We also found a synergistic role between the synthetic PLK1-inhibitor SBE13 and butyrate on the apoptosis of CRC cells. This study offered a novel p53-independent regulation of PLK1 during CRC differentiation and apoptosis. Down-regulation of PLK1 is one of the mechanisms underlying the anti-cancer role of dietary fibre-derived butyrate in CRC. Copyright © 2017 Elsevier B.V. All rights reserved.

Rosmarinic Acid Activates AMPK to Inhibit Metastasis of Colorectal Cancer

PubMed Central

Han, Yo-Han; Kee, Ji-Ye; Hong, Seung-Heon

2018-01-01

Rosmarinic acid (RA) has been used as an anti-inflammatory, anti-diabetic, and anti-cancer agent. Although RA has also been shown to exert an anti-metastatic effect, the mechanism of this effect has not been reported to be associated with AMP-activated protein kinase (AMPK). The aim of this study was to elucidate whether RA could inhibit the metastatic properties of colorectal cancer (CRC) cells via the phosphorylation of AMPK. RA inhibited the proliferation of CRC cells through the induction of cell cycle arrest and apoptosis. In several metastatic phenotypes of CRC cells, RA regulated epithelial–mesenchymal transition (EMT) through the upregulation of an epithelial marker, E-cadherin, and the downregulation of the mesenchymal markers, N-cadherin, snail, twist, vimentin, and slug. Invasion and migration of CRC cells were inhibited and expressions of matrix metalloproteinase (MMP)-2 and MMP-9 were decreased by RA treatment. Adhesion and adhesion molecules such as ICAM-1 and integrin β1 expressions were also reduced by RA treatment. In particular, the effects of RA on EMT and MMPs expressions were due to the activation of AMPK. Moreover, RA inhibited lung metastasis of CRC cells by activating AMPK in mouse model. Collectively, these results proved that RA could be potential therapeutic agent against metastasis of CRC. PMID:29459827

The distinct role of strand-specific miR-514b-3p and miR-514b-5p in colorectal cancer metastasis.

PubMed

Ren, Lin-Lin; Yan, Ting-Ting; Shen, Chao-Qin; Tang, Jia-Yin; Kong, Xuan; Wang, Ying-Chao; Chen, Jinxian; Liu, Qiang; He, Jie; Zhong, Ming; Chen, Hao-Yan; Hong, Jie; Fang, Jing-Yuan

2018-06-07

The abnormal expression of microRNAs (miRNAs) in colorectal cancer (CRC) progression has been widely investigated. It was reported that the same hairpin RNA structure could generate mature products from each strand, termed 5p and 3p, which binds different target mRNAs. Here, we explored the expression, functions, and mechanisms of miR-514b-3p and miR-514b-5p in CRC cells and tissues. We found that miR-514b-3p was significantly down-regulated in CRC samples, and the ratio of miR-514b-3p/miR-514b-5p increased from advanced CRC, early CRC to matched normal colorectal tissues. Follow-up functional experiments illustrated that miR-514b-3p and miR-514b-5p had distinct effects through interacting with different target genes: MiR-514b-3p reduced CRC cell migration, invasion and drug resistance through increasing epithelial marker and decreasing mesenchymal marker expressions, conversely, miR-514b-5p exerted its pro-metastatic properties in CRC by promoting EMT progression. MiR-514b-3p overexpressing CRC cells developed tumors more slowly in mice compared with control cells, however, miR-514b-5p accelerated tumor metastasis. Overall, our data indicated that though miR-514b-3p and miR-514b-5p were transcribed from the same RNA hairpin, each microRNA has distinct effect on CRC metastasis.

Activated human primary NK cells efficiently kill colorectal cancer cells in 3D spheroid cultures irrespectively of the level of PD-L1 expression.

PubMed

Lanuza, Pilar M; Vigueras, Alan; Olivan, Sara; Prats, Anne C; Costas, Santiago; Llamazares, Guillermo; Sanchez-Martinez, Diego; Ayuso, José María; Fernandez, Luis; Ochoa, Ignacio; Pardo, Julián

2018-01-01

Haploidentical Natural Killer (NK) cells have been shown as an effective and safe alternative for the treatment of haematological malignancies with poor prognosis for which traditional therapies are ineffective. In contrast to haematological cancer cells, that mainly grow as single suspension cells, solid carcinomas are characterised by a tridimensional (3D) architecture that provide specific surviving advantages and resistance against chemo- and radiotherapy. However, little is known about the impact of 3D growth on solid cancer immunotherapy especially adoptive NK cell transfer. We have recently developed a protocol to activate ex vivo human primary NK cells using B lymphoblastic cell lines, which generates NK cells able to overcome chemoresistance in haematological cancer cells. Here we have analysed the activity of these allogeneic NK cells against colorectal (CRC) human cell lines growing in 3D spheroid culture and correlated with the expression of some of the main ligands regulating NK cell activity. Our results indicate that activated NK cells efficiently kill colorectal tumour cell spheroids in both 2D and 3D cultures. Notably, although 3D CRC cell cultures favoured the expression of the inhibitory immune checkpoint PD-L1, it did not correlate with increased resistance to NK cells. Finally, we have analysed in detail the infiltration of NK cells in 3D spheroids by microscopy and found that at low NK cell density, cell death is not observed although NK cells are able to infiltrate into the spheroid. In contrast, higher densities promote tumoural cell death before infiltration can be detected. These findings show that highly dense activated human primary NK cells efficiently kill colorectal carcinoma cells growing in 3D cultures independently of PD-L1 expression and suggest that the use of allogeneic activated NK cells could be beneficial for the treatment of colorectal carcinoma.

Mapping the HLA ligandome of Colorectal Cancer Reveals an Imprint of Malignant Cell Transformation.

PubMed

Löffler, Markus W; Kowalewski, Daniel J; Backert, Linus; Bernhardt, Jörg; Adam, Patrick; Schuster, Heiko; Dengler, Florian; Backes, Daniel; Kopp, Hans-Georg; Beckert, Stefan; Wagner, Silvia; Königsrainer, Ingmar; Kohlbacher, Oliver; Kanz, Lothar; Königsrainer, Alfred; Rammensee, Hans-Georg; Stevanovic, Stefan; Haen, Sebastian P

2018-05-22

Immune cell infiltrates have proven highly relevant for colorectal carcinoma (CRC) prognosis, making CRC a promising candidate for immunotherapy. Since tumors interact with the immune system via HLA-presented peptide ligands, exact knowledge of the peptidome constitution is fundamental for understanding this relationship. Here we comprehensively describe the naturally presented HLA-ligandome of CRC and corresponding non-malignant colon (NMC) tissue. Mass spectrometry identified 35,367 and 28,132 HLA-class I ligands on CRC and NMC, attributable to 7,684 and 6,312 distinct source proteins, respectively. Cancer-exclusive peptides were assessed on source protein level using Kyoto Encyclopedia of Genes and Genomes (KEGG) and protein analysis through evolutionary relationships (PANTHER), revealing pathognomonic CRC-associated pathways including Wnt, TGF-β, PI3K, p53, and RTK-RAS. Relative quantitation of peptide presentation on paired CRC and NMC tissue further identified source proteins from cancer- and infection-associated pathways to be over-represented merely within the CRC ligandome. From the pool of tumor-exclusive peptides, a selected HLA-ligand subset was assessed for immunogenicity, with the majority exhibiting an existing T cell repertoire. Overall, these data show that the HLA-ligandome reflects cancer-associated pathways implicated in CRC oncogenesis, suggesting that alterations in tumor cell metabolism could result in cancer-specific, albeit not mutation-derived tumor-antigens. Hence, a defined pool of unique tumor peptides, attributable to complex cellular alterations that are exclusive to malignant cells might comprise promising candidates for immunotherapeutic applications. Copyright ©2018, American Association for Cancer Research.

High Myeloperoxidase Positive Cell Infiltration in Colorectal Cancer Is an Independent Favorable Prognostic Factor

PubMed Central

Eppenberger-Castori, Serenella; Zlobec, Inti; Viehl, Carsten T.; Frey, Daniel M.; Nebiker, Christian A.; Rosso, Raffaele; Zuber, Markus; Amicarella, Francesca; Iezzi, Giandomenica; Sconocchia, Giuseppe; Heberer, Michael; Lugli, Alessandro; Tornillo, Luigi; Oertli, Daniel

2013-01-01

Background Colorectal cancer (CRC) infiltration by adaptive immune system cells correlates with favorable prognosis. The role of the innate immune system is still debated. Here we addressed the prognostic impact of CRC infiltration by neutrophil granulocytes (NG). Methods A TMA including healthy mucosa and clinically annotated CRC specimens (n = 1491) was stained with MPO and CD15 specific antibodies. MPO+ and CD15+ positive immune cells were counted by three independent observers. Phenotypic profiles of CRC infiltrating MPO+ and CD15+ cells were validated by flow cytometry on cell suspensions derived from enzymatically digested surgical specimens. Survival analysis was performed by splitting randomized data in training and validation subsets. Results MPO+ and CD15+ cell infiltration were significantly correlated (p<0.0001; r = 0.76). However, only high density of MPO+ cell infiltration was associated with significantly improved survival in training (P = 0.038) and validation (P = 0.002) sets. In multivariate analysis including T and N stage, vascular invasion, tumor border configuration and microsatellite instability status, MPO+ cell infiltration proved an independent prognostic marker overall (P = 0.004; HR = 0.65; CI:±0.15) and in both training (P = 0.048) and validation (P = 0.036) sets. Flow-cytometry analysis of CRC cell suspensions derived from clinical specimens showed that while MPO+ cells were largely CD15+/CD66b+, sizeable percentages of CD15+ and CD66b+ cells were MPO−. Conclusions High density MPO+ cell infiltration is a novel independent favorable prognostic factor in CRC. PMID:23734221

A complex of α6 integrin and E-cadherin drives liver metastasis of colorectal cancer cells through hepatic angiopoietin-like 6

PubMed Central

Marchiò, Serena; Soster, Marco; Cardaci, Sabrina; Muratore, Andrea; Bartolini, Alice; Barone, Vanessa; Ribero, Dario; Monti, Maria; Bovino, Paola; Sun, Jessica; Giavazzi, Raffaella; Asioli, Sofia; Cassoni, Paola; Capussotti, Lorenzo; Pucci, Piero; Bugatti, Antonella; Rusnati, Marco; Pasqualini, Renata; Arap, Wadih; Bussolino, Federico

2012-01-01

Homing of colorectal cancer (CRC) cells to the liver is a non-random process driven by a crosstalk between tumour cells and components of the host tissue. Here we report the isolation of a liver metastasis-specific peptide ligand (CGIYRLRSC) that binds a complex of E-cadherin and α6 integrin on the surface of CRC cells. We identify angiopoietin-like 6 protein as a peptide-mimicked natural ligand enriched in hepatic blood vessels of CRC patients. We demonstrate that an interaction between hepatic angiopoietin-like 6 and tumoural α6 integrin/E-cadherin drives liver homing and colonization by CRC cells, and that CGIYRLRSC inhibits liver metastasis through interference with this ligand/receptor system. Our results indicate a mechanism for metastasis whereby a soluble factor accumulated in normal vessels functions as a specific ligand for circulating cancer cells. Consistently, we show that high amounts of coexpressed α6 integrin and E-cadherin in primary tumours represent a poor prognostic factor for patients with advanced CRC. PMID:23070965

Prognostic, predictive and pharmacogenomic assessments of CDX2 refine stratification of colorectal cancer.

PubMed

Bruun, Jarle; Sveen, Anita; Barros, Rita; Eide, Peter W; Eilertsen, Ina; Kolberg, Matthias; Pellinen, Teijo; David, Leonor; Svindland, Aud; Kallioniemi, Olli; Guren, Marianne G; Nesbakken, Arild; Almeida, Raquel; Lothe, Ragnhild A

2018-06-14

We aimed to refine the value of CDX2 as an independent prognostic and predictive biomarker in colorectal cancer (CRC) according to disease stage and chemotherapy sensitivity in preclinical models. CDX2 expression was evaluated in 1045 stage I-IV primary CRCs by gene expression (n=403) or immunohistochemistry (n=642) and in relation to 5-year relapse-free survival (RFS), overall survival (OS), and chemotherapy. Pharmacogenomic associations between CDX2 expression and 69 chemotherapeutics were assessed by drug screening of 35 CRC cell lines. CDX2 expression was lost in 11.6% of cases and showed independent poor prognostic value in multivariable models. For individual stages, CDX2 was prognostic only in stage IV, independent of chemotherapy. Among stage I-III patients not treated in an adjuvant setting, CDX2 loss was associated with a particularly poor survival in the BRAF-mutated subgroup, but prognostic value was independent of microsatellite instability status and the consensus molecular subtypes In stage III, the 5-year RFS rate was higher among patients with loss of CDX2 who received adjuvant chemotherapy than among patients who did not. The CDX2-negative cell lines were significantly more sensitive to chemotherapeutics than CDX2-positive cells, and the multidrug resistance genes MDR1 and CFTR were significantly downregulated both in CDX2-negative cells and patient tumors. Molecular Oncology (2018) © 2018 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

Low levels of Caspase-3 predict favourable response to 5FU-based chemotherapy in advanced colorectal cancer: Caspase-3 inhibition as a therapeutic approach.

PubMed

Flanagan, L; Meyer, M; Fay, J; Curry, S; Bacon, O; Duessmann, H; John, K; Boland, K C; McNamara, D A; Kay, E W; Bantel, H; Schulze-Bergkamen, H; Prehn, J H M

2016-02-04

Colorectal cancer (CRC) is one of the most common cancers in the Western world. 5-Fluorouracil (5FU)-based chemotherapy (CT) remains the mainstay treatment of CRC in the advanced setting, and activates executioner caspases in target cells. Executioner caspases are key proteins involved in cell disassembly during apoptosis. Activation of executioner caspases also has a role in tissue regeneration and repopulation by stimulating signal transduction and cell proliferation in neighbouring, non-apoptotic cells as reported recently. Tissue microarrays (TMAs) consisting of tumour tissue from 93 stage II and III colon cancer patients were analysed by immunohistochemistry. Surprisingly, patients with low levels of active Caspase-3 had an increased disease-free survival time. This was particularly pronounced in patients who received 5FU-based adjuvant CT. In line with this observation, lower serum levels of active Caspase-3 were found in patients with metastasised CRC who revealed stable disease or tumour regression compared with those with disease progression. The role of Caspase-3 in treatment responses was explored further in primary human tumour explant cultures from fresh patient tumour tissue. Exposure of explant cultures to 5FU-based CT increased the percentage of cells positive for active Caspase-3 and Terminal Deoxynucleotidyl Transferase dUTP Nick end Labelling (TUNEL), but also the expression of regeneration and proliferation markers β-Catenin and Ki-67, as well as cyclooxygenase-2 (COX-2). Of note, selective inhibition of Caspase-3 with Ac-DNLD-CHO, a selective, reversible inhibitor of Caspase-3, significantly reduced the expression of proliferation markers as well as COX-2. Inhibition of COX-2 with aspirin or celecoxib did not affect Caspase-3 levels but also reduced Ki-67 and β-Catenin levels, suggesting that Caspase-3 acted via COX-2 to stimulate cell proliferation and tissue regeneration. This indicates that low levels of active Caspase-3 may represent a new predictor of CT responsiveness, and inhibition of Caspase-3, or antagonising downstream effectors of Caspase-3 paracrine signalling, such as COX-2 may improve patient outcomes following CT in advanced CRC.

Non-coding effects of circular RNA CCDC66 promote colon cancer growth and metastasis

PubMed Central

Hsiao, Kuei-Yang; Lin, Ya-Chi; Gupta, Sachin Kumar; Chang, Ning; Yen, Laising; Sun, H. Sunny; Tsai, Shaw-Jenq

2018-01-01

Circular RNA (circRNA) is a class of non-coding RNA whose functions remain mostly unknown. Recent studies indicate circRNA may be involved in disease pathogenesis, but direct evidence is scarce. Here we characterize the functional role of a novel circRNA, circCCDC66, in colorectal cancer (CRC). RNA-Seq data from matched normal and tumor colon tissue samples identified numerous circRNAs specifically elevated in cancer cells, several of which were verified by quantitative RT-PCR. CircCCDC66 expression was elevated in polyps and colon cancer and was associated with poor prognosis. Gain-of-function and loss-of-function studies in CRC cell-lines demonstrated that circCCDC66 controlled multiple pathological processes, including cell proliferation, migration, invasion, and anchorage-independent growth. In-depth characterization revealed that circCCDC66 exerts its function via regulation of a subset of oncogenes, and knockdown of circCCDC66 inhibited tumor growth and cancer invasion in xenograft and orthotopic mouse models, respectively. Taken together, these findings highlight a novel oncogenic function of circRNA in cancer progression and metastasis. PMID:28249903

Up-regulated EMMPRIN/CD147 protein expression might play a role in colorectal carcinogenesis and its subsequent progression without an alteration of its glycosylation and mRNA level.

PubMed

Zheng, Hua-chuan; Wang, Wei; Xu, Xiao-yan; Xia, Pu; Yu, Miao; Sugiyama, Toshiro; Takano, Yasuo

2011-04-01

Extracellular matrix metalloproteinase inducer (EMMPRIN) was reported to involve in the invasion and metastasis of malignancies by regulating the expression of vascular endothelial growth factor (VEGF) in stromal and cancer cells. The study aimed to clarify the role of EMMPRIN expression in tumorigenesis and progression of colorectal carcinomas (CRC). EMMPRIN expression was examined on tissue microarray containing colorectal carcinomas, adenoma and non-neoplastic mucosa (NNM) by immunohistochemistry and in situ hybridization (ISH). Colorectal carcinoma cell lines (DLD-1, HCT-15, SW480 and WiDr) and tissues were studied for EMMPRIN expression by Western blot or RT-PCR, followed by sequencing. All carcinoma cell lines showed EMMPRIN expression at both mRNA and protein levels. Two synonymous mutations were found in carcinoma cell lines at codon109 (GCT → GCC: Ala) or 179 (GAT → GAC: Asp). Frozen CRC tissues displayed higher EMMPRIN expression than paired NNM (P < 0.05). EMMPRIN expression was immunohistochemically stronger in colorectal high-grade adenoma, adenocarcinoma and metastatic carcinoma than non-neoplastic superficial epithelium and low-grade adenoma (P < 0.05). In contrast, its mRNA level was similar from colorectal NNM, adenoma to adenocarcinoma by ISH, in line with the findings of RT-PCR (P > 0.05). Immunohistochemically, EMMPRIN expression was positively correlated with tumor size, depth of invasion, vascular or lymphatic invasion, grade of infiltration (INF), ki-67 and VEGF expression of CRCs (P < 0.05). Among them, depth of invasion was an independent associated factor for EMMPRIN expression in CRCs (P < 0.05). Up-regulated EMMPRIN protein expression might contribute to colorectal carcinogenesis without the alteration of its glycosylation and mRNA level. Aberrant EMMPRIN protein expression might promote growth or invasion of CRCs possibly through increased ki-67 expression and inducible angiogenesis via up-regulating VEGF expression.

Effect of Primary Tumor Location on Second- or Later-line Treatment Outcomes in Patients With RAS Wild-type Metastatic Colorectal Cancer and All Treatment Lines in Patients With RAS Mutations in Four Randomized Panitumumab Studies.

PubMed

Boeckx, Nele; Koukakis, Reija; Op de Beeck, Ken; Rolfo, Christian; Van Camp, Guy; Siena, Salvatore; Tabernero, Josep; Douillard, Jean-Yves; André, Thierry; Peeters, Marc

2018-03-08

The primary tumor location has a prognostic impact in metastatic colorectal cancer (mCRC). We report the results from retrospective analyses assessing the effect of tumor location on prognosis and efficacy of second- and later-line panitumumab treatment in patients with RAS wild-type (WT) mCRC and on prognosis in all lines of treatment in patients with RAS mutant (MT) mCRC. RAS WT data (n = 483) from 2 randomized phase III panitumumab trials (ClinicalTrials.gov identifiers, NCT00339183 and NCT00113763) were analyzed for treatment outcomes stratified by tumor location. The second analysis assessed the effect of tumor location in RAS MT patients (n = 1205) from 4 panitumumab studies (ClinicalTrials.gov identifiers, NCT00364013, NCT00819780, NCT00339183, and NCT00113763). Primary tumors located in the cecum to transverse colon were coded as right-sided; those located from the splenic flexure to the rectum were coded as left-sided. Of all patients, the tumor location was ascertained for 83% to 88%; 71% to 77% of patients had left-sided tumors. RAS WT patients with right-sided tumors did worse for all efficacy parameters compared with those with left-sided tumors. The patients with left-sided tumors had better outcomes with panitumumab than with the comparator treatment. Because of the low patient numbers, no conclusions could be drawn for right-sided mCRC. The prognostic effect of tumor location on survival was unclear for RAS MT patients. These retrospective analyses have confirmed that RAS WT right-sided mCRC is associated with a poor prognosis, regardless of the treatment. RAS WT patients with left-sided tumors benefitted from the addition of panitumumab in second or later treatment lines. Further research is warranted to determine the optimum management of right-sided mCRC and RAS MT tumors. Copyright © 2018 Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hogan, Niamh M.; Joyce, Myles R.; Murphy, J. Mary

Highlights: •MSCs were directly co-cultured with colorectal cancer (CRC) cells on 3D scaffolds. •MSCs influence CRC protein/gene expression, proliferation and migration. •We report a significant functional role of MSC-secreted PAI-1 in colon cancer. -- Abstract: Mesenchymal Stem Cells are known to engraft and integrate into the architecture of colorectal tumours, with little known regarding their fate following engraftment. This study aimed to investigate mediators of Mesenchymal Stem Cell (MSC) and colon cancer cell (CCC) interactions. Mesenchymal Stem Cells and colon cancer cells (HT29 and HCT-116) were cultured individually or in co-culture on 3-dimensional scaffolds. Conditioned media containing all secreted factorsmore » was harvested at day 1, 3 and 7. Chemokine secretion and expression were analyzed by Chemi-array, ELISA (Macrophage migration inhibitory factor (MIF), plasminogen activator inhibitor type 1 (PAI-1)) and RQ-PCR. Colon cancer cell migration and proliferation in response to recombinant PAI-1, MSCs and MSCs + antibody to PAI-1 was analyzed using Transwell inserts and an MTS proliferation assay respectively. Chemi-array revealed secretion of a wide range of factors by each cell population, including PAI-1and MIF. ELISA analysis revealed Mesenchymal Stem Cells to secrete the highest levels of PAI-1 (MSC mean 10.6 ng/mL, CCC mean 1.01 ng/mL), while colon cancer cells were the principal source of MIF. MSC-secreted PAI-1 stimulated significant migration of both CCC lines, with an antibody to the chemokine shown to block this effect (67–88% blocking,). A cell-line dependant effect on CCC proliferation was shown for Mesenchymal Stem Cell-secreted PAI-1 with HCT-116 cells showing decreased proliferation at all concentrations, and HT29 cells showing increased proliferation in the presence of higher PAI-1 levels. This is the first study to identify PAI-1 as an important mediator of Mesenchymal Stem Cell/colon cancer cell interactions and highlights the significant functional impact of Mesenchymal Stem Cell-secreted PAI-1 on colon cancer cells.« less

The infiltration, and prognostic importance, of Th1 lymphocytes vary in molecular subgroups of colorectal cancer

PubMed Central

Ling, Agnes; Lundberg, Ida V; Eklöf, Vincy; Wikberg, Maria L; Öberg, Åke; Palmqvist, Richard

2015-01-01

Abstract Giving strong prognostic information, T‐cell infiltration is on the verge of becoming an additional component in the routine clinical setting for classification of colorectal cancer (CRC). With a view to further improving the tools for prognostic evaluation, we have studied how Th1 lymphocyte infiltration correlates with prognosis not only by quantity, but also by subsite, within CRCs with different molecular characteristics (microsatellite instability, CpG island methylator phenotype status, and BRAF and KRAS mutational status). We evaluated the Th1 marker T‐bet by immunohistochemistry in 418 archival tumour tissue samples from patients who underwent surgical resection for CRC. We found that a high number of infiltrating Th1 lymphocytes is strongly associated with an improved prognosis in patients with CRC, irrespective of intratumoural subsite, and that both extent of infiltration and patient outcome differ according to molecular subgroup. In brief, microsatellite instability, CpG island methylator phenotype‐high and BRAF mutated tumours showed increased infiltration of Th1 lymphocytes, and the most pronounced prognostic effect of Th1 infiltration was found in these tumours. Interestingly, BRAF mutated tumours were found to be more highly infiltrated by Th1 lymphocytes than BRAF wild‐type tumours whereas the opposite was seen for KRAS mutated tumours. These differences could be explained at least partly by our finding that BRAF mutated, in contrast to KRAS mutated, CRC cell lines and tumour specimens expressed higher levels of the Th1‐attracting chemokine CXCL10, and reduced levels of CCL22 and TGFB1, stimulating Th2/Treg recruitment and polarisation. In conclusion, the strong prognostic importance of Th1 lymphocyte infiltration in CRC was found at all subsites evaluated, and it remained significant in multivariable analyses, indicating that T‐bet may be a valuable marker in the clinical setting. Our results also indicate that T‐bet is of value when analysed in molecular subgroups of CRC, allowing identification of patients with especially poor prognosis who are in need of extended treatment. PMID:27499912

HIF-1α activates hypoxia-induced BCL-9 expression in human colorectal cancer cells

PubMed Central

Chen, Tian-Rui; Wei, Hai-feng; Song, Dian-Wen; Liu, Tie-Long; Yang, Xing-Hai; Fu, Chuan-Gang; Hu, Zhi-qian; Zhou, Wang; Yan, Wang-Jun; Xiao, Jian-Ru

2017-01-01

B-cell CLL/lymphoma 9 protein (BCL-9), a multi-functional co-factor in Wnt signaling, induced carcinogenesis as well as promoting tumor progression, metastasis and chemo-resistance in colorectal cancer (CRC). However, the mechanisms for increased BCL-9 expression in CRC were not well understood. Here, we report that hypoxia, a hallmark of solid tumors, induced BCL-9 mRNA expression in human CRC cells. Analysis of BCL-9 promoter revealed two functional hypoxia-responsive elements (HRE-B and HRE-C) that can be specifically bound with and be transactivated by hypoxia inducible factors (HIF) -1α but not HIF-2α. Consistently, ectopic expression of HIF-1α but not HIF-2α transcriptionally induced BCL-9 expression levels in cells. Knockdown of endogenous HIF-1α but not HIF-2α by siRNA largely abolished the induction of HIF by hypoxia. Furthermore, there was a strong association of HIF-1α expression with BCL-9 expression in human CRC specimens. In summary, results from this study demonstrated that hypoxia induced BCL-9 expression in human CRC cells mainly through HIF-1α, which could be an important underlying mechanism for increased BCL-9 expression in CRC. PMID:27121066

CHK1-targeted therapy to deplete DNA replication-stressed, p53-deficient, hyperdiploid colorectal cancer stem cells

PubMed Central

Russo, Giorgio; Corradi, Francesca; Siteni, Silvia; Musella, Martina; Vitale, Sara; De Angelis, Maria Laura; Pallocca, Matteo; Amoreo, Carla Azzurra; Sperati, Francesca; Di Franco, Simone; Barresi, Sabina; Policicchio, Eleonora; De Luca, Gabriele; De Nicola, Francesca; Mottolese, Marcella; Zeuner, Ann; Fanciulli, Maurizio; Stassi, Giorgio; Maugeri-Saccà, Marcello; Baiocchi, Marta; Tartaglia, Marco

2018-01-01

Objective Cancer stem cells (CSCs) are responsible for tumour formation and spreading, and their targeting is required for tumour eradication. There are limited therapeutic options for advanced colorectal cancer (CRC), particularly for tumours carrying RAS-activating mutations. The aim of this study was to identify novel CSC-targeting strategies. Design To discover potential therapeutics to be clinically investigated as single agent, we performed a screening with a panel of FDA-approved or investigational drugs on primary CRC cells enriched for CSCs (CRC-SCs) isolated from 27 patients. Candidate predictive biomarkers of efficacy were identified by integrating genomic, reverse-phase protein microarray (RPPA) and cytogenetic analyses, and validated by immunostainings. DNA replication stress (RS) was increased by employing DNA replication-perturbing or polyploidising agents. Results The drug-library screening led to the identification of LY2606368 as a potent anti-CSC agent acting in vitro and in vivo in tumour cells from a considerable number of patients (∼36%). By inhibiting checkpoint kinase (CHK)1, LY2606368 affected DNA replication in most CRC-SCs, including RAS-mutated ones, forcing them into premature, lethal mitoses. Parallel genomic, RPPA and cytogenetic analyses indicated that CRC-SCs sensitive to LY2606368 displayed signs of ongoing RS response, including the phosphorylation of RPA32 and ataxia telangiectasia mutated serine/threonine kinase (ATM). This was associated with mutation(s) in TP53 and hyperdiploidy, and made these CRC-SCs exquisitely dependent on CHK1 function. Accordingly, experimental increase of RS sensitised resistant CRC-SCs to LY2606368. Conclusions LY2606368 selectively eliminates replication-stressed, p53-deficient and hyperdiploid CRC-SCs independently of RAS mutational status. These results provide a strong rationale for biomarker-driven clinical trials with LY2606368 in patients with CRC. PMID:28389531

LACTB, a novel epigenetic silenced tumor suppressor, inhibits colorectal cancer progression by attenuating MDM2-mediated p53 ubiquitination and degradation.

PubMed

Zeng, Kaixuan; Chen, Xiaoxiang; Hu, Xiuxiu; Liu, Xiangxiang; Xu, Tao; Sun, Huiling; Pan, Yuqin; He, Bangshun; Wang, Shukui

2018-06-13

Colorectal cancer (CRC) is one of the most common aggressive malignancies. Like other solid tumors, inactivation of tumor suppressor genes and activation of oncogenes occur during CRC development and progression. Recently, a novel tumor suppressor, LACTB, was proposed to inhibit tumor progression, but the functional and clinical significance of this tumor suppressor in CRC remains unexplored. Herein, we found LACTB was significantly downregulated in CRC due to promoter methylation and histone deacetylation, which was associated with metastasis and advanced clinical stage. CRC patients with low LACTB expression had poorer overall survival and LACTB also determined to be an independent prognostic factor for poorer outcome. Ectopic expression of LACTB suppressed CRC cells proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) in vitro and inhibited CRC growth and metastasis in vivo, while knockout of LACTB by CRISPR/Cas9 gene editing technique resulted in an opposite phenotype. Interestingly, LACTB could exert antitumorigenic effect only in HCT116 and HCT8 cells harboring wild-type TP53, but not in HT29 and SW480 cells harboring mutant TP53 or HCT116 p53 -/- cells. Mechanistic studies demonstrated that LACTB could directly bind to the C terminus of p53 to inhibit p53 degradation by preventing MDM2 from interacting with p53. Moreover, ablation of p53 attenuated the antitumorigenic effects of LACTB overexpression in CRC. Collectively, our findings successfully demonstrate for the first time that LACTB is a novel epigenetic silenced tumor suppressor through modulating the stability of p53, supporting the pursuit of LACTB as a potential therapeutic target for CRC.

High expression of WWP1 predicts poor prognosis and associates with tumor progression in human colorectal cancer

PubMed Central

Chen, Jian-Jun; Zhang, Wei

2018-01-01

WWP1 (WW domain-containing E3 ubiquitin protein ligase 1), which is frequently up-regulated in multiple human malignancies, has been demonstrated to play a critical function in cell proliferation, apoptosis and invasion. However, limited knowledge is known about the expression pattern and prognostic value of WWP1 in colorectal cancer (CRC). In this study, we firstly observed that WWP1 mRNA and protein is commonly up-regulated in CRC tissues compared with normal counterparts. Furthermore, by immunohistochemical analysis in 348 cases of CRC specimens, we demonstrated that the WWP1 protein expression is up-regulated in 58.91% (205/348) samples and detected increasing WWP1 expression is closely correlated with enhanced tumor size (P=0.022), CEA level (P=0.021), T classification (P=0.010), distant metastasis (P=0.021) and TNM stage (P=0.005). Meanwhile, Kaplan-Meier survival analysis showed CRC patients with a high WWP1 expression have a poorer overall survival (P<0.001) and disease-free survival (P=0.001) than those with a low WWP1 expression. Multivariate Cox regression analysis revealed WWP1 is the independent prognostic factors for overall survival rate of CRC patients. What’s more, by CCK-8 assays and Transwell assays, we found WWP1 depletion markedly inhibited tumor proliferation and invasion in CRC cells, and cells with WWP1 overexpression had a prominently higher proliferative and invasive capacity. Most notably, we illuminated WWP1 downregulation inactivated PTEN/Akt pathway in CRC cells. Taken together, our studies revealed the prognostic value of WWP1 in CRC and support that WWP1 may act as a molecular target for CRC treatment. PMID:29511596

Characterization of an adaptive immune response in microsatellite-instable colorectal cancer

PubMed Central

Boissière-Michot, Florence; Lazennec, Gwendal; Frugier, Hélène; Jarlier, Marta; Roca, Lise; Duffour, Jacqueline; Du Paty, Emilie; Laune, Daniel; Blanchard, France; Le Pessot, Florence; Sabourin, Jean-Christophe; Bibeau, Frédéric

2014-01-01

Sporadic or hereditary colorectal cancer (CRC) with microsatellite instability (MSI) is frequently characterized by inflammatory lymphocytic infiltration and tends to be associated with a better outcome than microsatellite stable (MSS) CRC, probably reflecting a more effective immune response. We investigated inflammatory mechanisms in 48 MSI CRCs and 62 MSS CRCs by analyzing: (1) the expression of 48 cytokines using Bio-Plex multiplex cytokine assays, and (2) the in situ immune response by immunohistochemical analysis with antibodies against CD3 (T lymphocytes), CD8 (cytotoxic T lymphocytes), CD45RO (memory T lymphocytes), T-bet (Th1 CD4 cells), and FoxP3 (regulatory T cells). MSI CRC exhibited significantly higher expression of CCL5 (RANTES), CXCL8 (IL-8), CXCL9 (MIG), IL-1β, CXCL10 (IP-10), IL-16, CXCL1 (GROα), and IL-1ra, and lower expression of MIF, compared with MSS CRC. Immunohistochemistry combined with image analysis indicated that the density of CD3+, CD8+, CD45RO+, and T-bet+ T lymphocytes was higher in MSI CRC than in MSS CRC, whereas the number of regulatory T cells (FoxP3+) was not statistically different between the groups. These results indicate that MSI CRC is associated with a specific cytokine expression profile that includes CCL5, CXCL10, and CXCL9, which are involved in the T helper type 1 (Th1) response and in the recruitment of memory CD45RO+ T cells. Our findings highlight the major role of adaptive immunity in MSI CRC and provide a possible explanation for the more favorable prognosis of this CRC subtype. PMID:25101223

PUMA mediates the anti-cancer effect of osimertinib in colon cancer cells.

PubMed

Guo, Lingchuan; Huang, Shan; Wang, Xinwei

2017-01-01

Osimertinib, an irreversible EGFR/HER2 inhibitor, has been found to be effective in the cancer cell with EGFR gene mutations in preclinical lung cancer models. However, the effect of osimertinib in colorectal cancer (CRC) cells is unclear. In the present study, we investigated how osimertinib suppresses CRC cells growth and potentiates effects of other chemotherapeutic drugs. We found that p73-mediated osimertinib-induced p53 upregulated modulator of apoptosis (PUMA) expression irrespective of p53 status following PI3K/AKT pathway inhibition in CRC cells. Furthermore, PUMA is required for osimertinib-induced apoptosis. In addition, osimertinib also synergized with 5-FU to induce significant apoptosis via PUMA in CRC cells. These results demonstrated a critical role of PUMA in mediating the anticancer effects of osimertinib and suggest that PUMA induction can be used as an indicator of osimertinib sensitivity.

PUMA mediates the anti-cancer effect of osimertinib in colon cancer cells

PubMed Central

Wang, Xinwei

2017-01-01

Osimertinib, an irreversible EGFR/HER2 inhibitor, has been found to be effective in the cancer cell with EGFR gene mutations in preclinical lung cancer models. However, the effect of osimertinib in colorectal cancer (CRC) cells is unclear. In the present study, we investigated how osimertinib suppresses CRC cells growth and potentiates effects of other chemotherapeutic drugs. We found that p73-mediated osimertinib-induced p53 upregulated modulator of apoptosis (PUMA) expression irrespective of p53 status following PI3K/AKT pathway inhibition in CRC cells. Furthermore, PUMA is required for osimertinib-induced apoptosis. In addition, osimertinib also synergized with 5-FU to induce significant apoptosis via PUMA in CRC cells. These results demonstrated a critical role of PUMA in mediating the anticancer effects of osimertinib and suggest that PUMA induction can be used as an indicator of osimertinib sensitivity. PMID:29138581

Lactate calcium salt affects the viability of colorectal cancer cells via betaine homeostasis.

PubMed

Jang, Yeong-Su; Jo, Young-Kwon; Sim, Jae Jun; Ji, Eunhee; Jeong, Keun-Yeong; Kim, Hwan Mook

2016-02-15

Betaine plays an important role in cellular homeostasis. However, the physiological roles of betaine-γ-aminobutyric acid (GABA) transporter (BGT-1) are still being disputed in cancer. In this study, we tried to find the possibility of the antitumor effect on colorectal cancer (CRC) cell via lactate calcium salt (CaLa)-induced BGT-1 downregulation. The CRC cell viability and clonogenic assay was performed using different doses of BGT-1 inhibitor. The expression level of BGT-1 was measured following the treatment of 2.5mM CaLa. Betaine was treated to confirm the resistance of the antitumor activity by CaLa. Tumor growth was also measured using a xenograft animal model. Long-term exposure of 2.5mM CaLa clearly decreased the expression of BGT-1 in the CRC cells. As a result of the downregulation of BGT-1 expression, the clonogenic ability of CRC cells was also decreased in the 2.5mM CaLa-treated group. Reversely, the number of colonies and cell viability was increased by combination treatment with betaine and 2.5mM CaLa, as compared with a single treatment of 2.5mM CaLa. Tumor growth was significantly inhibited in the xenograft model depending on BGT-1 downregulation by 2.5mM CaLa treatment. These results support the idea that long-lasting calcium supplementation via CaLa contributes to disruption of betaine homeostasis in the CRC cells and is hypothesized to reduce the risk of CRC. In addition, it indicates the possibility of CaLa being a potential incorporating agent with existing therapeutics against CRC. Copyright © 2016 Elsevier Inc. All rights reserved.

Macranthoidin B Modulates Key Metabolic Pathways to Enhance ROS Generation and Induce Cytotoxicity and Apoptosis in Colorectal Cancer.

PubMed

Fan, Xing; Rao, Jun; Zhang, Ziwei; Li, Dengfeng; Cui, Wenhao; Zhang, Jun; Wang, Hua; Tou, Fangfang; Zheng, Zhi; Shen, Qiang

2018-01-01

Induction of oxidative stress and reactive oxygen species (ROS) mediated-apoptosis have been utilized as effective strategies in anticancer therapy. Macranthoidin B (MB) is a potent inducer of ROS-mediated apoptosis in cancer, but its mechanism of action is poorly understood. Superoxide production with MB exposure in colorectal cancer (CRC) cells was measured using lucigenin chemiluminescence and real-time PCR. MB's inhibitory effect on proliferation and viability of CRC cells was determined by proliferation assays. MB's effect on apoptosis of CRC cells was determined by Western blotting and annexin V-FITC/PI staining. MB's effect on the growth of CRC xenografts in mice was assessed. An established metabolomics profiling platform combining ultra-performance liquid chromatography-tandem mass spectrometry (LC-MS) with gas chromatography-mass spectrometry (GC-MS) was performed to determine MB's effect on total metabolite variation in CRC cells. We found that MB increases ROS generation via modulating key metabolic pathways. Using metabolomics profiling platform combining LC-MS with GC-MS, a total of 236 metabolites were identified in HCT-116 cells in which 31 metabolites were determined to be significantly regulated (p ≤ 0.05) after MB exposure. A number of key metabolites revealed by metabolomics analysis include glucose, fructose, citrate, arginine, phenylalanine, and S-adenosylhomocysteine (SAH), suggesting specific modulation of metabolism on carbohydrates, amino acids and peptides, lipids, nucleotide, cofactors and vitamins in HCT-116 CRC cells with MB treatment highly associated with apoptosis triggered by enhanced ROS and activated caspase-3. Our results demonstrate that MB represses CRC cell proliferation by inducing ROS-mediated apoptosis. © 2018 The Author(s). Published by S. Karger AG, Basel.

miR-598 inhibits metastasis in colorectal cancer by suppressing JAG1/Notch2 pathway stimulating EMT.

PubMed

Chen, Jia; Zhang, Haichen; Chen, Ying; Qiao, Guanglei; Jiang, Weihua; Ni, Peihua; Liu, Xiangfan; Ma, Lijun

2017-03-01

MicroRNAs (miRNAs) are a class of endogenous, evolutionarily conserved small non-coding RNA molecules that mediate the posttranscriptional process of target gene, leading to translational repression or degradation of target mRNAs. A series of studies have indicated that miRNAs play an important role in tumor initiation, development and progression. In this study, we found that down regulation of miR-598 was a frequent event in CRC tissues compared to the paracarcinoma tissues. And the study demonstrated that miR-598 was implicated in CRC metastasis. Transwell migration assay revealed that elevated miR-598 expression reduces CRC cell migration. Moreover, our study showed that suppression of miR-598 expression induces CRC cell epithelialmesenchymal transition(EMT) and overexpression of miR-598 inhibits CRC cell EMT. In addition, bioinformatics target prediction identified JAG1 as a putative target of miR-598. Knockdown of miR-598 was shown to upregulate JAG1 expression. Furthermore, overexpression of miR-598 suppressed the expression of JAG1. Consistent results were also obtained when the regulation of JAG1 expression by miR-598 was further specified in CRC tissues. Moreover, overexpression of JAG1 induces epithelialmesenchymal transition(EMT) and promotes the metastasis of CRC cells. Decreased Notch2 expression suppresses CRC cells metastasis and EMT. Together, these results indicate that miR-598 is a novel regulator of colorectal cancer metastasis. Our data suggest miR-598 is implicated in regulating Epithelial-mesenchymal transitions by directly suppressing its downstream target gene JAG1 to inactivate Notch signaling pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

Rice Varietal Differences in Bioactive Bran Components for Inhibition of Colorectal Cancer Cell Growth

PubMed Central

Forster, Genevieve M.; Raina, Komal; Kumar, Ajay; Kumar, Sushil; Agarwal, Rajesh; Chen, Ming-Hsuan; Bauer, John E.; McClung, Anna M.; Ryan, Elizabeth P.

2013-01-01

Rice bran chemical profiles differ across rice varieties and have not yet been analyzed for differential chemopreventive bioactivity. A diverse panel of 7 rice bran varieties was analyzed for growth inhibition of human colorectal cancer (CRC) cells. Inhibition varied from 0–99%, depending on the variety of bran used. Across varieties, total lipid content ranged 5–16%, individual fatty acids had 1.4 to 1.9 fold differences, vitamin E isoforms (α-, γ-, δ- tocotrienols and tocopherols) showed 1.3 to 15.2 fold differences, and differences in γ- oryzanol and total phenolics ranged between 100–275 ng/mg and 57–146 ng GAE/mg, respectively. Spearman correlation analysis was used to identify bioactive compounds implicated in CRC cell growth inhibitory activity. Total phenolics and γ- tocotrienol were positively correlated with reduced CRC cell growth (p < 0.05). Stoichiometric variation in rice bran components and differential effects on CRC viability merit further evaluation elucidate their role in dietary CRC chemoprevention. PMID:23790950

Rice varietal differences in bioactive bran components for inhibition of colorectal cancer cell growth.

PubMed

Forster, Genevieve M; Raina, Komal; Kumar, Ajay; Kumar, Sushil; Agarwal, Rajesh; Chen, Ming-Hsuan; Bauer, John E; McClung, Anna M; Ryan, Elizabeth P

2013-11-15

Rice bran chemical profiles differ across rice varieties and have not yet been analysed for differential chemopreventive bioactivity. A diverse panel of seven rice bran varieties was analysed for growth inhibition of human colorectal cancer (CRC) cells. Inhibition varied from 0% to 99%, depending on the variety of bran used. Across varieties, total lipid content ranged 5-16%, individual fatty acids had 1.4- to 1.9-fold differences, vitamin E isoforms (α-, γ-, δ-tocotrienols, and tocopherols) showed 1.3- to 15.2-fold differences, and differences in γ-oryzanol and total phenolics ranged between 100-275ng/mg and 57-146ngGAE/mg, respectively. Spearman correlation analysis was used to identify bioactive compounds implicated in CRC cell growth inhibitory activity. Total phenolics and γ-tocotrienol were positively correlated with reduced CRC cell growth (p<0.05). Stoichiometric variation in rice bran components and differential effects on CRC viability merit further evaluation elucidate their role in dietary CRC chemoprevention. Copyright © 2013 Elsevier Ltd. All rights reserved.

Chrysin Attenuates Cell Viability of Human Colorectal Cancer Cells through Autophagy Induction Unlike 5-Fluorouracil/Oxaliplatin.

PubMed

Lin, Yueh-Ming; Chen, Chih-I; Hsiang, Yi-Ping; Hsu, Yung-Chia; Cheng, Kung-Chuan; Chien, Pei-Hsuan; Pan, Hsiao-Lin; Lu, Chien-Chang; Chen, Yun-Ju

2018-06-14

Chemotherapeutic 5-fluorouracil (5-FU) combined with oxaliplatin is often used as the standard treatment for colorectal cancer (CRC). The disturbing side effects and drug resistance commonly observed in chemotherapy motivate us to develop alternative optimal therapeutic options for CRC treatment. Chrysin, a natural and biologically active flavonoid abundant in propolis, is reported to have antitumor effects on a few CRCs. However, whether and how chrysin achieves similar effectiveness to the 5-FU combination is not clear. In this study, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), western blotting, fluorescence microscopy, and reactive oxygen species (ROS) production were assayed. We found that chrysin exhibited similar inhibition of cell viability as the 5-FU combination in a panel of human CRC cells. Furthermore, the results showed that chrysin significantly increased the levels of LC3-II, an autophagy-related marker, in CRC cells, which was not observed with the 5-FU combination. More importantly, blockage of autophagy induction restored chrysin-attenuated CRC cell viability. Further mechanistic analysis revealed that chrysin, not the 5-FU combination, induced ROS generation, and in turn, inhibited the phosphorylation of protein kinase B (Akt) and mammalian target of rapamycin (mTOR). Collectively, these results imply that chrysin may be a potential replacement for the 5-FU and oxaliplatin combination to achieve antitumor activity through autophagy for CRC treatment in the future.

Lipoic acid induces p53-independent cell death in colorectal cancer cells and potentiates the cytotoxicity of 5-fluorouracil.

PubMed

Dörsam, Bastian; Göder, Anja; Seiwert, Nina; Kaina, Bernd; Fahrer, Jörg

2015-10-01

Alpha-lipoic acid (LA), which plays a pivotal role in mitochondrial energy metabolism, is an endogenous dithiol compound with an array of antioxidative functions. It has been shown that LA triggers cell death in tumor cell lines, whereas non-transformed cells are hardly affected. In the present study, we analyzed the cytotoxicity of LA on colorectal cancer (CRC) cells differing in their p53 status and investigated a putative synergistic effect with the anticancer drug 5-fluorouracil (5-FU). We show that LA induces a dose-dependent decrease in cell viability, which was independent of the p53 status as attested in isogenic p53-proficient and p53-deficient cell lines. This effect was largely attributable to cell death induction as revealed by Annexin-V/PI staining. LA-treated HCT116 cells underwent caspase-dependent and caspase-independent cell death, which was blocked by the pan-caspase inhibitor zVAD and the RIP-kinase inhibitor Necrostatin-1, respectively. In CaCO-2 and HT29 cells, LA induced caspase-dependent cell demise via activation of caspase-9, caspase-3 and caspase-7 with subsequent PARP-1 cleavage as demonstrated by immunoblot analysis, activity assays and pan-caspase inhibition. Interestingly, LA treatment did neither activate p53 nor induced genotoxic effects as shown by lack of DNA strand breaks and phosphorylation of histone 2AX. Finally, we provide evidence that LA increases the cytotoxic effect induced by the anticancer drug 5-FU as revealed by significantly enhanced cell death rates in HCT116 and CaCO-2 cells. Collectively, these findings demonstrate that LA induces CRC cell death independent of their p53 status and potentiates the cytotoxicity of 5-FU without causing DNA damage on its own, which makes it a candidate for tumor therapy.

Cost-Effectiveness of Cetuximab as First-line Treatment for Metastatic Colorectal Cancer in the United States.

PubMed

Shankaran, Veena; Ortendahl, Jesse D; Purdum, Anna G; Bolinder, Bjorn; Anene, Ayanna M; Sun, Gordon H; Bentley, Tanya G K

2018-01-01

We conducted a cost-effectiveness analysis incorporating recent phase III clinical trial (FIRE-3) data to evaluate clinical and economic tradeoffs associated with first-line treatments of KRAS wild-type (WT) metastatic colorectal cancer (mCRC). A cost-effectiveness model was developed using FIRE-3 data to project survival and lifetime costs of FOLFIRI plus either cetuximab or bevacizumab. Hypothetical KRAS-WT mCRC patients initiated first-line treatment and could experience adverse events, disease progression warranting second-line treatment, or clinical response and hepatic metastasectomy. Model inputs were derived from FIRE-3 and published literature. Incremental cost-effectiveness ratios (ICERs) were reported as US$ per life year (LY) and quality-adjusted life year (QALY). Scenario analyses considered patients with extended RAS mutations and CALGB/SWOG 80405 data; 1-way and probabilistic sensitivity analyses were conducted. Compared with bevacizumab, KRAS-WT patients receiving first-line cetuximab gained 5.7 months of life at a cost of $46,266, for an ICER of $97,223/LY ($122,610/QALY). For extended RAS-WT patients, the ICER was $77,339/LY ($99,584/QALY). Cetuximab treatment was cost-effective 80.3% of the time, given a willingness-to-pay threshold of $150,000/LY. Results were sensitive to changes in survival, treatment duration, and product costs. Our analysis of FIRE-3 data suggests that first-line treatment with cetuximab and FOLFIRI in KRAS (and extended RAS) WT mCRC patients may improve health outcomes and use financial resources more efficiently than bevacizumab and FOLFIRI. This information, in combination with other studies investigating comparative effectiveness of first-line options, can be useful to clinicians, payers, and policymakers in making treatment and resource allocation decisions for mCRC patients.

G9a stimulates CRC growth by inducing p53 Lys373 dimethylation-dependent activation of Plk1.

PubMed

Zhang, Jie; Wang, Yafang; Shen, Yanyan; He, Pengxing; Ding, Jian; Chen, Yi

2018-01-01

Rationale: G9a is genetically deregulated in various tumor types and is important for cell proliferation; however, the mechanism underlying G9a-induced carcinogenesis, especially in colorectal cancer (CRC), is unclear. Here, we investigated if G9a exerts oncogenic effects in CRC by increasing polo-like kinase 1 (Plk1) expression. Thus, we further characterized the detailed molecular mechanisms. Methods: The role of Plk1 in G9a aberrant CRC was determined by performing different in vitro and in vivo assays, including assessment of cell growth by performing cell viability assay and assessment of signaling transduction profiles by performing immunoblotting, in the cases of pharmacological inhibition or short RNA interference-mediated suppression of G9a. Detailed molecular mechanisms underlying the effect of G9a on Plk1 expression were determined by performing point mutation analysis, chromatin immunoprecipitation analysis, and luciferase reporter assay. Correlation between G9a and Plk1 expression was determined by analyzing clinical samples of patients with CRC by performing immunohistochemistry. Results: Our study is the first to report a significant positive correlation between G9a and Plk1 levels in 89 clinical samples of patients with CRC. Moreover, G9a depletion decreased Plk1 expression and suppressed CRC cell growth both in vitro and in vivo , thus confirming the significant correlation between G9a and Plk1 levels. Further, we observed that G9a-induced Plk1 regulation depended on p53 inhibition. G9a dimethylated p53 at lysine 373, which in turn increased Plk1 expression and promoted CRC cell growth. Conclusions: These results indicate that G9a-induced and p53-dependent epigenetic programing stimulates the growth of colon cancer, which also suggests that G9a inhibitors that restore p53 activity are promising therapeutic agents for treating colon cancer, especially for CRC expressing wild-type p53.

Effect of β,β-dimethylacrylshikonin on inhibition of human colorectal cancer cell growth in vitro and in vivo.

PubMed

Fan, Yingying; Jin, Shaoju; He, Jun; Shao, Zhenjun; Yan, Jiao; Feng, Ting; Li, Hong

2012-01-01

In traditional Chinese medicine, shikonin and its derivatives, has been used in East Asia for several years for the prevention and treatment of several diseases, including cancer. We previously identified that β,β-dimethylacrylshikonin (DA) could inhibit hepatocellular carcinoma growth. In the present study, we investigated the inhibitory effects of DA on human colorectal cancer (CRC) cell line HCT-116 in vitro and in vivo. A viability assay showed that DA could inhibit tumor cell growth in a time- and dose-dependent manner. Flow cytometry showed that DA blocks the cell cycle at G(0)/G(1) phase. Western blotting results demonstrated that the induction of apoptosis by DA correlated with the induction of pro-apoptotic proteins Bax, and Bid, and a decrease in the expression of anti-apoptotic proteins Bcl-2 and Bcl-xl. Furthermore, treatment of HCT-116 bearing nude mice with DA significantly retarded the growth of xenografts. Consistent with the results in vitro, the DA-mediated suppression of HCT-116 xenografts correlated with Bax and Bcl-2. Taken together, these results suggest that DA could be a novel and promising approach to the treatment of CRC.

LPA Induces Colon Cancer Cell Proliferation through a Cooperation between the ROCK and STAT-3 Pathways.

PubMed

Leve, Fernanda; Peres-Moreira, Rubem J; Binato, Renata; Abdelhay, Eliana; Morgado-Díaz, José A

2015-01-01

Lysophosphatidic acid (LPA) plays a critical role in the proliferation and migration of colon cancer cells; however, the downstream signaling events underlying these processes remain poorly characterized. The aim of this study was to investigate the signaling pathways triggered by LPA to regulate the mechanisms involved in the progression of colorectal cancer (CRC). We have used three cell line models of CRC, and initially analyzed the expression profile of LPA receptors (LPAR). Then, we treated the cells with LPA and events related to their tumorigenic potential, such as migration, invasion, anchorage-independent growth, proliferation as well as apoptosis and cell cycle were evaluated. We used the Chip array technique to analyze the global gene expression profiling that occurs after LPA treatment, and we identified cell signaling pathways related to the cell cycle. The inhibition of these pathways verified the conclusions of the transcriptomic analysis. We found that the cell lines expressed LPAR1, -2 and -3 in a differential manner and that 10 μM LPA did not affect cell migration, invasion and anchorage-independent growth, but it did induce proliferation and cell cycle progression in HCT-116 cells. Although LPA in this concentration did not induce transcriptional activity of β-catenin, it promoted the activation of Rho and STAT-3. Moreover, ROCK and STAT-3 inhibitors prevented LPA-induced proliferation, but ROCK inhibition did not prevent STAT-3 activation. Finally, we observed that LPA regulates the expression of genes related to the cell cycle and that the combined inhibition of ROCK and STAT-3 prevented cell cycle progression and increased the LPA-induced expression of cyclins E1, A2 and B1 to a greater degree than either inhibitor alone. Overall, these results demonstrate that LPA increases the proliferative potential of colon adenocarcinoma HCT-116 cells through a mechanism involving cooperation between the Rho-ROCK and STAT3 pathways involved in cell cycle control.

MicroRNA-20a-5p promotes colorectal cancer invasion and metastasis by downregulating Smad4.

PubMed

Cheng, Dantong; Zhao, Senlin; Tang, Huamei; Zhang, Dongyuan; Sun, Hongcheng; Yu, Fudong; Jiang, Weiliang; Yue, Ben; Wang, Jingtao; Zhang, Meng; Yu, Yang; Liu, Xisheng; Sun, Xiaofeng; Zhou, Zongguang; Qin, Xuebin; Zhang, Xin; Yan, Dongwang; Wen, Yugang; Peng, Zhihai

2016-07-19

Tumor metastasis is one of the leading causes of poor prognosis for colorectal cancer (CRC) patients. Loss of Smad4 contributes to aggression process in many human cancers. However, the underlying precise mechanism of aberrant Smad4 expression in CRC development is still little known. miR-20a-5p negatively regulated Smad4 by directly targeting its 3'UTR in human colorectal cancer cells. miR-20a-5p not only promoted CRC cells aggression capacity in vitro and liver metastasis in vivo, but also promoted the epithelial-to-mesenchymal transition process by downregulating Smad4 expression. In addition, tissue microarray analysis obtained from 544 CRC patients' clinical characters showed that miR-20a-5p was upregulated in human CRC tissues, especially in the tissues with metastasis. High level of miR-20a-5p predicted poor prognosis in CRC patients. Five miRNA target prediction programs were applied to identify potential miRNA(s) that target(s) Smad4 in CRC. Luciferase reporter assay and transfection technique were used to validate the correlation between miR-20a-5p and Smad4 in CRC. Wound healing, transwell and tumorigenesis assays were used to explore the function of miR-20a-5p and Smad4 in CRC progression in vitro and in vivo. The association between miR-20a-5p expression and the prognosis of CRC patients was evaluated by Kaplan-Meier analysis and multivariate cox proportional hazard analyses based on tissue microarray data. miR-20a-5p, as an onco-miRNA, promoted the invasion and metastasis ability by suppressing Smad4 expression in CRC cells, and high miR-20a-5p predicted poor prognosis for CRC patients, providing a novel and promising therapeutic target in human colorectal cancer.

MicroRNA-20a-5p promotes colorectal cancer invasion and metastasis by downregulating Smad4

PubMed Central

Zhang, Dongyuan; Sun, Hongcheng; Yu, Fudong; Yue, Ben; Wang, Jingtao; Zhang, Meng; Yu, Yang; Liu, Xisheng; Sun, Xiaofeng; Zhou, Zongguang; Qin, Xuebin; Zhang, Xin; Yan, Dongwang; Wen, Yugang; Peng, Zhihai

2016-01-01

Background Tumor metastasis is one of the leading causes of poor prognosis for colorectal cancer (CRC) patients. Loss of Smad4 contributes to aggression process in many human cancers. However, the underlying precise mechanism of aberrant Smad4 expression in CRC development is still little known. Results miR-20a-5p negatively regulated Smad4 by directly targeting its 3′UTR in human colorectal cancer cells. miR-20a-5p not only promoted CRC cells aggression capacity in vitro and liver metastasis in vivo, but also promoted the epithelial-to-mesenchymal transition process by downregulating Smad4 expression. In addition, tissue microarray analysis obtained from 544 CRC patients’ clinical characters showed that miR-20a-5p was upregulated in human CRC tissues, especially in the tissues with metastasis. High level of miR-20a-5p predicted poor prognosis in CRC patients. Methods Five miRNA target prediction programs were applied to identify potential miRNA(s) that target(s) Smad4 in CRC. Luciferase reporter assay and transfection technique were used to validate the correlation between miR-20a-5p and Smad4 in CRC. Wound healing, transwell and tumorigenesis assays were used to explore the function of miR-20a-5p and Smad4 in CRC progression in vitro and in vivo. The association between miR-20a-5p expression and the prognosis of CRC patients was evaluated by Kaplan–Meier analysis and multivariate cox proportional hazard analyses based on tissue microarray data. Conclusions miR-20a-5p, as an onco-miRNA, promoted the invasion and metastasis ability by suppressing Smad4 expression in CRC cells, and high miR-20a-5p predicted poor prognosis for CRC patients, providing a novel and promising therapeutic target in human colorectal cancer. PMID:27286257

TS and ERCC-1 mRNA expressions and clinical outcome in patients with metastatic colon cancer in CONFIRM-1 and -2 clinical trials.

PubMed

Grimminger, P P; Shi, M; Barrett, C; Lebwohl, D; Danenberg, K D; Brabender, J; Vigen, C L P; Danenberg, P V; Winder, T; Lenz, H-J

2012-10-01

To validate established cutoff levels of thymidylate synthase (TS) and excision repair cross-complementing (ERCC-1) intratumoral mRNA expressions in tumor samples from metastatic colorectal cancer (mCRC) patients treated with PTK787/ZK222584 (PTK/ZK). From 122 samples of patients with mCRC enrolled in CONFIRM-1 (Colorectal Oral Novel Therapy for the Inhibition of Angiogenesis and Retarding of Metastases) or CONFIRM-2, mRNA was isolated of microdissected formalin-fixed paraffin-embedded samples and quantitated using TaqMan-based technology. Existing TS and ERCC-1 cutoff levels were tested for their prognostic value in first-line and second-line therapy. TS expression was associated with overall survival (OS) in first-line, but not second-line therapy. ERCC-1 was associated with OS in patients treated with first-line and second-line FOLFOX4. In first-line FOLFOX4, combination of high TS and/or high ERCC-1 was associated with shorter OS. A correlation was observed between ERCC-1 expression and benefit from PTK/ZK+FOLFOX4 treatment. TS and ERCC-1 expression is associated with clinical outcome in mCRC. Baseline TS and ERCC-1 levels may allow the selection of patients who benefit from FOLFOX4 chemotherapy.

A complex of α6 integrin and E-cadherin drives liver metastasis of colorectal cancer cells through hepatic angiopoietin-like 6.

PubMed

Marchiò, Serena; Soster, Marco; Cardaci, Sabrina; Muratore, Andrea; Bartolini, Alice; Barone, Vanessa; Ribero, Dario; Monti, Maria; Bovino, Paola; Sun, Jessica; Giavazzi, Raffaella; Asioli, Sofia; Cassoni, Paola; Capussotti, Lorenzo; Pucci, Piero; Bugatti, Antonella; Rusnati, Marco; Pasqualini, Renata; Arap, Wadih; Bussolino, Federico

2012-11-01

Homing of colorectal cancer (CRC) cells to the liver is a non-random process driven by a crosstalk between tumour cells and components of the host tissue. Here we report the isolation of a liver metastasis-specific peptide ligand (CGIYRLRSC) that binds a complex of E-cadherin and α(6) integrin on the surface of CRC cells. We identify angiopoietin-like 6 protein as a peptide-mimicked natural ligand enriched in hepatic blood vessels of CRC patients. We demonstrate that an interaction between hepatic angiopoietin-like 6 and tumoural α(6) integrin/E-cadherin drives liver homing and colonization by CRC cells, and that CGIYRLRSC inhibits liver metastasis through interference with this ligand/receptor system. Our results indicate a mechanism for metastasis whereby a soluble factor accumulated in normal vessels functions as a specific ligand for circulating cancer cells. Consistently, we show that high amounts of coexpressed α(6) integrin and E-cadherin in primary tumours represent a poor prognostic factor for patients with advanced CRC. Copyright © 2012 The Authors. Published by John Wiley and Sons, Ltd on behalf of EMBO.

Predictive Biomarkers of Radiation Sensitivity in Rectal Cancer

NASA Astrophysics Data System (ADS)

Tut, Thein Ga

Colorectal cancer (CRC) is the third most common cancer in the world. Australia, New Zealand, Canada, the United States, and parts of Europe have the highest incidence rates of CRC. China, India, South America and parts of Africa have the lowest risk of CRC. CRC is the second most common cancer in both sexes in Australia. Even though the death rates from CRC involving the colon have diminished, those arising from the rectum have revealed no improvement. The greatest obstacle in attaining a complete surgical resection of large rectal cancers is the close anatomical relation to surrounding structures, as opposed to the free serosal surfaces enfolding the colon. To assist complete resection, pre-operative radiotherapy (DXT) can be applied, but the efficacy of ionising radiation (IR) is extremely variable between individual tumours. Reliable predictive marker/s that enable patient stratification in the application of this otherwise toxic therapy is still not available. Current therapeutic management of rectal cancer can be improved with the availability of better predictive and prognostic biomarkers. Proteins such as Plk1, gammaH2AX and MMR proteins (MSH2, MSH6, MLH1 and PMS2), involved in DNA damage response (DDR) pathway may be possible biomarkers for radiation response prediction and prognostication of rectal cancer. Serine/threonine protein kinase Plk1 is overexpressed in most of cancers including CRC. Plk1 functional activity is essential in the restoration of DNA damage following IR, which causes DNA double strand break (DSB). The earliest manifestation of this reparative process is histone H2AX phosphorylation at serine 139, leading to gammaH2AX. Colorectal normal mucosa showed the lowest level of gammaH2AX with gradually increasing levels in early adenoma and then in advanced malignant colorectal tissues, leading to the possibility that gammaH2AX may be a prospective biomarker in rectal cancer management. There are numerous publications regarding DNA mismatch repair (MMR) proteins, the insufficiency of which is characteristic of CRCs with microsatellite instability (MSI). MSI may enable unlimited replicative potential of malignant cell that leads to radiation injury resistance. Therefore, these proteins were characterized in both CRC cell lines (MMR proteins) and different (core and invasive front) rectal cancer tissues (Plk1, gammaH2AX and MMR proteins) exposed to radiation. Histopathological grading of tumour regression was performed following radiotherapy in rectal cancer as a marker of radiotherapy response and a surrogate indicator of patient prognosis. Though MMR protein expressions correlated with improved in vitro cell survival following radiation, these findings could only be partially replicated in patient tissue samples. This may not be entirely unexpected, given intratumoural heterogeneity in genetic profiles and oxygenation between individual cancer cells, their interaction with stromal environment and a multitude of other factors that cannot be adequately replicated in cell line experiments. In our rectal cancer patient cohort, histopathological regression following radiotherapy did appear to correlate with better clinical outcome, but certainly no replacement for the routine pTNM staging with which it was compared. Overexpression of Plk1 in the primary rectal cancer also correlates with poor tumour regression and reduced overall survival. High level of gammaH2AX correlates with higher tumour stage, perineural invasion and vascular invasion. However, interpretation of the results is limited by the small number of positivity amongst the cohort, with respect to gammaH2AX and MMR proteins. The combined analysis of all the proteins examined in this thesis revealed no interactions, possibly suggesting these biomarkers act individually within the DDR pathway, rather than in a demonstrably interdependent manner. Though our results are mixed, finding biomarkers predictive of radiation response is nonetheless critical. Enhancing the radiosensitivity of cancers through manipulating the functional activity and/or expression of prospective biomarkers could conceivably enhance tumour response to the level that the extent of consequent surgical resection can be minimized.

Interplay between Trx-1 and S100P promotes colorectal cancer cell epithelial-mesenchymal transition by up-regulating S100A4 through AKT activation.

PubMed

Zuo, Zhigui; Zhang, Peili; Lin, Feiyan; Shang, Wenjing; Bi, Ruichun; Lu, Fengying; Wu, Jianbo; Jiang, Lei

2018-04-01

We previously reported a novel positive feedback loop between thioredoxin-1 (Trx-1) and S100P, which promotes the invasion and metastasis of colorectal cancer (CRC). However, the underlying molecular mechanisms remain poorly understood. In this study, we examined the roles of Trx-1 and S100P in CRC epithelial-to-mesenchymal transition (EMT) and their underlying mechanisms. We observed that knockdown of Trx-1 or S100P in SW620 cells inhibited EMT, whereas overexpression of Trx-1 or S100P in SW480 cells promoted EMT. Importantly, S100A4 and the phosphorylation of AKT were identified as potential downstream targets of Trx-1 and S100P in CRC cells. Silencing S100A4 or inhibition of AKT phosphorylation eliminated S100P- or Trx-1-mediated CRC cell EMT, migration and invasion. Moreover, inhibition of AKT activity reversed S100P- or Trx-1-induced S100A4 expression. The expression of S100A4 was higher in human CRC tissues compared with their normal counterpart tissues and was significantly correlated with lymph node metastasis and poor survival. The overexpression of S100A4 protein was also positively correlated with S100P or Trx-1 protein overexpression in our cohort of CRC tissues. In addition, overexpression of S100P reversed the Trx-1 knockdown-induced inhibition of S100A4 expression, EMT and migration and invasion in SW620 cells. The data suggest that interplay between Trx-1 and S100P promoted CRC EMT as well as migration and invasion by up-regulating S100A4 through AKT activation, thus providing further potential therapeutic targets for suppressing the EMT in metastatic CRC. © 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

ZEB1 Mediates Drug Resistance and EMT in p300-Deficient CRC.

PubMed

Lazarova, Darina; Bordonaro, Michael

2017-01-01

We discuss the hypothesis that ZEB1-Wnt-p300 signaling integrates epithelial to mesenchymal transition (EMT) and resistance to histone deacetylase inhibitors (HDACis) in colorectal cancer (CRC) cells. The HDACi butyrate, derived from dietary fiber, has been linked to CRC prevention, and other HDACis have been proposed as therapeutic agents against CRC. We have previously discussed that resistance to butyrate likely contributes to colonic carcinogenesis, and we have demonstrated that butyrate resistance leads to cross-resistance to cancer therapeutic HDACis. Deregulated Wnt signaling is the major initiating event in most CRC cases. One mechanism whereby butyrate and other HDACis exert their anti-CRC effects is via Wnt signaling hyperactivation, which promotes CRC cell apoptosis. The histone acetylases (HATs) CBP and p300 are mediators of Wnt transcriptional activity, and play divergent roles in the downstream consequences of Wnt signaling. CBP-mediated Wnt signaling is associated with cell proliferation and stem cell maintenance; whereas, p300-mediated Wnt activity is associated with differentiation. We have found that CBP and p300 differentially affect the ability of butyrate to influence Wnt signaling, apoptosis, and proliferation. ZEB 1 is a Wnt signaling-targeted gene, whose product is a transcription factor expressed at the invasive front of carcinomas where it promotes malignant progression and EMT. ZEB1 is typically a transcriptional repressor; however, when associated with p300, ZEB1 enhances transcription. These changes in ZEB1 activity likely affect the cancer cell phenotype. ZEB1 has been shown to promote resistance to chemotherapeutic agents, and expression of ZEB1 is upregulated in butyrate-resistant CRC cells that lack p300 expression. Since the expression of ZEB1 correlates with poor outcomes in cancer, ZEB represents a relevant therapeutic target. Here we propose that targeting the signaling network established by ZEB1, Wnt signaling, and p300 signaling can reverse HDACi resistance and inhibit EMT.

EphA2 signaling is impacted by carcinoembryonic antigen cell adhesion molecule 1-L expression in colorectal cancer liver metastasis in a cell context-dependent manner.

PubMed

Arabzadeh, Azadeh; McGregor, Kevin; Breton, Valérie; Van Der Kraak, Lauren; Akavia, Uri David; Greenwood, Celia M T; Beauchemin, Nicole

2017-11-28

We have shown that carcinoembryonic antigen cell adhesion molecule 1 long isoform (CEACAM1-L) expression in MC38 metastatic colorectal cancer (CRC) cells results in liver metastasis inhibition via CCL2 and STAT3 signaling. But other molecular mechanisms orchestrating CEACAM1-L-mediated metastasis inhibition remain to be defined. We screened a panel of mouse and human CRC cells and evaluated their metastatic outcome after CEACAM1 overexpression or downregulation. An unbiased transcript profiling and a phospho-receptor tyrosine kinase screen comparing MC38 CEACAM1-L-expressing and non-expressing (CT) CRC cells revealed reduced ephrin type-A receptor 2 (EPHA2) expression and activity. An EPHA2-specific inhibitor reduced EPHA2 downstream signaling in CT cells similar to that in CEACAM1-L cells with decreased proliferation and migration. Human CRC patients exhibiting high CEACAM1 in combination with low EPHA2 expression benefited from longer time to first recurrence/metastasis compared to those with high EPHA2 expression. With the added interaction of CEACAM6 , we denoted that CEACAM1 high- and EPHA2 low-expressing patient samples with lower CEACAM6 expression also exhibited a longer time to first recurrence/metastasis. In HT29 human CRC cells, down-regulation of CEACAM1 along with CEA and CEACAM6 up-regulation led to higher metastatic burden. Overall, CEACAM1-L expression in poorly differentiated CRC can inhibit liver metastasis through cell context-dependent EPHA2-mediated signaling. However, CEACAM1's role should be considered in the presence of other CEACAM family members.

Black Raspberries Enhance Natural Killer Cell Infiltration into the Colon and Suppress the Progression of Colorectal Cancer

PubMed Central

Pan, Pan; Kang, Siwen; Wang, Youwei; Liu, Ka; Oshima, Kiyoko; Huang, Yi-Wen; Zhang, Jianying; Yearsley, Martha; Yu, Jianhua; Wang, Li-Shu

2017-01-01

Natural killer (NK) cells are an essential component of innate immunity against cancer development. Many studies have been conducted to evaluate immune-modulating effects using dietary compounds. Our laboratory has been investigating the chemopreventive potential of black raspberries (BRBs) and previously demonstrated their beneficial modulation of genetic and epigenetic biomarkers in patients with colorectal cancer (CRC). The current study investigated their potential on modulating NK cells. To avoid the excessive inflammation caused by the dextran sulfate sodium (DSS) treatment that leads to colitis, we treated the mice with overnight DSS so that it would slightly irritate the colon but still promote colon carcinogenesis with 100% incidence in both the ApcMin/+ mice and azoxymethane (AOM)-treated mice. A significant decrease of tissue-infiltrating NK cells along the progression of microadenoma-to-adenoma and adenoma-to-adenocarcinoma was observed in the ApcMin/+/DSS and AOM/DSS mice, respectively. Depletion of NK cells significantly promoted the development of CRC, suggesting a critical role of NK cells in combating CRC progression. BRBs significantly suppressed the CRC progression and increased the number of tissue-infiltrating NK cells in both mouse models. Moreover, we further determined BRBs’ effects on NK cells in the human biopsy specimens collected from our previously completed clinical trial, in which CRC patients consumed BRBs for an average of 4 weeks during a presurgical window. We observed an increased number and an enhanced cytotoxicity of NK cells by BRB intervention. The current study provides evidence that BRBs have the potential to enhance the tumor immunesurveillance of NK cells that can be beneficial in the setting of CRC prevention and treatment. PMID:28861089

Overproduction of Three Genes Leads to Camphor Resistance and Chromosome Condensation in Escherichia Coli

PubMed Central

Hu, K. H.; Liu, E.; Dean, K.; Gingras, M.; DeGraff, W.; Trun, N. J.

1996-01-01

We isolated and characterized three genes, crcA, cspE and crcB, which when present in high copy confer camphor resistance on a cell and suppress mutations in the chromosomal partition gene mukB. Both phenotypes require the same genes. Unlike chromosomal camphor resistant mutants, high copy number crcA, cspE and crcB do not result in an increase in the ploidy of the cells. The cspE gene has been previously identified as a cold shock-like protein with homologues in all organisms tested. We also demonstrate that camphor causes the nucleoids to decondense in vivo and when the three genes are present in high copy, the chromosomes do not decondense. Our results implicate camphor and mukB mutations as interfering with chromosome condensation and high copy crcA, cspE and crcB as promoting or protecting chromosome folding. PMID:8844142

Jolkinolide B induces apoptosis of colorectal carcinoma through ROS-ER stress-Ca2+-mitochondria dependent pathway

PubMed Central

Zhang, Jing; Wang, Yang; Zhou, Ye; He, Qing-Yu

2017-01-01

Colorectal carcinoma (CRC) remains one of the leading causes of death in cancer-related diseases. In this study, we aimed to investigate the anticancer effect of Jolkinolide B (JB), a bioactive diterpenoid component isolated from the dried roots of Euphorbia fischeriana Steud, on CRC cells and its underlying mechanisms. We found that JB suppressed the cell viability and colony formation of CRC cells, HT29 and SW620. Annexin V/PI assay revealed that JB induced apoptosis in CRC cells, which was further confirmed by the increased expression of cleaved-caspase3 and cleaved-PARP. iTRAQ-based quantitative proteomics was performed to identify JB-regulated proteins in CRC cells. Gene Ontology (GO) analysis revealed that these JB-regulated proteins were mainly involved in ER stress response, which was evidenced by the expression of ER stress marker proteins, HSP90, Bip and PDI. Moreover, we found that JB provoked the generation of reactive oxygen species (ROS), and that inhibition of the ROS generation with N-acetyl L-cysteine could reverse the JB-induced apoptosis. Confocal microscopy and flow cytometry showed that JB treatment enhanced intracellular and mitochondrial Ca2+ level and JC-1 assay revealed a loss of mitochondrial membrane potential in CRC after JB treatment. The mitochondrial Ca2+ uptake and depolarization can be blocked by Ruthenium Red (RuRed), an inhibitor of mitochondrial Ca2+ uniporter. Taken together, we demonstrated that JB exerts its anticancer effect by ER stress-Ca2+-mitochondria signaling, suggesting the promising chemotherapeutic potential of JB for the treatment of CRC. PMID:29207638

Jolkinolide B induces apoptosis of colorectal carcinoma through ROS-ER stress-Ca2+-mitochondria dependent pathway.

PubMed

Zhang, Jing; Wang, Yang; Zhou, Ye; He, Qing-Yu

2017-10-31

Colorectal carcinoma (CRC) remains one of the leading causes of death in cancer-related diseases. In this study, we aimed to investigate the anticancer effect of Jolkinolide B (JB), a bioactive diterpenoid component isolated from the dried roots of Euphorbia fischeriana Steud, on CRC cells and its underlying mechanisms. We found that JB suppressed the cell viability and colony formation of CRC cells, HT29 and SW620. Annexin V/PI assay revealed that JB induced apoptosis in CRC cells, which was further confirmed by the increased expression of cleaved-caspase3 and cleaved-PARP. iTRAQ-based quantitative proteomics was performed to identify JB-regulated proteins in CRC cells. Gene Ontology (GO) analysis revealed that these JB-regulated proteins were mainly involved in ER stress response, which was evidenced by the expression of ER stress marker proteins, HSP90, Bip and PDI. Moreover, we found that JB provoked the generation of reactive oxygen species (ROS), and that inhibition of the ROS generation with N-acetyl L-cysteine could reverse the JB-induced apoptosis. Confocal microscopy and flow cytometry showed that JB treatment enhanced intracellular and mitochondrial Ca 2+ level and JC-1 assay revealed a loss of mitochondrial membrane potential in CRC after JB treatment. The mitochondrial Ca 2+ uptake and depolarization can be blocked by Ruthenium Red (RuRed), an inhibitor of mitochondrial Ca 2+ uniporter. Taken together, we demonstrated that JB exerts its anticancer effect by ER stress-Ca 2+ -mitochondria signaling, suggesting the promising chemotherapeutic potential of JB for the treatment of CRC.

Anti-tumor effects of ONC201 in combination with VEGF-inhibitors significantly impacts colorectal cancer growth and survival in vivo through complementary non-overlapping mechanisms.

PubMed

Wagner, Jessica; Kline, C Leah; Zhou, Lanlan; Khazak, Vladimir; El-Deiry, Wafik S

2018-01-22

Small molecule ONC201 is an investigational anti-tumor agent that upregulates intra-tumoral TRAIL expression and the integrated stress response pathway. A Phase I clinical trial using ONC201 therapy in advanced cancer patients has been completed and the drug has progressed into Phase II trials in several cancer types. Colorectal cancer (CRC) remains one of the leading causes of cancer worldwide and metastatic disease has a poor prognosis. Clinical trials in CRC and other tumor types have demonstrated that therapeutics targeting the vascular endothelial growth factor (VEGF) pathway, such as bevacizumab, are effective in combination with certain chemotherapeutic agents. We investigated the potential combination of VEGF inhibitors such as bevacizumab and its murine-counterpart; along with other anti-angiogenic agents and ONC201 in both CRC xenograft and patient-derived xenograft (PDX) models. We utilized non-invasive imaging and immunohistochemistry to determine potential mechanisms of action. Our results demonstrate significant tumor regression or complete tumor ablation in human xenografts with the combination of ONC201 with bevacizumab, and in syngeneic MC38 colorectal cancer xenografts using a murine VEGF-A inhibitor. Imaging demonstrated the impact of this combination on decreasing tumor growth and tumor metastasis. Our results indicate that ONC201 and anti-angiogenic agents act through distinct mechanisms while increasing tumor cell death and inhibiting proliferation. With the use of both a murine VEGF inhibitor in syngeneic models, and bevacizumab in human cell line-derived xenografts, we demonstrate that ONC201 in combination with anti-angiogenic therapies such as bevacizumab represents a promising approach for further testing in the clinic for the treatment of CRC.

Gene Therapy Using Therapeutic and Diagnostic Recombinant Oncolytic Vaccinia Virus GLV-1h153 for Management of Colorectal Peritoneal Carcinomatosis

PubMed Central

Eveno, Clarisse; Mojica, Kelly; Ady, Justin W.; Thorek, Daniel L.J.; Longo, Valerie; Belin, Laurence J.; Gholami, Sepideh; Johnsen, Clark; Zanzonico, Pat; Chen, Nanhai; Yu, Tony; Szalay, Aladar A.; Fong, Yuman

2015-01-01

Background Peritoneal carcinomatosis (PC) is a terminal progression of colorectal cancer (CRC). Poor response to cytoreductive surgery and chemotherapy, coupled with the inability to reliably track disease progression using established diagnostic methods make this a deadly disease. This paper examines the effectiveness of the oncolytic vaccinia virus GLV-1h153 as a therapeutic and diagnostic vehicle. We believe that viral expression of the human sodium iodide transporter (hNIS) can provide both real-time monitoring of viral therapy and effective treatment of colorectal peritoneal carcinomatosis (CRPC). Methods Infectivity and cytotoxic effect of GLV-1h153 on CRC cell lines was assayed in-vitro. Viral replication was examined by standard viral plaque assays. Orthotopic CRPC xenografts were generated in athymic nude mice, and subsequently administered GLV-1h153 intraperitoneally. Reduction of tumor burden was assessed by mass. Orthotopic tumors were visualized by SPECT/CT after Iodine (131I) administration and by fluorescence optical imaging. Results GLV-1h153 infected and killed CRC cells in a time and concentration dependent manner. Viral replication demonstrated greater than a 2.35 log increase in titer over 4 days. Intraperitoneal treatment of orthotopic CRPC xenografts resulted in a significant reduction of tumor burden. Infection of orthotopic xenografts was both therapeutic and facilitated monitoring by 131I-SPECT/CT via expression of hNIS in infected tissue. Conclusions GLV-1h153 effectively kills CRC in-vitro and dramatically reduces tumor burden in-vivo. We demonstrate that GLV-1h153 can be used as an agent to provide accurate delineation of tumor burden in-vivo. These findings indicate that GLV-1h153 has significant potential for use as theragnostic agent in the treatment of CRPC. PMID:25616946

Expression of metastasis suppressor gene AES driven by a Yin Yang (YY) element in a CpG island promoter and transcription factor YY2.

PubMed

Kakizaki, Fumihiko; Sonoshita, Masahiro; Miyoshi, Hiroyuki; Itatani, Yoshiro; Ito, Shinji; Kawada, Kenji; Sakai, Yoshiharu; Taketo, M Mark

2016-11-01

We recently found that the product of the AES gene functions as a metastasis suppressor of colorectal cancer (CRC) in both humans and mice. Expression of amino-terminal enhancer of split (AES) protein is significantly decreased in liver metastatic lesions compared with primary colon tumors. To investigate its downregulation mechanism in metastases, we searched for transcriptional regulators of AES in human CRC and found that its expression is reduced mainly by transcriptional dysregulation and, in some cases, by additional haploidization of its coding gene. The AES promoter-enhancer is in a typical CpG island, and contains a Yin-Yang transcription factor recognition sequence (YY element). In human epithelial cells of normal colon and primary tumors, transcription factor YY2, a member of the YY family, binds directly to the YY element, and stimulates expression of AES. In a transplantation mouse model of liver metastases, however, expression of Yy2 (and therefore of Aes) is downregulated. In human CRC metastases to the liver, the levels of AES protein are correlated with those of YY2. In addition, we noticed copy-number reduction for the AES coding gene in chromosome 19p13.3 in 12% (5/42) of human CRC cell lines. We excluded other mechanisms such as point or indel mutations in the coding or regulatory regions of the AES gene, CpG methylation in the AES promoter enhancer, expression of microRNAs, and chromatin histone modifications. These results indicate that Aes may belong to a novel family of metastasis suppressors with a CpG-island promoter enhancer, and it is regulated transcriptionally. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Down-regulation of MutS homolog 3 by hypoxia in human colorectal cancer

PubMed Central

Li, Jie; Koike, Junichi; Kugoh, Hiroyuki; Arita, Michitsune; Ohhira, Takahito; Kikuchi, Yoshinori; Funahashi, Kimihiko; Takamatsu, Ken; Boland, C. Richard; Koi, Minoru; Hemmi, Hiromichi

2013-01-01

Down-regulation of hMSH3 is associated with elevated microsatellite alterations at selected tetranucleotide repeats and low levels of microsatellite instability in colorectal cancer (CRC). However, the mechanism that down-regulates hMSH3 in CRC is not known. In this study, a significant association between over-expression of glucose transporter 1, a marker for hypoxia, and down-regulation of hMSH3 in CRC tissues was observed. Therefore, we examined the effect of hypoxia on the expression of hMSH3 in human cell lines. When cells with wild type p53 (wt-p53) were exposed to hypoxia, rapid down-regulation of both hMSH2 and hMSH3 occurred. In contrast, when null or mutated p53 (null/mut-p53) cells were exposed to hypoxia, only hMSH3 was down-regulated, and at slower rate than wt-p53 cells. Using a reporter assay, we found that disruption of the two putative hypoxia response elements (HREs) located within the promoter region of the hMSH3 abrogated the suppressive effect of hypoxia on reporter activity regardless of p53 status. In an EMSA, two different forms of HIF-1α complexes that specifically bind to these HREs were detected. A larger complex containing HIF-1α predominantly bound to the HREs in hypoxic null/mut-p53 cells whereas a smaller complex predominated in wt-p53 cells. Finally, HIF-1α knockdown by siRNA significantly inhibited down-regulation of hMSH3 by hypoxia in both wt-p53 and mut-p53 cells. Taken together, our results suggest that the binding of HIF-1α complexes to HRE sites is necessary for down-regulation of hMSH3 in both wt-p53 and mut-p53 cells. PMID:22343000

Targeting the Epidermal Growth Factor Receptor Can Counteract the Inhibition of Natural Killer Cell Function Exerted by Colorectal Tumor-Associated Fibroblasts

PubMed Central

Costa, Delfina; Venè, Roberta; Benelli, Roberto; Romairone, Emanuele; Scabini, Stefano; Catellani, Silvia; Rebesco, Barbara; Mastracci, Luca; Grillo, Federica; Minghelli, Simona; Loiacono, Fabrizio; Zocchi, Maria Raffaella; Poggi, Alessandro

2018-01-01

Mesenchymal stromal cells (MSC) present in the tumor microenvironment [usually named tumor-associated fibroblasts (TAF)] can exert immunosuppressive effects on T and natural killer (NK) lymphocytes, favoring tumor immune escape. We have analyzed this mechanism in colorectal carcinoma (CRC) and found that co-culture of NK cells with TAF can prevent the IL-2-mediated NKG2D upregulation. This leads to the impairment of NKG2D-mediated recognition of CRC cells, sparing the NK cell activation through DNAM1 or FcγRIIIA (CD16). In situ, TAF express detectable levels of epidermal growth factor receptor (EGFR); thus, the therapeutic anti-EGFR humanized antibody cetuximab can trigger the antibody-dependent cellular cytotoxicity of TAF, through the engagement of FcγRIIIA on NK cells. Importantly, in the tumor, we found a lymphoid infiltrate containing NKp46+CD3− NK cells, enriched in CD16+ cells. This population, sorted and cultured with IL-2, could be triggered via CD16 and via NKG2D. Of note, ex vivo NKp46+CD3− cells were able to kill autologous TAF; in vivo, this might represent a control mechanism to reduce TAF-mediated regulatory effect on NK cell function. Altogether, these findings suggest that MSC from the neoplastic mucosa (TAF) of CRC patients can downregulate the immune cell recognition of CRC tumor cells. This immunosuppression can be relieved by the anti-EGFR antibody used in CRC immunotherapy. PMID:29910806

Genome-wide mRNA and miRNA expression profiling reveal multiple regulatory networks in colorectal cancer

PubMed Central

Vishnubalaji, R; Hamam, R; Abdulla, M-H; Mohammed, M A V; Kassem, M; Al-Obeed, O; Aldahmash, A; Alajez, N M

2015-01-01

Despite recent advances in cancer management, colorectal cancer (CRC) remains the third most common cancer and a major health-care problem worldwide. MicroRNAs have recently emerged as key regulators of cancer development and progression by targeting multiple cancer-related genes; however, such regulatory networks are not well characterized in CRC. Thus, the aim of this study was to perform global messenger RNA (mRNA) and microRNA expression profiling in the same CRC samples and adjacent normal tissues and to identify potential miRNA-mRNA regulatory networks. Our data revealed 1273 significantly upregulated and 1902 downregulated genes in CRC. Pathway analysis revealed significant enrichment in cell cycle, integrated cancer, Wnt (wingless-type MMTV integration site family member), matrix metalloproteinase, and TGF-β pathways in CRC. Pharmacological inhibition of Wnt (using XAV939 or IWP-2) or TGF-β (using SB-431542) pathways led to dose- and time-dependent inhibition of CRC cell growth. Similarly, our data revealed up- (42) and downregulated (61) microRNAs in the same matched samples. Using target prediction and bioinformatics, ~77% of the upregulated genes were predicted to be targeted by microRNAs found to be downregulated in CRC. We subsequently focused on EZH2 (enhancer of zeste homolog 2 ), which was found to be regulated by hsa-miR-26a-5p and several members of the let-7 (lethal-7) family in CRC. Significant inverse correlation between EZH2 and hsa-miR-26a-5p (R2=0.56, P=0.0001) and hsa-let-7b-5p (R2=0.19, P=0.02) expression was observed in the same samples, corroborating the belief of EZH2 being a bona fide target for these two miRNAs in CRC. Pharmacological inhibition of EZH2 led to significant reduction in trimethylated histone H3 on lysine 27 (H3K27) methylation, marked reduction in cell proliferation, and migration in vitro. Concordantly, small interfering RNA-mediated knockdown of EZH2 led to similar effects on CRC cell growth in vitro. Therefore, our data have revealed several hundred potential miRNA-mRNA regulatory networks in CRC and suggest targeting relevant networks as potential therapeutic strategy for CRC. PMID:25611389

Genome-wide mRNA and miRNA expression profiling reveal multiple regulatory networks in colorectal cancer.

PubMed

Vishnubalaji, R; Hamam, R; Abdulla, M-H; Mohammed, M A V; Kassem, M; Al-Obeed, O; Aldahmash, A; Alajez, N M

2015-01-22

Despite recent advances in cancer management, colorectal cancer (CRC) remains the third most common cancer and a major health-care problem worldwide. MicroRNAs have recently emerged as key regulators of cancer development and progression by targeting multiple cancer-related genes; however, such regulatory networks are not well characterized in CRC. Thus, the aim of this study was to perform global messenger RNA (mRNA) and microRNA expression profiling in the same CRC samples and adjacent normal tissues and to identify potential miRNA-mRNA regulatory networks. Our data revealed 1273 significantly upregulated and 1902 downregulated genes in CRC. Pathway analysis revealed significant enrichment in cell cycle, integrated cancer, Wnt (wingless-type MMTV integration site family member), matrix metalloproteinase, and TGF-β pathways in CRC. Pharmacological inhibition of Wnt (using XAV939 or IWP-2) or TGF-β (using SB-431542) pathways led to dose- and time-dependent inhibition of CRC cell growth. Similarly, our data revealed up- (42) and downregulated (61) microRNAs in the same matched samples. Using target prediction and bioinformatics, ~77% of the upregulated genes were predicted to be targeted by microRNAs found to be downregulated in CRC. We subsequently focused on EZH2 (enhancer of zeste homolog 2 ), which was found to be regulated by hsa-miR-26a-5p and several members of the let-7 (lethal-7) family in CRC. Significant inverse correlation between EZH2 and hsa-miR-26a-5p (R(2)=0.56, P=0.0001) and hsa-let-7b-5p (R(2)=0.19, P=0.02) expression was observed in the same samples, corroborating the belief of EZH2 being a bona fide target for these two miRNAs in CRC. Pharmacological inhibition of EZH2 led to significant reduction in trimethylated histone H3 on lysine 27 (H3K27) methylation, marked reduction in cell proliferation, and migration in vitro. Concordantly, small interfering RNA-mediated knockdown of EZH2 led to similar effects on CRC cell growth in vitro. Therefore, our data have revealed several hundred potential miRNA-mRNA regulatory networks in CRC and suggest targeting relevant networks as potential therapeutic strategy for CRC.

miR-300 promotes proliferation and EMT-mediated colorectal cancer migration and invasion by targeting p53.

PubMed

Wang, Lin; Yu, Peiwu

2016-12-01

p53 mutations in tumors can induce the loss of wild-type tumor-suppressing p53 function, which results in the increase in proliferation, migration and invasion ability in cancer cells. Studies have shown that the expression of p53 is regulated by several microRNAs (miRNAs). In the present study, we found that miR-300 and p53 were significantly increased in colorectal cancer (CRC) tissues when compared with levels noted in adjacent colorectal tissues. Both miR-300 and p53 were significantly correlated with lymphatic metastasis and TNM stage. Both miR-300 and p53 promoted CRC cell (SW480 and HT29) proliferation, migration, and invasion, respectively, in vitro. In addition, we found that miR-300 is a direct positive regulator of p53 through binding to the binding site in the 3'UTR of the p53 gene in human CRC cells. Moreover, both miR-300 and p53 induced CRC cell epithelial‑mesenchymal transition (EMT) respectively. Taken together, we demonstrated that miR-300 promoted proliferation and EMT-mediated CRC migration and invasion by targeting p53. These findings provide a new theoretical basis and potential therapeutic targets, and thus lays the foundation for exploring the pathogenesis of CRC.

Detection of adult T-cell leukemia virus (ATLV) bearing lymphocytes in concentrated red blood cells derived from ATL associated antibody (ATLA-Ab) positive donors.

PubMed

Morishima, Y; Ohya, K; Ueda, R; Fukuda, T

1986-01-01

Adult T cell leukemia associated antibody (ATLA-Ab) positive persons were screened by indirect immunofluorescence (IF) testing. Their lymphocytes were collected from concentrated red blood cells (CRC), and cultured in vitro with and without phytohemagglutinin (PHA) for 10 days. The expression of ATL virus (ATLV) positive lymphocytes during the in vitro culture was then analyzed by IF assay using mouse monoclonal antibody ATL-19 reactive to p19 core protein of ATLV. 97% of ATLA-Ab positive CRC (36 cases) demonstrated ATLV positive lymphocytes after being cultured for more than 10 days with PHA, whereas, none of ATLA-Ab negative CRC (22 cases) demonstrated ATLV positive lymphocytes. All of the 10 ATLA-Ab positive CRC that were stored for 2, 4, and 7 days contained lymphocytes which expressed ATLV after in vitro culture, while 7 of 10 CRC stored for 14 days and only 1 of 10 CRCs stored for 20 days, expressed ATLV positive lymphocytes. This data indicates that almost all of the ATLA-Ab positive blood contained ATLV positive lymphocytes, and that the in vitro appearance of these ATLV positive lymphocytes was reduced by storing the CRC for more than 14 days.

Cell-Free Circulating Methylated SEPT9 for Noninvasive Diagnosis and Monitoring of Colorectal Cancer

PubMed Central

Fu, Bo; Yan, Peng; Zhang, Shan; Lu, Yan; Pan, Li; Tang, Wenqiang; Chen, Shen; Chen, Shuangfeng

2018-01-01

Identification of early-stage tumor and monitoring therapeutic efficacy and recurrence or metastasis of colorectal cancer (CRC) are urgently warranted for improving the outcome of CRC patients and reducing the disease-related mortality. In this study, we evaluated the diagnostic value of cell-free circulating methylated SEPT9 (mSEPT9) for CRC and beyond CRC and examined the potentiality of mSEPT9 in assessing therapeutic efficacy and monitoring recurrence of CRC. Our results confirmed the favorable diagnostic value of plasma mSEPT9 for CRC, with a sensitivity of 61.22% (95% confidence interval (CI): 51.33%–70.27%) and specificity of 93.7% (95% CI: 91.09%–95.57%) using 2/3 algorithm. The positive rate of mSEPT9 in CRC was correlated with tumor size, histological grade, and general histological type (P < 0.05). Beyond CRC, gastric cancer patients also presented a high positive rate of plasma mSEPT9 (70%). The conversions between preoperative and postoperative plasma mSEPT9 reflected the therapeutic efficacy of curatively intended surgery for CRC patients. The persistent positivity of plasma mSEPT9 after surgery (within 7–14 days) was highly associated with impending recurrences or metastases (within one year), with a sensitivity of 100%. Postoperative mSEPT9 status during follow-up also provided valuable indication for CRC recurrence or metastases, with a good consistency (kappa = 0.818, P = 0.001). Our results verified the reliability of plasma mSEPT9 as a biomarker for noninvasive diagnosis of CRC. More significantly, we revealed its valuable role in appraising CRC therapeutic efficacy and monitoring CRC recurrences or metastases. Further studies with larger sample sizes are needed to verify and elucidate the clinical utility of the promising findings. PMID:29849824

Methylenetetrahydrofolate reductase gene polymorphisms and the risk of colorectal carcinoma in a sample of Egyptian individuals.

PubMed

El Awady, Mostafa K; Karim, Amr M; Hanna, Laila S; El Husseiny, Lamia A; El Sahar, Medhat; Menem, Hanan A Abdel; Meguid, Nagwa A

2009-01-01

The study was planned as a pilot study to investigate two common polymorphisms in the MTHFR gene c.677C > T and c.1298A > C and their association with enhanced risk of colorectal cancer (CRC) in a sample of Egyptian individuals. Venous blood samples were withdrawn from 35 cases of CRC and 68 healthy controls. Specimens from colonic and rectal carcinoma tissues in addition to cancer free tissues were obtained from all cases. Frequencies of MTHFR677T and 1298C alleles were significantly higher among cases of CRC tumor tissues (50% and 56%, respectively) than germ line alleles in CRC patients (33% and 41%, respectively) and healthy controls (21% and 35%, respectively). Frequencies of heterozygous and homoyzgous polymorphisms of MTHFR at positions 677 and 1298 in carcinoma tissues were always the highest. At position 677, TT and CT genotype frequencies were 17% and 66% with an odds ratio {OR} of 11 [95% confidence interval {CI} 2.39-50.59] and OR 8.34 [95%CI 2.97-23.92], respectively, in carcinoma tissues. While in the germ line of patients the genotype frequencies of 677TT and CT were 6% and 54% with OR 1.57 [95%CI 0.26-9.51] and 2.99 [95%CI 1.25-7.12], respectively, compared to controls (6% and 29%, respectively). The combined genotype MTHFR 1298CC + AC frequencies were 86% with OR 3.71 [95%CI 1.28-10.78] in carcinoma tissues, 69% with OR 1.35 [95%CI 0.57-3.21] in germ line of patients and 62% in controls. The combined genotype 677CT plus any of the following genotypes 1298AA, AC or CC enhanced risk of CRC, when comparing germ line DNA polymorphism of patients versus peripheral blood DNA of control subjects with OR 4.5 [95%CI 0.94-21.56], OR 3.12 [95%CI 0.79-12.36] and OR 18 [95%CI 1.56-207.5], respectively, suggesting strong genetic predisposition of certain Egyptian population to CRC. These results suggested that at least one C to T polymorphism at 677MTHFR gene is required to significantly increase the risk for CRC development. Further large scale studies are required to confirm the present findings.

Conserved Region C Functions To Regulate PD-1 Expression and Subsequent CD8 T Cell Memory.

PubMed

Bally, Alexander P R; Tang, Yan; Lee, Joshua T; Barwick, Benjamin G; Martinez, Ryan; Evavold, Brian D; Boss, Jeremy M

2017-01-01

Expression of programmed death 1 (PD-1) on CD8 T cells promotes T cell exhaustion during chronic Ag exposure. During acute infections, PD-1 is transiently expressed and has the potential to modulate CD8 T cell memory formation. Conserved region C (CR-C), a promoter proximal cis-regulatory element that is critical to PD-1 expression in vitro, responds to NFATc1, FoxO1, and/or NF-κB signaling pathways. Here, a CR-C knockout mouse was established to determine its role on PD-1 expression and the corresponding effects on T cell function in vivo. Deletion of CR-C decreased PD-1 expression on CD4 T cells and Ag-specific CD8 T cells during acute and chronic lymphocytic choriomeningitis virus challenges, but did not affect the ability to clear an infection. Following acute lymphocytic choriomeningitis virus infection, memory CD8 T cells in the CR-C knockout mouse were formed in greater numbers, were more functional, and were more effective at responding to a melanoma tumor than wild-type memory cells. These data implicate a critical role for CR-C in governing PD-1 expression, and a subsequent role in guiding CD8 T cell differentiation. The data suggest the possibility that titrating PD-1 expression during CD8 T cell activation could have important ramifications in vaccine development and clinical care. Copyright © 2016 by The American Association of Immunologists, Inc.

Surrogate Endpoints in Second-Line Trials of Targeted Agents in Metastatic Colorectal Cancer: A Literature-Based Systematic Review and Meta-Analysis.

PubMed

Cremolini, Chiara; Antoniotti, Carlotta; Pietrantonio, Filippo; Berenato, Rosa; Tampellini, Marco; Baratelli, Chiara; Salvatore, Lisa; Marmorino, Federica; Borelli, Beatrice; Nichetti, Federico; Bironzo, Paolo; Sonetto, Cristina; Bartolomeo, Maria Di; Braud, Filippo de; Loupakis, Fotios; Falcone, Alfredo; Di Maio, M

2017-07-01

The purpose of this study was to evaluate progression-free survival (PFS) and objective response rate (ORR) as surrogate endpoints of overall survival (OS) in modern clinical trials investigating the efficacy of targeted agents in the second-line treatment of metastatic colorectal cancer (mCRC). A systematic search of literature pertaining to randomized phase II and III trials evaluating targeted agents as second-line treatments for mCRC was performed. The strength of the correlation between both PFS and ORR and OS was assessed based on the Pearson's correlation coefficient (R) and the coefficient of determination (R 2 ). Twenty trials, including a total of 7,571 patients, met the search criteria. The median duration of post-progression survival (PPS) was 7.6 months. The median differences between experimental and control arms were 0.65 months (range, -2.4 to 3.4) for the median PFS and 0.7 months (range, -5.8 to 3.9) for the median OS. PFS and ORR showed moderate (R=0.734, R 2 =0.539, p < 0.001) and poor correlation (R=0.169, R 2 =0.029, p=0.476) with OS, respectively. No differences between anti-angiogenic agents and other drugs were evident. Targeted agents investigated in the second-line treatment of mCRC provided minimal PFS gains translating into modest OS improvements. Considering both the moderate correlation between PFS and OS and the short duration of PPS, the OS should remain the preferred primary endpoint for randomized clinical trials in the second-line treatment of mCRC.

Nanomagnetic Levitation 3-D Cultures of Breast and Colorectal Cancers

PubMed Central

Bumpers, Harvey L.; Janagama, Dasharatham G.; Manne, Upender; Basson, Marc D.; Katkoori, Venkat

2014-01-01

Background Innovative technologies for drug discovery and development, cancer models, stem cell research, tissue engineering, and drug testing in various cell-based platforms require an application similar to the in vivo system. Materials and Methods We developed for the first time nanomagnetically levitated three dimensional (3-D) cultures of breast cancer (BC) and colorectal cancer (CRC) cells using carbon encapsulated cobalt magnetic nanoparticles. BC and CRC xenografts grown in severe combined immunodeficient (SCID) mice were evaluated for N-cadherin and Epidermal growth factor receptor (EGFR) expressions. These phenotypes were compared with 2-D cultures and 3-D cultures grown in a gel matrix. Results The BC and CRC cells grown by magnetic levitation formed microtissues. The levitated cultures had high viability and were maintained in culture for long periods of time. It has been observed that N-cadherin and EGFR activities were highly expressed in the levitated 3-D tumor spheres and xenografts of CRC and BC cells. Conclusions Nanomagnetically levitated 3-D cultures tend to form stable microtissues of BC and CRC and may be more feasible for a range of applications in drug discovery or regenerative medicine. PMID:25617973

The role of RCAS1 as a biomarker in diagnosing CRC and monitoring tumor recurrence and metastasis.

PubMed

Han, Su-xia; Wang, Jing; Wang, Li-juan; Jin, Gui-hua; Ying, Xia; He, Chen-chen; Guo, Xi-jing; Zhang, Jian-ying; Zhang, Ying; Zhu, Qing

2014-06-01

Receptor-binding cancer antigen expressed on SiSo cells (RCAS1) plays an important role in tumor progression by helping tumor cell to escape from host immunological surveillance or modifying the characteristics of connective tissue around. RCAS1 may appropriately reflect the development and prognosis of tumor. In the study, we sought to identify the clinical significance of RCAS1 in colorectal cancer (CRC) diagnosis and tumor recurrence monitoring. Immunohistochemistry (IHC) with tissue array slides was preformed to analyze RCAS1 protein expression in CRC, colorectal polyps, and normal colon tissues. RCAS1 levels in colorectal cancer were significantly higher than those in colorectal polyps and normal colon tissues (P<0.001). Silencing RCAS1 gene in human colonic adenocarcinoma cells decreased cell proliferation and enhanced apoptosis through the p53 signaling pathway. Further analysis by an enzyme-linked immunosorbent assay (ELISA) showed that serum RCAS1 levels in CRC are significantly higher than in healthy controls and polyps (P<0.05), in which the highest serum RCAS1 level is reported in the recurrence group. The serum RCAS1 levels have a significant correlation with clinical stage and pathologic grading. Furthermore, the positive rate of serum RCAS1 in CRC was 82.1 %, which was higher than carcinoembryonic antigen (CEA). Especially in CEA-negative cases, the sensitivity of RCAS1 was 88.2 %. Finally, CRC patients who were followed up showed a serum RCAS1 level which significantly decreased after surgery (P<0.001) and obviously increased in the recurrence group. Taken together, our data demonstrated that RCAS1 is not only a supplementary serological biomarker for CRC diagnosis but also useful for monitoring tumor recurrence. RCAS1 might be a supplementary serological marker for CRC.

Weighted gene co-expression network analysis of colorectal cancer liver metastasis genome sequencing data and screening of anti-metastasis drugs.

PubMed

Gao, Bo; Shao, Qin; Choudhry, Hani; Marcus, Victoria; Dong, Kung; Ragoussis, Jiannis; Gao, Zu-Hua

2016-09-01

Approximately 9% of cancer-related deaths are caused by colorectal cancer (CRC). CRC patients are prone to liver metastasis, which is the most important cause for the high CRC mortality rate. Understanding the molecular mechanism of CRC liver metastasis could help us to find novel targets for the effective treatment of this deadly disease. Using weighted gene co-expression network analysis on the sequencing data of CRC with and with metastasis, we identified 5 colorectal cancer liver metastasis related modules which were labeled as brown, blue, grey, yellow and turquoise. In the brown module, which represents the metastatic tumor in the liver, gene ontology (GO) analysis revealed functions including the G-protein coupled receptor protein signaling pathway, epithelial cell differentiation and cell surface receptor linked signal transduction. In the blue module, which represents the primary CRC that has metastasized, GO analysis showed that the genes were mainly enriched in GO terms including G-protein coupled receptor protein signaling pathway, cell surface receptor linked signal transduction, and negative regulation of cell differentiation. In the yellow and turquoise modules, which represent the primary non-metastatic CRC, 13 downregulated CRC liver metastasis-related candidate miRNAs were identified (e.g. hsa-miR-204, hsa-miR-455, etc.). Furthermore, analyzing the DrugBank database and mining the literature identified 25 and 12 candidate drugs that could potentially block the metastatic processes of the primary tumor and inhibit the progression of metastatic tumors in the liver, respectively. Data generated from this study not only furthers our understanding of the genetic alterations that drive the metastatic process, but also guides the development of molecular-targeted therapy of colorectal cancer liver metastasis.

Overexpression of xeroderma pigmentosum group C decreases the chemotherapeutic sensitivity of colorectal carcinoma cells to cisplatin.

PubMed

Zhang, Yi; Cao, Jia; Meng, Yanni; Qu, Chunying; Shen, Feng; Xu, Leiming

2018-05-01

Xeroderma pigmentosum group C (XPC) is a DNA-damage-recognition gene active at the early stage of DNA repair. XPC also participates in regulation of cell-cycle checkpoint and DNA-damage-induced apoptosis. In the present study, the expression levels of genes involved in nucleotide excision repair (NER) were assessed in human colorectal cancer (CRC) tissue. This analysis revealed that expression of XPC mRNA significantly increased in colorectal carcinoma tissues compared with matched normal controls. Expression of XPC gradually increased along with the degree of progression of CRC. In vitro , an XTT assay demonstrated that small interfering RNA (siRNA) targeting XPC significantly increased the sensitivity of CRC SW480 cells to cisplatin, whereas cells transfected with a XPC-overexpression plasmid became more resistant to cisplatin. Furthermore, flow cytometry revealed that the proportion of apoptotic cells significantly increased in XPC-knockdown cells upon cisplatin treatment. However, the overexpression XPC significantly increased the resistance of cells to cisplatin. In vivo , tumor growth was significantly reduced in tumor-bearing mice when the XPC gene was knocked down. Upregulation of the expression of pro-apoptotic Bcl-associated X and downregulation of the anti-apoptotic B-cell lymphoma 2 proteins was observed in the implanted tumor tissue. In conclusion, XPC serves a key role in chemotherapeutic sensitivity of CRC to cisplatin, meaning that it may be a potential target for chemotherapy of CRC.

MicroRNA-302c represses epithelial-mesenchymal transition and metastasis by targeting transcription factor AP-4 in colorectal cancer.

PubMed

Ma, Wenqi; Liu, Bailing; Li, Jie; Jiang, Jue; Zhou, Ru; Huang, Lili; Li, Xiaopeng; He, Xin; Zhou, Qi

2018-06-12

MicroRNAs (miRNAs) contribute to tumorigenesis and progression via acting as tumor suppressors or oncogenes in human cancer. Aberrant expression of miR-302c has been reported in various types of cancer except colorectal cancer (CRC). Thus, our study was aimed to verify the expression of miR-302c and its functional role in CRC. We found a significant reduced expression of miR-302c in CRC tissues compared to tumor-adjacent tissues. Low miR-302c level was remarkably correlated with deeper tumor invasion, lymph node metastasis and advanced TNM stage. Importantly, low miR-302c expression was identified as an independent indicator for poor prognosis of CRC patients. Overexpression of miR-302c repressed migration and invasion capacities of SW620 and SW480 cells in vitro. Mechanistically, miR-302c inversely regulated transcription factor AP4 (TFAP4) abundance in both SW620 and SW480 cells, and it negatively correlated with TFAP4 mRNA expression in CRC samples. Herein, TFAP4, a regulator of epithelial-mesenchymal transition (EMT), was recognized as a direct target gene of miR-302c in CRC. Otherwise, miR-302c overexpression increased E-cadherin expression and reduced the levels of Vimentin and SNAI1, suggesting an inhibitory effect of miR-302c on EMT of CRC cells. Notably, our findings established that the EMT and metastasis of Caco-2 cells were enhanced by miR-302c knockdown, and subsequently reversed by TFAP4 silencing. Collectively, these data indicate that miR-302c represses EMT and CRC metastasis possibly by targeting TFAP4, and it may serve as a potential prognostic factor and therapeutic target for CRC. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Trichostatin A, a histone deacetylase inhibitor, suppresses JAK2/STAT3 signaling via inducing the promoter-associated histone acetylation of SOCS1 and SOCS3 in human colorectal cancer cells.

PubMed

Xiong, Hua; Du, Wan; Zhang, Yan-Jie; Hong, Jie; Su, Wen-Yu; Tang, Jie-Ting; Wang, Ying-Chao; Lu, Rong; Fang, Jing-Yuan

2012-02-01

Aberrant janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling is involved in the oncogenesis of several cancers. Suppressors of cytokine signaling (SOCS) genes and SH2-containing protein tyrosine phosphatase 1 (SHP1) proteins, which are negative regulators of JAK/STAT signaling, have been reported to have tumor suppressor functions. However, in colorectal cancer (CRC) cells, the mechanisms that regulate SOCS and SHP1 genes, and the cause of abnormalities in the JAK/STAT signaling pathway, remain largely unknown. The present study shows that trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, leads to the hyperacetylation of histones associated with the SOCS1 and SOCS3 promoters, but not the SHP1 promoter in CRC cells. This indicates that histone modifications are involved in the regulation of SOCS1 and SOCS3. Moreover, upregulation of SOCS1 and SOCS3 expression was achieved using TSA, which also significantly downregulated JAK2/STAT3 signaling in CRC cells. We also demonstrate that TSA suppresses the growth of CRC cells, and induces G1 cell cycle arrest and apoptosis through the regulation of downstream targets of JAK2/STAT3 signaling, including Bcl-2, survivin and p16(ink4a) . Therefore, our data demonstrate that TSA may induce SOCS1 and SOCS3 expression by inducing histone modifications and consequently inhibits JAK2/STAT3 signaling in CRC cells. These results also establish a mechanistic link between the inhibition of JAK2/STAT3 signaling and the anticancer action of TSA in CRC cells. Copyright © 2011 Wiley Periodicals, Inc.

Increased expression of interleukin-21 along colorectal adenoma-carcinoma sequence and its predicating significance in patients with sporadic colorectal cancer.

PubMed

Cui, Guanglin; Yuan, Aping; Zhu, Li; Florholmen, Jon; Goll, Rasmus

2017-10-01

The role and significance of interleukin (IL)-21 in the development of sporadic CRC have not been well defined. The aim of this study is therefore to investigate the dynamics of the IL-21 along colorectal adenoma-carcinoma sequence and to evaluate the impact of IL-21 on clinicopathological parameters and CRC prognosis. The real-time PCR results showed that the level of IL-21 in adenomas (n=50) and sporadic CRC (n=50) were significantly higher than that in normal controls (n=18), which were predominately observed in the adenoma/CRC stroma. Analysis revealed that IL-21 level was correlated with the overall survival time in CRC patients. Double immunofluorescence observations confirmed that IL-21 positive cells were mostly natural killer cells and T lymphocytes in the tumor stroma. These results indicate that significant increased IL-21 expression present within the adenoma/CRC microenvironment might have a potential predicating significance for survival time in patients with CRC. Copyright © 2017 Elsevier Inc. All rights reserved.

Carcinoma of the colon and rectum with deregulation of insulin-like growth factor 2 signaling: clinical and molecular implications.

PubMed

Belharazem, Djeda; Magdeburg, Julia; Berton, Ann-Kristin; Beissbarth, Li; Sauer, Christian; Sticht, Carsten; Marx, Alexander; Hofheinz, Ralf; Post, Stefan; Kienle, Peter; Ströbel, Philipp

2016-10-01

Loss of imprinting (LOI) of the insulin-like growth factor 2 (IGF2) is an early event in the development of colorectal cancer (CRC). Whether LOI of IGF2 denotes a molecular or clinical cancer subgroup is currently unknown. Tumor biopsies and paired normal mucosa from 399 patients with extensive clinical annotations were analyzed for LOI and IGF2 expression. LOI status in 140 informative cases was correlated with clinicopathologic parameters and outcome. LOI was frequent in normal mucosa and tumors and occurred throughout the large intestine. LOI was unrelated to microsatellite instability, KRAS mutation status, stage, and survival. However, CRC with LOI showed increased IGF2 protein levels and activation of AKT1. Gene expression analysis of tumors with and without LOI and knockdown of IGF2 in cell lines revealed that IGF2 induced distinct sets of activated and repressed genes, including Wnt5a, CEACAM6, IGF2BP3, KPN2A, BRCA2, and CDK1. Inhibition of AKT1 in IGF2-stimulated cells showed that the downstream effects of IGF2 on cell proliferation and gene expression were strictly AKT1-dependent. LOI of IGF2 is a frequent and early event in CRC that occurs both in the adenomatous polyposis coli (APC) gene-mutated and serrated route of carcinogenesis. LOI leads to overexpression of IGF2, activates IGF1R and AKT1, and is a powerful driver of cell proliferation. Moreover, our results suggest that IGF2 via AKT1 also contributes to non-canonical wnt signaling. Although LOI had no significant impact on major clinical parameters and outcome, its potential as a target for preventive and therapeutic interventions merits further investigation.

Natural compounds and combination therapy in colorectal cancer treatment.

PubMed

Rejhová, A; Opattová, A; Čumová, A; Slíva, D; Vodička, P

2018-01-20

Colorectal cancer (CRC) therapy using conventional chemotherapeutics represents a considerable burden for the patient's organism because of high toxicity while the response is relatively low. Our review summarizes the findings about natural compounds as chemoprotective agents for decreasing risk of CRC. It also identifies natural compounds which possess anti-tumor effects of various characteristics, mainly in vitro on colorectal cell lines or in vivo studies on experimental models, but also in a few clinical trials. Many of natural compounds suppress proliferation by inducing cell cycle arrest or induce apoptosis of CRC cells resulting in the inhibition of tumor growth. A novel employment of natural substances is a so-called combination therapy - administration of two or more substances - conventional chemotherapeutics and a natural compound or more natural compounds at a time. Some natural compounds may sensitize to conventional cytotoxic therapy, reinforce the drug effective concentration, intensify the combined effect of both administered therapeutics or exert cytotoxic effects specifically on tumor cells. Moreover, combined therapy by targeting multiple signaling pathways, uses various mechanisms to reduce the development of resistance to antitumor drugs. The desired effect could be to diminish burden on the patient's organism by replacing part of the dose of a conventional chemotherapeutic with a natural substance with a defined effect. Many natural compounds are well tolerated by the patients and do not cause toxic effects even at high doses. Interaction of conventional chemotherapeutics with natural compounds introduces a new aspect in the research and therapy of cancer. It could be a promising approach to potentially achieve improvements, while minimizing of adverse effects associated with conventional chemotherapy. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Ulcerative colitis-associated colorectal cancer

PubMed Central

Yashiro, Masakazu

2014-01-01

The association between ulcerative colitis (UC) and colorectal cancer (CRC) has been acknowledged. One of the most serious and life threatening consequences of UC is the development of CRC (UC-CRC). UC-CRC patients are younger, more frequently have multiple cancerous lesions, and histologically show mucinous or signet ring cell carcinomas. The risk of CRC begins to increase 8 or 10 years after the diagnosis of UC. Risk factors for CRC with UC patients include young age at diagnosis, longer duration, greater anatomical extent of colonic involvement, the degree of inflammation, family history of CRC, and presence of primary sclerosing cholangitis. CRC on the ground of UC develop from non-dysplastic mucosa to indefinite dysplasia, low-grade dysplasia, high-grade dysplasia and finally to invasive adenocarcinoma. Colonoscopy surveillance programs are recommended to reduce the risk of CRC and mortality in UC. Genetic alterations might play a role in the development of UC-CRC. 5-aminosalicylates might represent a favorable therapeutic option for chemoprevention of CRC. PMID:25469007

[Resting forms of gram negative chemolithoautotrophic bacteria Thioalkalivibrio versutus and Thioalkalimicrobium aerophilum].

PubMed

Loĭko, N G; Soina, V S; Sorokin, D Iu; Mitiushina, L L; El'-Registan, G I

2003-01-01

The haloalkaliphilic chemoautotrophic gram-negative bacteria Thioalkalivibrio versutus, strain AL2, and Thioalkalimicrobium aerophilum, strain AL3, were shown to possess the capacity to produce resting forms, namely cyst-like refractile cells (CRC), whose production was controlled by the level of the d1 extracellular factors, exhibiting the function of anabiosis autoinducers. The conditions were elucidated that promoted the formation of CRC in the developmental cycles of the cultures studied, in condensed cell suspensions undergoing autolysis, and under the action of exogenously introduced chemical analogues of anabiosis autoinducers (alkylhydroxybenzenes). The peculiarities of the fine structure of the resting cells obtained were studied. Distinctions were revealed (with respect to viability and thermotolerance) between the CRC formed under different conditions. The relationship between the growth strategy and survival strategy of extremophilic bacteria is discussed with taking into account the effect of the d1 autoregulatory factors. A new model of CRC formation is proposed: CRC production in the life cycle of bacteria developing under conditions of increased concentration of anabiosis autoinducers.

Novel approach to abuse the hyperactive K-Ras pathway for adenoviral gene therapy of colorectal cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Naumov, Inna; Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv; Kazanov, Dina

2012-01-15

Background: Functional activation of oncogenic K-Ras signaling pathway plays an important role in the early events of colorectal carcinogenesis (CRC). K-Ras proto-oncogene is involved in 35-40% of CRC cases. Mutations in the Ras gene trigger the transduction of proliferative and anti-apoptotic signals, even in the absence of extra cellular stimuli. The objective of the current study was to use a gene-targeting approach to kill human CRC cells selectively harboring mutated K-Ras. Results: A recombinant adenovirus that carries a lethal gene, PUMA, under the control of a Ras responsive promoter (Ad-Py4-SV40-PUMA) was used selectively to target CRC cells (HCT116, SW480, DLD1more » and RIE-Ras) that possess a hyperactive Ras pathway while using HT29 and RIE cells as a control that harbors wild type Ras and exhibit very low Ras activity. Control vector, without the Ras responsive promoter elements was used to assess the specificity of our 'gene therapy' approach. Both adenoviral vectors were assed in vitro and in xenograft model in vivo. Ad-Py4-SV40-PUMA showed high potency to induce {approx} 50% apoptosis in vitro, to abolish completely tumor formation by infecting cells with the Ad-Py4-SV40-PUMA prior xenografting them in nude mice and high ability to suppress by {approx} 35% tumor progression in vivo in already established tumors. Conclusions: Selective targeting of CRC cells with the activated Ras pathway may be a novel and effective therapy in CRC. The high potency of this adenoviral vector may help to overcome an undetectable micro metastasis that is the major hurdle in challenging with CRC.« less

Kefir exhibits anti‑proliferative and pro‑apoptotic effects on colon adenocarcinoma cells with no significant effects on cell migration and invasion.

PubMed

Khoury, Nathalie; El-Hayek, Stephany; Tarras, Omayr; El-Sabban, Marwan; El-Sibai, Mirvat; Rizk, Sandra

2014-11-01

Kefir, a fermented milk product, exhibits anti‑tumoral activity in vivo; yet its mechanism of action remains elusive. Recent studies have focused on the mechanism of action of kefir on cancer cells in vitro. The current study aims at examining the effect of kefir on cell survival, proliferation, and motility of colorectal cancer (CRC) cells. Kefir's anti‑cancer potential was tested on CRC cell lines, Caco‑2 and HT‑29, through cytotoxicity, proliferation, and apoptotic assays. The expression of certain genes involved in proliferation and apoptosis was measured using reverse transcriptase‑polymerase chain reaction (RT‑PCR) and western blotting. To assess the effect of kefir on cancer metastasis, wound‑healing and time‑lapse movies, in addition to collagen‑based invasion assay, were used. The results show that cell‑free fractions of kefir exhibit an anti‑proliferative effect on Caco‑2 and HT‑29 cells. Analysis of DNA content by flow cytometry revealed the ability of kefir to induce cell cycle arrest at the G1 phase. Kefir was also found to induce apoptosis, as seen by cell death ELISA. Results from RT‑PCR showed that kefir decreases the expression of transforming growth factor α (TGF‑α); and transforming growth factor‑β1 (TGF‑β1) in HT‑29 cells. Western blotting results revealed an upregulation in Bax:Bcl‑2 ratio, confirming the pro‑apoptotic effect of kefir, and an increase in p53 independent‑p21 expression upon kefir treatment. MMP expression was not altered by kefir treatment. Furthermore, results from time‑lapse motility movies, wound‑healing, and invasion assays showed no effect on the motility of colorectal as well as breast (MCF‑7 and MB‑MDA‑231) cancer cells upon kefir treatment. Our data suggest that kefir is able to inhibit the proliferation and induce apoptosis in HT‑29 and Caco‑2 CRC cells, yet it does not exhibit a significant effect on the motility and invasion of these cells in vitro.

Origin of Somatic Mutations in β-Catenin versus Adenomatous Polyposis Coli in Colon Cancer: Random Mutagenesis in Animal Models versus Nonrandom Mutagenesis in Humans.

PubMed

Yang, Da; Zhang, Min; Gold, Barry

2017-07-17

Wnt signaling is compromised early in the development of human colorectal cancer (CRC) due to truncating nonsense mutations in adenomatous polyposis coli (APC). CRC induced by chemical carcinogens, such as heterocyclic aromatic amines and azoxymethane, in mice also involves dysregulation of Wnt signaling but via activating missense mutations in the β-catenin oncogene despite the fact that genetically modified mice harboring an inactive APC allele efficiently develop CRC. In contrast, activating mutations in β-catenin are rarely observed in human CRC. Dysregulation of the Wnt signaling pathway by the two distinct mechanisms reveals insights into the etiology of human CRC. On the basis of calculations related to DNA adduct levels produced in mouse CRC models using mutagens, and the number of stem cells in the mouse colon, we show that two nonsense mutations required for biallelic disruption of APC are statistically unlikely to produce CRC in experiments using small numbers of mice. We calculate that an activating mutation in one allele near the critical GSK3β phosphorylation site on β-catenin is >10 5 -times more likely to produce CRC by random mutagenesis due to chemicals than inactivating two alleles in APC, yet it does not occur in humans. Therefore, the mutagenesis mechanism in human CRC cannot be random. We explain that nonsense APC mutations predominate in human CRC because of deamination at 5-methylcytosine at CGA and CAG codons, coupled with the number of human colonic stem cells and lifespan. Our analyses, including a comparison of mutation type and age at CRC diagnosis in U.S. and Chinese patients, also indicate that APC mutations in CRC are not due to environmental mutagens that randomly damage DNA.

Inhibitory effect of maple syrup on the cell growth and invasion of human colorectal cancer cells.

PubMed

Yamamoto, Tetsushi; Uemura, Kentaro; Moriyama, Kaho; Mitamura, Kuniko; Taga, Atsushi

2015-04-01

Maple syrup is a natural sweetener consumed by individuals of all ages throughout the world. Maple syrup contains not only carbohydrates such as sucrose but also various components such as organic acids, amino acids, vitamins and phenolic compounds. Recent studies have shown that these phenolic compounds in maple syrup may possess various activities such as decreasing the blood glucose level and an anticancer effect. In this study, we examined the effect of three types of maple syrup, classified by color, on the cell proliferation, migration and invasion of colorectal cancer (CRC) cells in order to investigate whether the maple syrup is suitable as a phytomedicine for cancer treatment. CRC cells that were administered maple syrup showed significantly lower growth rates than cells that were administered sucrose. In addition, administration of maple syrup to CRC cells caused inhibition of cell invasion, while there was no effect on cell migration. Administration of maple syrup clearly inhibited AKT phosphorylation, while there was no effect on ERK phosphorylation. These data suggest that maple syrup might inhibit cell proliferation and invasion through suppression of AKT activation and be suitable as a phytomedicine for CRC treatment, with fewer adverse effects than traditional chemotherapy.

Inhibitory effect of maple syrup on the cell growth and invasion of human colorectal cancer cells

PubMed Central

YAMAMOTO, TETSUSHI; UEMURA, KENTARO; MORIYAMA, KAHO; MITAMURA, KUNIKO; TAGA, ATSUSHI

2015-01-01

Maple syrup is a natural sweetener consumed by individuals of all ages throughout the world. Maple syrup contains not only carbohydrates such as sucrose but also various components such as organic acids, amino acids, vitamins and phenolic compounds. Recent studies have shown that these phenolic compounds in maple syrup may possess various activities such as decreasing the blood glucose level and an anticancer effect. In this study, we examined the effect of three types of maple syrup, classified by color, on the cell proliferation, migration and invasion of colorectal cancer (CRC) cells in order to investigate whether the maple syrup is suitable as a phytomedicine for cancer treatment. CRC cells that were administered maple syrup showed significantly lower growth rates than cells that were administered sucrose. In addition, administration of maple syrup to CRC cells caused inhibition of cell invasion, while there was no effect on cell migration. Administration of maple syrup clearly inhibited AKT phosphorylation, while there was no effect on ERK phosphorylation. These data suggest that maple syrup might inhibit cell proliferation and invasion through suppression of AKT activation and be suitable as a phytomedicine for CRC treatment, with fewer adverse effects than traditional chemotherapy. PMID:25647359

Peroxiredoxin 5 promotes the epithelial-mesenchymal transition in colon cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ahn, Hye-Mi; Yoo, Jin-Woo; College of Natural Sciences, Kyungpook National University, Daegu

Globally, colorectal cancer (CRC) is common cause of cancer-related deaths. The high mortality rate of patients with colon cancer is due to cancer cell invasion and metastasis. Initiation of the epithelial-to-mesenchymal transition (EMT) is essential for the tumorigenesis. Peroxiredoinxs (PRX1-6) have been reported to be overexpressed in various tumor tissues, and involved to be responsible for tumor progression. However, the exact role of PRX5 in colon cancer remains to be investigated enhancing proliferation and promoting EMT properties. In this study, we constructed stably overexpressing PRX5 and suppressed PRX5 expression in CRC cells. Our results revealed that PRX5 overexpression significantly enhancedmore » CRC cell proliferation, migration, and invasion. On the other hand, PRX5 suppression markedly inhibited these EMT properties. PRX5 was also demonstrated to regulate the expression of two hallmark EMT proteins, E-cadherin and Vimentin, and the EMT-inducing transcription factors, Snail and Slug. Moreover, in the xenograft mouse model, showed that PRX5 overexpression enhances tumor growth of CRC cells. Thus, our findings first provide evidence in CRC that PRX5 promotes EMT properties by inducing the expression of EMT-inducing transcription factors. Therefore, PRX5 can be used as a predictive biomarker and serves as a putative therapeutic target for the development of clinical treatments for human CRC. - Highlights: • PRX5 promoted colorectal cancer cell proliferation. • PRX5 enhanced EMT properties in colorectal cancer. • PRX5 mediated the EMT by inducing the expression of Snail and Slug. • PRX5 promoted tumor growth of colorectal cancer cells.« less

A Specific Mutational Signature Associated with DNA 8-Oxoguanine Persistence in MUTYH-defective Colorectal Cancer

DOE PAGES

Viel, Alessandra; Bruselles, Alessandro; Meccia, Ettore; ...

2017-04-13

8-Oxoguanine, a common mutagenic DNA lesion, generates G:C > T:A transversions via mispairing with adenine during DNA replication. When operating normally, the MUTYH DNA glycosylase prevents 8-oxoguanine-related mutagenesis by excising the incorporated adenine. Biallelic MUTYH mutations impair this enzymatic function and are associated with colorectal cancer (CRC) in MUTYH-Associated Polyposis (MAP) syndrome. Here in this paper, we perform whole-exome sequencing that reveals a modest mutator phenotype in MAP CRCs compared to sporadic CRC stem cell lines or bulk tumours. The excess G:C > T:A transversion mutations in MAP CRCs exhibits a novel mutational signature, termed Signature 36, with a strongmore » sequence dependence. The MUTYH mutational signature reflecting persistent 8-oxoG:A mismatches occurs frequently in the APC, KRAS, PIK3CA, FAT4, TP53, FAT1, AMER1, KDM6A, SMAD4 and SMAD2 genes that are associated with CRC. In conclusion, the occurrence of Signature 36 in other types of human cancer indicates that DNA 8-oxoguanine-related mutations might contribute to the development of cancer in other organs.« less

A Specific Mutational Signature Associated with DNA 8-Oxoguanine Persistence in MUTYH-defective Colorectal Cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Viel, Alessandra; Bruselles, Alessandro; Meccia, Ettore

8-Oxoguanine, a common mutagenic DNA lesion, generates G:C > T:A transversions via mispairing with adenine during DNA replication. When operating normally, the MUTYH DNA glycosylase prevents 8-oxoguanine-related mutagenesis by excising the incorporated adenine. Biallelic MUTYH mutations impair this enzymatic function and are associated with colorectal cancer (CRC) in MUTYH-Associated Polyposis (MAP) syndrome. Here in this paper, we perform whole-exome sequencing that reveals a modest mutator phenotype in MAP CRCs compared to sporadic CRC stem cell lines or bulk tumours. The excess G:C > T:A transversion mutations in MAP CRCs exhibits a novel mutational signature, termed Signature 36, with a strongmore » sequence dependence. The MUTYH mutational signature reflecting persistent 8-oxoG:A mismatches occurs frequently in the APC, KRAS, PIK3CA, FAT4, TP53, FAT1, AMER1, KDM6A, SMAD4 and SMAD2 genes that are associated with CRC. In conclusion, the occurrence of Signature 36 in other types of human cancer indicates that DNA 8-oxoguanine-related mutations might contribute to the development of cancer in other organs.« less

Consensus on management of metastatic colorectal cancer in Central America and the Caribbean: San José, Costa Rica, August 2016

PubMed Central

López, Roberto Ivan; Castro, Jenny Lissette; Cedeño, Heidy; Cisneros, Dagoberto; Corrales, Luis; González-Herrera, Ileana; Lima-Pérez, Mayté; Prestol, Rogelio; Salinas, Roberto; Soriano-García, Jorge Luis; T Zavala, Alejandra; Zetina, Luis Miguel; Zúñiga-Orlich, Carlos Eduardo

2018-01-01

Colorectal cancer (CRC) is the third most common cancer in men and the second most common in women worldwide. In Latin America and the Caribbean, it has a mortality of 56%. The median overall survival for patients with metastatic colorectal cancer (mCRC) is currently estimated as ~30 months, which has substantially improved through strategic changes in treatment and in the management of patients. As opposed to other metastatic cancers where first-line regimens are often determined, mCRC requires special attention because there is controversy in the possible combinations of the available drugs and the different periods of duration for each patient. Each combination must seek to be effective and to generate the minimum adverse effects as possible. Instead of giving the first-line regimen until the tumour progresses, treatment is often individualised. Furthermore, up to 60% of colorectal tumours are considered non-mutated or wild-type CRC. Not harbouring mutations in the RAS family of genes or mutations in the signalling pathways of the epidermal growth factor receptor causes a null response to anti-epidermal growth factor receptor antibody therapy, which implies even more complex considerations regarding its management. The primary objective of this consensus is to address the main scenarios of mCRC in order to warrant the most appropriate therapeutic intervention for these patients in the Central American and the Caribbean (CAC) region. This can lead to better clinical outcomes as well as quality of life for palliative patients. This document includes the formal expert consensus recommendations for scenarios of mutated and non-mutated mCRC, including synchronous or metachronous disease, management of mCRC with liver and lung metastasis, resectable, potentially resectable or non-resectable tumours and local in the CAC context. PMID:29636987

Colorectal cancer chemoprevention: the potential of a selective approach.

PubMed

Ben-Amotz, Oded; Arber, Nadir; Kraus, Sarah

2010-10-01

Colorectal cancer (CRC) is a leading cause of cancer death, and therefore demands special attention. Novel recent approaches for the chemoprevention of CRC focus on selective targeting of key pathways. We review the study by Zhang and colleagues, evaluating a selective approach targeting APC-deficient premalignant cells using retinoid-based therapy and TNF-related apoptosis-inducing ligand (TRAIL). This study demonstrates that induction of TRAIL-mediated death signaling contributes to the chemopreventive value of all-trans-retinyl acetate (RAc) by sensitizing premalignant adenoma cells for apoptosis without affecting normal cells. We discuss these important findings, raise few points that deserve consideration, and may further contribute to the development of RAc-based combination therapies with improved efficacy. The authors clearly demonstrate a synergistic interaction between TRAIL, RAc and APC, which leads to the specific cell death of premalignant target cells. The study adds to the growing body of literature related to CRC chemoprevention, and provides solid data supporting a potentially selective approach for preventing CRC using RAc and TRAIL.

SDHB downregulation facilitates the proliferation and invasion of colorectal cancer through AMPK functions excluding those involved in the modulation of aerobic glycolysis.

PubMed

Xiao, Zhiming; Liu, Shaojun; Ai, Feiyan; Chen, Xiong; Li, Xiayu; Liu, Rui; Ren, Weiguo; Zhang, Xuemei; Shu, Peng; Zhang, Decai

2018-01-01

Loss-of-function of succinate dehydrogenase-B (SDHB) is a predisposing factor of aerobic glycolysis and cancer progression. Adenosine monophosphate activated protein kinase (AMPK) is involved in the regulation of aerobic glycolysis and the diverse hallmarks of cancer. The present study investigated whether AMPK mediated the regulatory effects of SDHB in aerobic glycolysis and cancer growth. The expression of SDHB and AMPK in colorectal cancer (CRC) and normal tissues was assessed by western blotting. HT-29 CRC cells were used to establish in vitro models of ectopic overexpression and knockdown of SDHB. SDHB was downregulated, while AMPK and phosphorylated-AMPK (Thr172) were upregulated in CRC tissues. Experiments involving the loss- or gain-of-function of SDHB, revealed that this protein negatively regulated AMPK by influencing its expression and activity. However, SDHB and AMPK were identified to suppress lactic acid production in CRC cells, indicating that each had an inhibitory effect on aerobic glycolysis. Therefore, the regulation of aerobic glycolysis by SDHB is unlikely to be mediated via AMPK. SDHB knockdown promoted the viability, migration and invasion of HT-29 cells, whereas inhibition of AMPK demonstrated the opposite effect. SDHB overexpression impaired cell migration and invasion, and this effect was reversed following AMPK activation. These results indicate that AMPK may mediate the effects of SDHB in CRC cell proliferation and migration. In conclusion, SDHB downregulation in CRC cells may increase AMPK activity, which may subsequently facilitate the proliferation and invasion of these cancer cells. However, the regulation of aerobic glycolysis by SDHB may be independent of AMPK. Further studies are warranted to elucidate the mechanism by which SDHB regulates aerobic glycolysis.

Treatment of colorectal peritoneal carcinomatosis with systemic chemotherapy: a pooled analysis of north central cancer treatment group phase III trials N9741 and N9841.

PubMed

Franko, Jan; Shi, Qian; Goldman, Charles D; Pockaj, Barbara A; Nelson, Garth D; Goldberg, Richard M; Pitot, Henry C; Grothey, Axel; Alberts, Steven R; Sargent, Daniel J

2012-01-20

Symptoms and complications of metastatic colorectal cancer (mCRC) differ by metastatic sites. There is a paucity of prospective survival data for patients with peritoneal carcinomatosis colorectal cancer (pcCRC). We characterized outcomes of patients with pcCRC enrolled onto two prospective randomized trials of chemotherapy and contrasted that with other manifestations of mCRC (non-pcCRC). A total of 2,095 patients enrolled onto two prospective randomized trials were evaluated for overall survival (OS) and progression-free survival (PFS). A Cox proportional hazard model was used to assess the adjusted associations. The characteristics of the pcCRC group (n = 364) were similar to those of the non-pcCRC patients in median age (63 v 61 years, P = .23), sex (57% males v 61%, P = .23), and performance status (Eastern Cooperative Oncology Group performance status 0 or 1 94% v 96%, P = .06), but differed in frequency of liver (63% v 82%, P < .001) and lung metastases (27% v 34%, P = .01). Median OS (12.7 v 17.6 months, hazard ratio [HR] = 1.3; 95% CI, 1.2 to 1.5; P < .001) and PFS (5.8 v 7.2 months, HR = 1.2; 95% CI, 1.1 to 1.3; P = .001) were shorter for pcCRC versus non-pcCRC. The unfavorable prognostic influence of pcCRC remained after adjusting for age, PS, liver metastases, and other factors (OS: HR = 1.3, P < .001; PFS: HR = 1.1, P = .02). Infusional fluorouracil, leucovorin, and oxaliplatin was superior to irinotecan, leucovorin, and fluorouracil as a first-line treatment among pcCRC (HR for OS = 0.62, P = .005) and non-pcCRC patients (HR = 0.66, P < .001). pcCRC is associated with a significantly shorter OS and PFS as compared with other manifestations of mCRC. Future trials for mCRC should consider stratifying on the basis of pcCRC status.

The metastasis suppressor, NDRG1, inhibits “stemness” of colorectal cancer via down-regulation of nuclear β-catenin and CD44

PubMed Central

Wangpu, Xiongzhi; Yang, Xiao; Zhao, Jingkun; Lu, Jiaoyang; Guan, Shaopei; Lu, Jun; Kovacevic, Zaklina; Liu, Wensheng; Mi, Lan; Jin, Runsen; Sun, Jing; Yue, Fei; Ma, Junjun; Lu, Aiguo; Richardson, Des R.; Wang, Lishun; Zheng, Minhua

2015-01-01

N-myc downstream-regulated gene 1 (NDRG1), has been identified as an important metastasis suppressor for colorectal cancer (CRC). In this study, we investigated: (1) the effects of NDRG1 on CRC stemness and tumorigenesis; (2) the molecular mechanisms involved; and (3) the relationship between NDRG1 expression and colorectal cancer prognosis. Our investigation demonstrated that CRC cells with silenced NDRG1 showed more tumorigenic ability and stem cell-like properties, such as: colony and sphere formation, chemoresistance, cell invasion, high expression of CD44, and tumorigenicity in vivo. Moreover, NDRG1 silencing reduced β-catenin expression on the cell membrane, while increasing its nuclear expression. The anti-tumor activity of NDRG1 was demonstrated to be mediated by preventing β-catenin nuclear translocation, as silencing of this latter molecule could reverse the effects of silencing NDRG1 expression. NDRG1 expression was also demonstrated to be negatively correlated to CRC prognosis. In addition, there was a negative correlation between NDRG1 and nuclear β-catenin and also NDRG1 and CD44 expression in clinical CRC specimens. Taken together, our investigation demonstrates that the anti-metastatic activity of NDRG1 in CRC occurs through the down-regulation of nuclear β-catenin and suggests that NDRG1 is a significant therapeutic target. PMID:26418878

Interplay between diet, gut microbiota, epigenetic events, and colorectal cancer

PubMed Central

Bultman, Scott J.

2016-01-01

Despite the success of colonoscopy screening, colorectal cancer (CRC) remains one of the most common and deadly cancers, and CRC incidence is rising in some countries where screening is not routine and populations have recently switched from traditional diets to western diets. Diet and energy balance influence CRC by multiple mechanisms. They modulate the composition and function of gut microbiota, which have a prodigious metabolic capacity and can produce oncometabolites or tumor-suppressive metabolites depending, in part, on which dietary factors and digestive components are present in the GI tract. Gut microbiota also have a profound effect on immune cells in the lamina propria, which influences inflammation and subsequently CRC. Nutrient availability, which is an outcome of diet and energy balance, determines the abundance of certain energy metabolites that are essential co-factors for epigenetic enzymes and therefore impinges upon epigenetic regulation of gene expression. Aberrant epigenetic marks accumulate during CRC, and epimutations that are selected for drive tumorigenesis by causing transcriptome profiles to diverge from the cell of origin. In some instances, the above mechanisms are intertwined as exemplified by dietary fiber being metabolized by colonic bacteria into butyrate, which is both a short-chain fatty acid (SCFA) and a histone deacetylase (HDAC) inhibitor that epigenetically upregulates tumor-suppressor genes in CRC cells and anti-inflammatory genes in immune cells. PMID:27138454

High expression of PTBP1 promote invasion of colorectal cancer by alternative splicing of cortactin.

PubMed

Wang, Zhi-Na; Liu, Dan; Yin, Bin; Ju, Wen-Yi; Qiu, Hui-Zhong; Xiao, Yi; Chen, Yuan-Jia; Peng, Xiao-Zhong; Lu, Chong-Mei

2017-05-30

Polypyrimidine tract-binding protein 1 (PTBP1) involving in almost all steps of mRNA regulation including alternative splicing metabolism during tumorigenesis due to its RNA-binding activity. Initially, we found that high expressed PTBP1 and poor prognosis was interrelated in colorectal cancer (CRC) patients with stages II and III CRC, which widely different in prognosis and treatment, by immunohistochemistry. PTBP1 was also upregulated in colon cancer cell lines. In our study, knockdown of PTBP1 by siRNA transfection decreased cell proliferation and invasion in vitro. Denovirus shRNA knockdown of PTBP1 inhibited colorectal cancer growth in vivo. Furthermore, PTBP1 regulates alternative splicing of many target genes involving in tumorgenesis in colon cancer cells. We confirmed that the splicing of cortactin exon 11 which was only contained in cortactin isoform-a, as a PTBP1 target. Knockdown of PTBP1 decreased the expression of cortactin isoform-a by exclusion of exon 11. Also the mRNA levels of PTBP1 and cortactin isoform-a were cooperatively expressed in colorectal cancer tissues. Knocking down cortactin isoform-a significantly decreased cell migration and invasion in colorectal cancer cells. Overexpression of cortactin isoform-a could rescue PTBP1-knockdown effect of cell motility. In summary the study revealed that PTBP1 facilitates colorectal cancer migration and invasion activities by inclusion of cortactin exon 11.

Amphiregulin and PTEN evoke a multimodal mechanism of acquired resistance to PI3K inhibition

PubMed Central

Edgar, Kyle A.; Crocker, Lisa; Cheng, Eric; Wagle, Marie-Claire; Wongchenko, Matthew; Yan, Yibing; Wilson, Timothy R.; Dompe, Nicholas; Neve, Richard M.; Belvin, Marcia; Sampath, Deepak; Friedman, Lori S.; Wallin, Jeffrey J.

2014-01-01

Phosphoinositide-3 kinase (PI3K) signaling pathway alterations occur broadly in cancer and PI3K is a promising therapeutic target. Here, we investigated acquired resistance to GDC-0941, a PI3K inhibitor in clinical trials. Colorectal cancer (CRC) cells made to be resistant to GDC-0941 were discovered to secrete amphiregulin, which resulted in increased EGFR/MAPK signaling. Moreover, prolonged PI3K pathway inhibition in cultured cells over a period of months led to a secondary loss of PTEN in 40% of the CRC lines with acquired resistance to PI3K inhibition. In the absence of PI3K inhibitor, these PTEN-null PI3K inhibitor-resistant clones had elevated PI3K pathway signaling and decreased sensitivity to MAPK pathway inhibitors. Importantly, PTEN loss was not able to induce resistance to PI3K inhibitors in the absence of amphiregulin, indicating a multimodal mechanism of acquired resistance. The combination of PI3K and MAPK pathway inhibitors overcame acquired resistance in vitro and in vivo. PMID:25053989

Amphiregulin and PTEN evoke a multimodal mechanism of acquired resistance to PI3K inhibition.

PubMed

Edgar, Kyle A; Crocker, Lisa; Cheng, Eric; Wagle, Marie-Claire; Wongchenko, Matthew; Yan, Yibing; Wilson, Timothy R; Dompe, Nicholas; Neve, Richard M; Belvin, Marcia; Sampath, Deepak; Friedman, Lori S; Wallin, Jeffrey J

2014-03-01

Phosphoinositide-3 kinase (PI3K) signaling pathway alterations occur broadly in cancer and PI3K is a promising therapeutic target. Here, we investigated acquired resistance to GDC-0941, a PI3K inhibitor in clinical trials. Colorectal cancer (CRC) cells made to be resistant to GDC-0941 were discovered to secrete amphiregulin, which resulted in increased EGFR/MAPK signaling. Moreover, prolonged PI3K pathway inhibition in cultured cells over a period of months led to a secondary loss of PTEN in 40% of the CRC lines with acquired resistance to PI3K inhibition. In the absence of PI3K inhibitor, these PTEN-null PI3K inhibitor-resistant clones had elevated PI3K pathway signaling and decreased sensitivity to MAPK pathway inhibitors. Importantly, PTEN loss was not able to induce resistance to PI3K inhibitors in the absence of amphiregulin, indicating a multimodal mechanism of acquired resistance. The combination of PI3K and MAPK pathway inhibitors overcame acquired resistance in vitro and in vivo.

Left Versus Right: Does Location Matter for Refractory Metastatic Colorectal Cancer Patients in Phase 1 Clinical Trials?

PubMed

Arora, Sukeshi Patel; Ketchum, Norma S; Michalek, Joel; Gelfond, Jonathon; Mahalingam, Devalingam

2017-04-22

Location of the primary tumor is prognostic and predictive of efficacy with VEGF-inhibitors (I) versus EGFR-I given first-line to metastatic colorectal cancer (mCRC) patients. However, little is known regarding the effect of location on prognosis and prediction in refractory mCRC. We assessed the efficacy of VEGF-I and EGFR-I in regards to location of the primary tumor in patients with refractory mCRC enrolled in early phase studies. A historical cohort analysis of mCRC patients, including 44 phase I trials our institution, from March 2004 to September 2012. Median Progression free survival (mPFS) and overall survival (mOS) were estimated from Kaplan-Meier curves and groups were statistically compared with the log-rank test. One hundred thirty-nine patients with a median age 59 (33-81). 73.9% received 3+ lines of therapy. All KRAS wild-type patients had received prior EGFR-I. right 20.9%, left 61.9%, and transverse 4.3%. For survival analysis, transverse CRC were included with right. Of the 112 patients, mOS was left (N = 80) 6.6 months versus right (N = 32) 5.9 months, P = 0.18. mPFS was left (n = 86) 2.0 months versus right (N = 35) 2.0 months, P = 0.76. In subgroup analysis, survival was significant for KRAS wild-type patients with left-sided mCRC had mOS of 6.2 months with other agents versus 9.4 months with EGFR-I (P = 0.03). In phase 1 clinical trials, although location alone was not prognostic in heavily pretreated patients, left-sided mCRC had improved survival with EGFR-I. Despite progression on EGFR-I, left-sided KRAS wild mCRC patients should be considered for phase 1 studies of agents targeting growth factor pathways.

Overexpression of xeroderma pigmentosum group C decreases the chemotherapeutic sensitivity of colorectal carcinoma cells to cisplatin

PubMed Central

Zhang, Yi; Cao, Jia; Meng, Yanni; Qu, Chunying; Shen, Feng; Xu, Leiming

2018-01-01

Xeroderma pigmentosum group C (XPC) is a DNA-damage-recognition gene active at the early stage of DNA repair. XPC also participates in regulation of cell-cycle checkpoint and DNA-damage-induced apoptosis. In the present study, the expression levels of genes involved in nucleotide excision repair (NER) were assessed in human colorectal cancer (CRC) tissue. This analysis revealed that expression of XPC mRNA significantly increased in colorectal carcinoma tissues compared with matched normal controls. Expression of XPC gradually increased along with the degree of progression of CRC. In vitro, an XTT assay demonstrated that small interfering RNA (siRNA) targeting XPC significantly increased the sensitivity of CRC SW480 cells to cisplatin, whereas cells transfected with a XPC-overexpression plasmid became more resistant to cisplatin. Furthermore, flow cytometry revealed that the proportion of apoptotic cells significantly increased in XPC-knockdown cells upon cisplatin treatment. However, the overexpression XPC significantly increased the resistance of cells to cisplatin. In vivo, tumor growth was significantly reduced in tumor-bearing mice when the XPC gene was knocked down. Upregulation of the expression of pro-apoptotic Bcl-associated X and downregulation of the anti-apoptotic B-cell lymphoma 2 proteins was observed in the implanted tumor tissue. In conclusion, XPC serves a key role in chemotherapeutic sensitivity of CRC to cisplatin, meaning that it may be a potential target for chemotherapy of CRC. PMID:29616110

In Hyperthermia Increased ERK and WNT Signaling Suppress Colorectal Cancer Cell Growth

PubMed Central

Bordonaro, Michael; Shirasawa, Senji; Lazarova, Darina L.

2016-01-01

Although neoplastic cells exhibit relatively higher sensitivity to hyperthermia than normal cells, hyperthermia has had variable success as an anti-cancer therapy. This variable outcome might be due to the fact that cancer cells themselves have differential degrees of sensitivity to high temperature. We hypothesized that the varying sensitivity of colorectal cancer (CRC) cells to hyperthermia depends upon the differential induction of survival pathways. Screening of such pathways revealed that Extracellular Signal-Regulated Kinase (ERK) signaling is augmented by hyperthermia, and the extent of this modulation correlates with the mutation status of V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS). Through clonal growth assays, apoptotic analyses and transcription reporter assays of CRC cells that differ only in KRAS mutation status we established that mutant KRAS cells are more sensitive to hyperthermia, as they exhibit sustained ERK signaling hyperactivation and increased Wingless/Integrated (WNT)/beta-catenin signaling. We propose that whereas increased levels of WNT and ERK signaling and a positive feedback between the two pathways is a major obstacle in anti-cancer therapy today, under hyperthermia the hyperinduction of the pathways and their positive crosstalk contribute to CRC cell death. Ascertaining the causative association between types of mutations and hyperthermia sensitivity may allow for a mutation profile-guided application of hyperthermia as an anti-cancer therapy. Since KRAS and WNT signaling mutations are prevalent in CRC, our results suggest that hyperthermia-based therapy might benefit a significant number, but not all, CRC patients. PMID:27187477

MicroRNA-466 (miR-466) functions as a tumor suppressor and prognostic factor in colorectal cancer (CRC).

PubMed

Tong, Feng; Ying, Youhua; Pan, Haihua; Zhao, Wei; Li, Hongchen; Zhan, Xiaoli

2018-01-17

MicroRNAs (miRNAs) have an important role in the regulation of tumor development and metastasis. In this study, we investigated the clinical and prognostic value as well as biological function of miR-466 in colorectal cancer (CRC). Tumor and adjacent healthy tissues were obtained from 100 patients diagnosed with CRC. miR-466 expression was determined by quantitative reverse transcription polymerase chain reaction (qRT-PCR). mRNA and protein levels of cyclin D1, apoptosis regulator BAX (BAX), and matrix metalloproteinase-2 (MMP-2) were analyzed by qRT-PCR and Western blot, respectively, in SW-620 CRC cells transfected with miR-466 mimics or negative control miRNA. Effects of miR-466 on SW-620 cell proliferation, cell cycle and apoptosis, and invasion were investigated using CCK-8 assay, flow cytometry and Transwell assay, respectively. miR-466 expression was significantly downregulated in tumor tissues compared to matched adjacent non-tumor tissues. Low expression of miR-466 was significantly correlated with the tumor size, Tumor Node Metastasis stage, lymph node metastasis, and distant metastasis. The overall survival of CRC patients with low miR-466 expression was significantly shorter compared to high-miR-466 expression group (log-rank test: p = 0.0103). Multivariate analysis revealed that low miR-466 expression was associated with poor prognosis in CRC patients. The ectopic expression of miR-466 suppressed cell proliferation and migration/invasion, as well as induced G0/G1 arrest and apoptosis in SW-620 cells. Moreover, the ectopic expression of miR-466 decreased the expression of cyclin D1 and MMP-2, but increased BAX expression in SW-620 cells. In conclusion, our findings demonstrated that miR-466 functions as a suppressor miRNA in CRC and may be used as a prognostic factor in these patients.

The epigenetic modifier PBRM1 restricts the basal activity of the innate immune system by repressing retinoic acid-inducible gene-I-like receptor signalling and is a potential prognostic biomarker for colon cancer.

PubMed

Shu, Xing-Sheng; Zhao, Yingying; Sun, Yanmei; Zhong, Lan; Cheng, Yingduan; Zhang, Yixiang; Ning, Kaile; Tao, Qian; Wang, Yejun; Ying, Ying

2018-01-01

It has long been known that patients suffering from inflammatory bowel disease (IBD) have an increased risk of developing colorectal cancer (CRC). The innate immune system of host cells provides a first-line defence against pathogenic infection, whereas an uncontrolled inflammatory response under homeostatic conditions usually leads to pathological consequences, as exemplified by the chronic inflammation of IBD. The key molecules and pathways keeping innate immunity in check are still poorly defined. Here, we report that the chromatin remodeller polybromo-1 (PBRM1) is a repressor of innate immune signalling mediated by retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs). Knockdown of PBRM1 in colon cancer cells increased the expression of two receptor genes (RIG-I and MDA5) and upregulated interferon (IFN)-related and inflammation-related gene signatures. The innate immune signal stimulated by a double-stranded RNA viral mimic was exaggerated by PBRM1 suppression. PBRM1 cooperated with polycomb protein EZH2 to directly bind the cis-regulatory elements of RIG-I and MDA5, thereby suppressing their transcription. Moreover, upregulation of RIG-I and MDA5 is required for IFN response activation induced by PBRM1 silencing. TRIM25, a protein stimulated by the RLR pathway and IFN production, physically interacted with PBRM1 and induced PBRM1 protein destabilization by promoting its ubiquitination. These findings reveal a PBRM1-RLR regulatory circuit that can keep innate immune activity at a minimal level in resting cells, and also ensure a robust inflammatory response in the case of pathogen invasion. PBRM1 was found to be downregulated in primary tissues from patients with CRC or IBD, and its expression correlated negatively with that of RLR genes and interferon-stimulated genes in CRC samples. Lower PBRM1 expression was associated with advanced pathological grade and poorer survival of CRC patients, indicating that PBRM1 could serve as a potential prognostic biomarker for CRC. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Pseudomonas putida growing at low temperature shows increased levels of CrcZ and CrcY sRNAs, leading to reduced Crc-dependent catabolite repression.

PubMed

Fonseca, Pilar; Moreno, Renata; Rojo, Fernando

2013-01-01

The Crc protein of Pseudomonas inhibits the expression of genes involved in the transport and assimilation of a number of non-preferred carbon sources when preferred substrates are available, thus coordinating carbon metabolism. Crc acts by binding to target mRNAs, inhibiting their translation. In Pseudomonas putida, the amount of free Crc available is controlled by two sRNAs, CrcY and CrcZ, which bind to and sequester Crc. The levels of these sRNAs vary according to metabolic conditions. Pseudomonas putida grows optimally at 30°C, but can also thrive at 10°C. The present work shows that when cells grow exponentially at 10°C, the repressive effect of Crc on many genes is significantly reduced compared with that seen at 30°C. Total Crc levels were similar at both temperatures, but those of CrcZ and CrcY were significantly higher at 10°C. Therefore, Crc-mediated repression may, at least in part, be reduced at 10°C because the fraction of Crc protein sequestered by CrcZ and CrcY is larger, reducing the amount of free Crc available to bind its targets. This may help P. putida to face cold stress. The results reported might help understanding the behaviour of this bacterium in bioremediation or rhizoremediation strategies at low temperatures. © 2012 Society for Applied Microbiology and Blackwell Publishing Ltd.

Colorectal cancer cell-derived exosomes containing miR-10b regulate fibroblast cells via the PI3K/Akt pathway.

PubMed

Dai, Guangyao; Yao, Xiaoguang; Zhang, Yubin; Gu, Jianbin; Geng, Yunfeng; Xue, Fei; Zhang, Jingcheng

2018-04-01

Cancer-associated fibroblasts (CAFs) contribute to the proliferation of colorectal cancer(CRC) cells. However, the mechanism by which CAFs develop in the tumor microenvironment remains unknown. Exosomes may be involved in activating CAFs. Using a miRNA expression profiling array, we determined the miRNA expression profile of secretory exosomes in CRC cells and then identified potential miRNAs with significant differential expression compared to normal cells via enrichment analysis. Predicted targets of candidate miRNAs were then assessed via bioinformatics analysis. Realtime qPCR, western blot, and cell cycle analyses were performed to evaluate the role of candidate exosomal miRNAs. Luciferase reporter assays were applied to confirm whether candidate exosomal miRNAs control target pathway expression. A CRC xenograft mouse model was constructed to evaluate tumor growth in vivo. Exosomes from CRC cells contained significantly higher levels of miR-10b than did exosomes from normal colorectal epithelial cells. Moreover, exosomes containing miR-10b were transferred to fibroblasts. Bioinformatics analysis identified PIK3CA, as a potential target of miR-10b. Luciferase reporter assays confirmed that miR-10b directly inhibited PIK3CA expression. Co-culturing fibroblasts with exosomes containing miR-10b significantly suppressed PIK3CA expression and decreased PI3K/Akt/mTOR pathway activity. Finally, exosomes containing miR-10b reduced fibroblast proliferation but promoted expression of TGF-β and SM α-actin, suggesting that exosomal miR-10b may activate fibroblasts to become CAFs that express myofibroblast markers. These activated fibroblasts were able to promote CRC growth in vitro and in vivo. CRC-derived exosomes actively promote disease progression by modulating surrounding stromal cells, which subsequently acquire features of CAFs. Copyright © 2018 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.

Methylation status of the APC and RASSF1A promoter in cell-free circulating DNA and its prognostic role in patients with colorectal cancer.

PubMed

Matthaios, Dimitrios; Balgkouranidou, Ioanna; Karayiannakis, Anastasios; Bolanaki, Helen; Xenidis, Nikolaos; Amarantidis, Kyriakos; Chelis, Leonidas; Romanidis, Konstantinos; Chatzaki, Aikaterini; Lianidou, Evi; Trypsianis, Grigorios; Kakolyris, Stylianos

2016-07-01

DNA methylation is the most frequent epigenetic alteration. Using methylation-specific polymerase chain reaction (MSP), the methylation status of the adenomatous polyposis coli ( APC ) and Ras association domain family 1 isoform A ( RASSF1A ) genes was examined in cell-free circulating DNA from 155 plasma samples obtained from patients with early and advanced colorectal cancer (CRC). APC and RASSF1A hypermethylation was frequently observed in both early and advanced disease, and was significantly associated with a poorer disease outcome. The methylation status of the APC and RASSF1A promoters was investigated in cell-free DNA of patients with CRC. Using MSP, the promoter methylation status of APC and RASSF1A was examined in 155 blood samples obtained from patients with CRC, 88 of whom had operable CRC (oCRC) and 67 had metastatic CRC (mCRC). The frequency of APC methylation in patients with oCRC was 33%. Methylated APC promoter was significantly associated with older age (P=0.012), higher stage (P=0.014) and methylated RASSF1A status (P=0.050). The frequency of APC methylation in patients with mCRC was 53.7%. In these patients, APC methylation was significantly associated with methylated RASSF1A status (P=0.016). The frequency of RASSF1A methylation in patients with oCRC was 25%. Methylated RASSF1A in oCRC was significantly associated with higher stage (P=0.021). The frequency of RASSF1A methylation in mCRC was 44.8%. Methylated RASSF1A in mCRC was associated with moderate differentiation (P=0.012), high levels of carcinoembryonic antigen (P=0.023) and methylated APC status (P=0.016). Patients with an unmethylated APC gene had better survival in both early (81±5 vs. 27±4 months, P<0.001) and advanced disease (37±7 vs. 15±3 months, P<0.001), compared with patients with methylated APC . Patients with an unmethylated RASSF1A gene had better survival in both early (71±6 vs. 46±8 months, P<0.001) and advanced disease (28±4 vs. 16±3 months, P<0.001) than patients with methylated RASSF1A . The observed significant correlations between APC and RASSF1A promoter methylation status and survival may be indicative of a prognostic role for these genes in CRC, which requires additional testing in larger studies.

Reduced migration of MLH1 deficient colon cancer cells depends on SPTAN1.

PubMed

Hinrichsen, Inga; Ernst, Benjamin Philipp; Nuber, Franziska; Passmann, Sandra; Schäfer, Dieter; Steinke, Verena; Friedrichs, Nicolaus; Plotz, Guido; Zeuzem, Stefan; Brieger, Angela

2014-01-24

Defects in the DNA mismatch repair (MMR) protein MLH1 are frequently observed in sporadic and hereditary colorectal cancers (CRC). Affected tumors generate much less metastatic potential than the MLH1 proficient forms. Although MLH1 has been shown to be not only involved in postreplicative MMR but also in several MMR independent processes like cytoskeletal organization, the connection between MLH1 and metastasis remains unclear. We recently identified non-erythroid spectrin αII (SPTAN1), a scaffolding protein involved in cell adhesion and motility, to interact with MLH1. In the current study, the interaction of MLH1 and SPTAN1 and its potential consequences for CRC metastasis was evaluated. Nine cancer cell lines as well as fresh and paraffin embedded colon cancer tissue from 12 patients were used in gene expression studies of SPTAN1 and MLH1. Co-expression of SPTAN1 and MLH1 was analyzed by siRNA knock down of MLH1 in HeLa, HEK293, MLH1 positive HCT116, SW480 and LoVo cells. Effects on cellular motility were determined in MLH1 deficient HCT116 and MLH1 deficient HEK293T compared to their MLH1 proficient sister cells, respectively. MLH1 deficiency is clearly associated with SPTAN1 reduction. Moreover, siRNA knock down of MLH1 decreased the mRNA level of SPTAN1 in HeLa, HEK293 as well as in MLH1 positive HCT116 cells, which indicates a co-expression of SPTAN1 by MLH1. In addition, cellular motility of MLH1 deficient HCT116 and MLH1 deficient HEK293T cells was impaired compared to the MLH1 proficient sister clones. Consequently, overexpression of SPTAN1 increased migration of MLH1 deficient cells while knock down of SPTAN1 decreased cellular mobility of MLH1 proficient cells, indicating SPTAN1-dependent migration ability. These data suggest that SPTAN1 levels decreased in concordance with MLH1 reduction and impaired cellular mobility in MLH1 deficient colon cancer cells. Therefore, aggressiveness of MLH1-positive CRC might be related to SPTAN1.

Reduced migration of MLH1 deficient colon cancer cells depends on SPTAN1

PubMed Central

2014-01-01

Introduction Defects in the DNA mismatch repair (MMR) protein MLH1 are frequently observed in sporadic and hereditary colorectal cancers (CRC). Affected tumors generate much less metastatic potential than the MLH1 proficient forms. Although MLH1 has been shown to be not only involved in postreplicative MMR but also in several MMR independent processes like cytoskeletal organization, the connection between MLH1 and metastasis remains unclear. We recently identified non-erythroid spectrin αII (SPTAN1), a scaffolding protein involved in cell adhesion and motility, to interact with MLH1. In the current study, the interaction of MLH1 and SPTAN1 and its potential consequences for CRC metastasis was evaluated. Methods Nine cancer cell lines as well as fresh and paraffin embedded colon cancer tissue from 12 patients were used in gene expression studies of SPTAN1 and MLH1. Co-expression of SPTAN1 and MLH1 was analyzed by siRNA knock down of MLH1 in HeLa, HEK293, MLH1 positive HCT116, SW480 and LoVo cells. Effects on cellular motility were determined in MLH1 deficient HCT116 and MLH1 deficient HEK293T compared to their MLH1 proficient sister cells, respectively. Results MLH1 deficiency is clearly associated with SPTAN1 reduction. Moreover, siRNA knock down of MLH1 decreased the mRNA level of SPTAN1 in HeLa, HEK293 as well as in MLH1 positive HCT116 cells, which indicates a co-expression of SPTAN1 by MLH1. In addition, cellular motility of MLH1 deficient HCT116 and MLH1 deficient HEK293T cells was impaired compared to the MLH1 proficient sister clones. Consequently, overexpression of SPTAN1 increased migration of MLH1 deficient cells while knock down of SPTAN1 decreased cellular mobility of MLH1 proficient cells, indicating SPTAN1-dependent migration ability. Conclusions These data suggest that SPTAN1 levels decreased in concordance with MLH1 reduction and impaired cellular mobility in MLH1 deficient colon cancer cells. Therefore, aggressiveness of MLH1-positive CRC might be related to SPTAN1. PMID:24456667

Effect of API-1 and FR180204 on cell proliferation and apoptosis in human DLD-1 and LoVo colorectal cancer cells

PubMed Central

Saglam, Atiye Seda Yar; Alp, Ebru; Elmazoglu, Zubeyir; Menevse, Emine Sevda

2016-01-01

The activation of the phosphatidylinositol-3 kinase/v-akt murine thymoma viral oncogene homolog (Akt) and mitogen activated protein kinase kinase/extracellular signal-regulated kinase (ERK) pathways are implicated in the majority of cancers. Selective inhibition of Akt and ERK represents a potential approach for cancer therapy. Therefore, the present study aimed to investigate the apoptotic and anti-proliferative effects of the novel and selective Akt inhibitor 4-amino-5,8-dihydro-5-oxo-8-β-D-ribofuranosyl-pyrido[2,3-d]pyrimidine-6-carboxamide (API-1) and selective ERK1/2 inhibitor FR180204 (FR) alone and in combination on colorectal cancer (CRC) cells (DLD-1 and LoVo). In addition, the effects of API-1 and FR on Akt and ERK signaling pathways were also investigated. The effects of the agents on DLD-1 and LoVo cells were evaluated in terms of cell viability, cytotoxicity, DNA synthesis rate, DNA fragmentation and caspase-3 activity levels. In addition, quantitative reverse transcription-polymerase chain reaction and western blot analysis were performed to examine relevant mRNA and protein levels. The present study observed that the combination of FR with API-1 resulted in significant apoptosis and cytotoxicity compared with any single agent alone in a time-dependent manner in these cells. Also, treatment with FR and API-1 in combination decreased the expression levels of B-cell lymphoma-2 (BCL2), Bcl-2-like1, cyclin D1 and cMYC, and increased the expression levels of BCL2-associated X protein and BCL2 antagonist/killer via phosphorylated Akt and phosphorylated ERK1/2 downregulation. The combination of Akt and ERK1/2 inhibitors resulted in enhanced apoptotic and anti-proliferative effects against CRC cells. The present study hypothesizes that the combination of FR and API-1 in CRC cells may contribute toward potential anti-carcinogenic effects. Additional analyses using other cancer cell lines and animal models are required to confirm these findings in vitro and in vivo. PMID:27698814

Effect of API-1 and FR180204 on cell proliferation and apoptosis in human DLD-1 and LoVo colorectal cancer cells.

PubMed

Saglam, Atiye Seda Yar; Alp, Ebru; Elmazoglu, Zubeyir; Menevse, Emine Sevda

2016-10-01

The activation of the phosphatidylinositol-3 kinase/v-akt murine thymoma viral oncogene homolog (Akt) and mitogen activated protein kinase kinase/extracellular signal-regulated kinase (ERK) pathways are implicated in the majority of cancers. Selective inhibition of Akt and ERK represents a potential approach for cancer therapy. Therefore, the present study aimed to investigate the apoptotic and anti-proliferative effects of the novel and selective Akt inhibitor 4-amino-5,8-dihydro-5-oxo-8-β-D-ribofuranosyl-pyrido[2,3-d]pyrimidine-6-carboxamide (API-1) and selective ERK1/2 inhibitor FR180204 (FR) alone and in combination on colorectal cancer (CRC) cells (DLD-1 and LoVo). In addition, the effects of API-1 and FR on Akt and ERK signaling pathways were also investigated. The effects of the agents on DLD-1 and LoVo cells were evaluated in terms of cell viability, cytotoxicity, DNA synthesis rate, DNA fragmentation and caspase-3 activity levels. In addition, quantitative reverse transcription-polymerase chain reaction and western blot analysis were performed to examine relevant mRNA and protein levels. The present study observed that the combination of FR with API-1 resulted in significant apoptosis and cytotoxicity compared with any single agent alone in a time-dependent manner in these cells. Also, treatment with FR and API-1 in combination decreased the expression levels of B-cell lymphoma-2 (BCL2), Bcl-2-like1, cyclin D1 and cMYC, and increased the expression levels of BCL2-associated X protein and BCL2 antagonist/killer via phosphorylated Akt and phosphorylated ERK1/2 downregulation. The combination of Akt and ERK1/2 inhibitors resulted in enhanced apoptotic and anti-proliferative effects against CRC cells. The present study hypothesizes that the combination of FR and API-1 in CRC cells may contribute toward potential anti-carcinogenic effects. Additional analyses using other cancer cell lines and animal models are required to confirm these findings in vitro and in vivo .

Effect of Crc and Hfq proteins on the transcription, processing, and stability of the Pseudomonas putida CrcZ sRNA

PubMed Central

Hernández-Arranz, Sofía; Sánchez-Hevia, Dione; Rojo, Fernando; Moreno, Renata

2016-01-01

In Pseudomonas putida, the Hfq and Crc proteins regulate the expression of many genes in response to nutritional and environmental cues, by binding to mRNAs that bear specific target motifs and inhibiting their translation. The effect of these two proteins is antagonized by the CrcZ and CrcY small RNAs (sRNAs), the levels of which vary greatly according to growth conditions. The crcZ and crcY genes are transcribed from promoters PcrcZ and PcrcY, respectively, a process that relies on the CbrB transcriptional activator and the RpoN σ factor. Here we show that crcZ can also be transcribed from the promoter of the immediate upstream gene, cbrB, a weak constitutive promoter. The cbrB-crcZ transcript was processed to render a sRNA very similar in size to the CrcZ produced from promoter PcrcZ. The processed sRNA, termed CrcZ*, was able to antagonize Hfq/Crc because, when provided in trans, it relieved the deregulated Hfq/Crc-dependent hyperrepressing phenotype of a ΔcrcZΔcrcY strain. CrcZ* may help in attaining basal levels of CrcZ/CrcZ* that are sufficient to protect the cell from an excessive Hfq/Crc-dependent repression. Since a functional sRNA can be produced from PcrcZ, an inducible strong promoter, or by cleavage of the cbrB-crcZ mRNA, crcZ can be considered a 3′-untranslated region of the cbrB-crcZ mRNA. In the absence of Hfq, the processed form of CrcZ was not observed. In addition, we show that Crc and Hfq increase CrcZ stability, which supports the idea that these proteins can form a complex with CrcZ and protect it from degradation by RNases. PMID:27777366

Effect of Crc and Hfq proteins on the transcription, processing, and stability of the Pseudomonas putida CrcZ sRNA.

PubMed

Hernández-Arranz, Sofía; Sánchez-Hevia, Dione; Rojo, Fernando; Moreno, Renata

2016-12-01

In Pseudomonas putida, the Hfq and Crc proteins regulate the expression of many genes in response to nutritional and environmental cues, by binding to mRNAs that bear specific target motifs and inhibiting their translation. The effect of these two proteins is antagonized by the CrcZ and CrcY small RNAs (sRNAs), the levels of which vary greatly according to growth conditions. The crcZ and crcY genes are transcribed from promoters PcrcZ and PcrcY, respectively, a process that relies on the CbrB transcriptional activator and the RpoN σ factor. Here we show that crcZ can also be transcribed from the promoter of the immediate upstream gene, cbrB, a weak constitutive promoter. The cbrB-crcZ transcript was processed to render a sRNA very similar in size to the CrcZ produced from promoter PcrcZ The processed sRNA, termed CrcZ*, was able to antagonize Hfq/Crc because, when provided in trans, it relieved the deregulated Hfq/Crc-dependent hyperrepressing phenotype of a ΔcrcZΔcrcY strain. CrcZ* may help in attaining basal levels of CrcZ/CrcZ* that are sufficient to protect the cell from an excessive Hfq/Crc-dependent repression. Since a functional sRNA can be produced from PcrcZ, an inducible strong promoter, or by cleavage of the cbrB-crcZ mRNA, crcZ can be considered a 3'-untranslated region of the cbrB-crcZ mRNA. In the absence of Hfq, the processed form of CrcZ was not observed. In addition, we show that Crc and Hfq increase CrcZ stability, which supports the idea that these proteins can form a complex with CrcZ and protect it from degradation by RNases. © 2016 Hernández-Arranz et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

COL1A1 promotes metastasis in colorectal cancer by regulating the WNT/PCP pathway

PubMed Central

Zhang, Zheying; Wang, Yongxia; Zhang, Jinghang; Zhong, Jiateng; Yang, Rui

2018-01-01

Colorectal cancer (CRC) is the third leading cause of cancer-associated mortality, and is a major health problem. Collagen type I α 1 (COL1A1) is a major component of collagen type I. Recently, it was reported to be overexpressed in a variety of tumor tissues and cells. However, the function of COL1A1 in CRC remains unclear. Herein, the present study demonstrated that COL1A1 was upregulated in CRC tissues and the paired lymph node tissues. Transwell assays showed that COL1A1 promoted CRC cell migration in vitro. Moreover, it was revealed that COL1A1 levels were correlated with those of WNT/planar cell polarity (PCP) signaling pathway genes; inhibition of COL1A1 decreased the expression levels of Ras-related C3 botulinum toxin substrate 1-GTP, phosphorylated-c-Jun N-terminal kinase, and RhoA-GTP, all of which are key genes in the WNT/PCP signaling pathway. These results may indicate the mechanisms underlying the oncogenic role of COL1A1 in CRC. In summary, the present data indicated that COL1A1 may serve as an oncoprotein, and that it may be used as a potential therapeutic target in CRC. PMID:29393423

SRC activates TAZ for intestinal tumorigenesis and regeneration.

PubMed

Byun, Mi Ran; Hwang, Jun-Ha; Kim, A Rum; Kim, Kyung Min; Park, Jung Il; Oh, Ho Taek; Hwang, Eun Sook; Hong, Jeong-Ho

2017-12-01

Proto-oncogene tyrosine-protein kinase Src (cSRC) is involved in colorectal cancer (CRC) development and damage-induced intestinal regeneration, although the cellular mechanisms involved are poorly understood. Here, we report that transcriptional coactivator with PDZ binding domain (TAZ) is activated by cSRC, regulating CRC cell proliferation and tumor formation, where cSRC overexpression increases TAZ expression in CRC cells. In contrast, knockdown of cSRC decreases TAZ expression. Additionally, direct phosphorylation of TAZ at Tyr316 by cSRC stimulates nuclear localization and facilitates transcriptional enhancer factor TEF-3 (TEAD4)-mediated transcription. However, a TAZ phosphorylation mutant significantly decreased cell proliferation, wound healing, colony forming, and tumor formation. In a CRC mouse model, Apc Min/+ , activated SRC expression was associated with increased TAZ expression in polyps and TAZ depletion decreased polyp formation. Moreover, intestinal TAZ knockout mice had intestinal regeneration defects following γ-irradiation. Finally, significant correspondence between SRC activation and TAZ overexpression was observed in CRC patients. These results suggest that TAZ is a critical factor for SRC-mediated intestinal tumor formation and regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

The effect of individualized nutritional counseling on muscle mass and treatment outcome in patients with metastatic colorectal cancer undergoing chemotherapy: a randomized controlled trial protocol.

PubMed

van der Werf, Anne; Blauwhoff-Buskermolen, Susanne; Langius, Jacqueline A E; Berkhof, Johannes; Verheul, Henk M W; de van der Schueren, Marian A E

2015-03-05

A low muscle mass is prevalent in patients with metastatic colorectal cancer (mCRC) and has been associated with poor treatment outcome. Chemotherapeutic treatment has an additional unfavorable effect on muscle mass. Sufficient protein intake and physical activity are known to induce muscle protein anabolism in healthy individuals, however it is unclear whether optimal nutrition is effective to preserve muscle mass in patients with mCRC during first-line chemotherapy as well. We hypothesize that individual nutritional counseling by a trained dietitian during first-line chemotherapy is effective in preserving muscle mass and may improve clinical outcomes in patients with mCRC. In this multi-center single-blind randomized controlled trial, patients with mCRC scheduled for first-line combination chemotherapy consisting of oxaliplatin and fluoropyrimidine, with or without bevacizumab (n = 110), will be randomized to receive either individualized nutritional counseling by a trained dietitian to achieve a sufficient dietary intake and an adequate physical activity level, or usual care. Outcome measures will be assessed at baseline and after two and four months of treatment. The primary endpoint will be the change in skeletal muscle area (measured by CT-scan) at the first treatment evaluation. Secondary endpoints will be quality of life, physical functioning, treatment toxicity, treatment intensity and survival. Statistical analyses include one-sided t-tests for the primary endpoint and mixed models and the Kaplan-Meier method for secondary endpoints. This randomized controlled trial will provide evidence whether individualized nutritional counseling during chemotherapy is effective in preventing loss of muscle mass in patients with mCRC. ClinicalTrials.gov NCT01998152 ; Netherlands Trial Register NTR4223.

First-Line Cetuximab Monotherapy in KRAS/NRAS/BRAF Mutation-Negative Colorectal Cancer Patients.

PubMed

Moiseyenko, Vladimir M; Moiseyenko, Fedor V; Yanus, Grigoriy A; Kuligina, Ekatherina Sh; Sokolenko, Anna P; Bizin, Ilya V; Kudriavtsev, Alexey A; Aleksakhina, Svetlana N; Volkov, Nikita M; Chubenko, Vyacheslav A; Kozyreva, Kseniya S; Kramchaninov, Mikhail M; Zhuravlev, Alexandr S; Shelekhova, Kseniya V; Pashkov, Denis V; Ivantsov, Alexandr O; Venina, Aigul R; Sokolova, Tatyana N; Preobrazhenskaya, Elena V; Mitiushkina, Natalia V; Togo, Alexandr V; Iyevleva, Aglaya G; Imyanitov, Evgeny N

2018-06-01

Colorectal carcinomas (CRCs) are sensitive to treatment by anti-epidermal growth factor receptor (EGFR) antibodies only if they do not carry activating mutations in down-stream EGFR targets (KRAS/NRAS/BRAF). Most clinical trials for chemo-naive CRC patients involved combination of targeted agents and chemotherapy, while single-agent cetuximab or panitumumab studies included either heavily pretreated patients or subjects who were not selected on the basis of molecular tests. We hypothesized that anti-EGFR therapy would have significant efficacy in chemo-naive patients with KRAS/NRAS/BRAF mutation-negative CRC. Nineteen patients were prospectively included in the study. Two (11%) patients experienced partial response (PR) and 11 (58%) subjects showed stable disease (SD). Median time to progression approached 6.1 months (range 1.6-15.0 months). Cetuximab efficacy did not correlate with RNA expression of EGFR and insulin-like growth factor 2 (IGF2). Only one tumor carried PIK3CA mutation, and this CRC responded to cetuximab. Exome analysis of patients with progressive disease (PD) revealed 1 CRC with high-level microsatellite instability and 1 instance of HER2 oncogene amplification; 3 of 4 remaining patients with PD had allergic reactions to cetuximab, while none of the subjects with PR or SD had this complication. Comparison with 19 retrospective KRAS/NRAS/BRAF mutation-negative patients receiving first-line fluoropyrimidines revealed no advantages or disadvantages of cetuximab therapy. Cetuximab demonstrates only modest efficacy when given as a first-line monotherapy to KRAS/NRAS/BRAF mutation-negative CRC patients. It is of question, why meticulous patient selection, which was undertaken in the current study, did not result in the improvement of outcomes of single-agent cetuximab treatment.

Par3L enhances colorectal cancer cell survival by inhibiting Lkb1/AMPK signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Taiyuan; Liu, Dongning; Lei, Xiong

Partitioning defective 3-like protein (Par3L) is a recently identified cell polarity protein that plays an important role in mammary stem cell maintenance. Previously, we showed that high expression of Par3L is associated with poor survival in malignant colorectal cancer (CRC), but the underlying mechanism remained unknown. To this end, we established a Par3L knockout colorectal cancer cell line using the CRISPR/Cas system. Interestingly, reduced proliferation, enhanced cell death and caspase-3 activation were observed in Par3L knockout (KO) cells as compared with wildtype (WT) cells. Consistent with previous studies, we showed that Par3L interacts with a tumor suppressor protein liver kinasemore » B1 (Lkb1). Moreover, Par3L depletion resulted in abnormal activation of Lkb1/AMPK signaling cascade. Knockdown of Lkb1 in these cells could significantly reduce AMPK activity and partially rescue cell death caused by Par3L knockdown. Furthermore, we showed that Par3L KO cells were more sensitive to chemotherapies and irradiation. Together, these results suggest that Par3L is essential for colorectal cancer cell survival by inhibiting Lkb1/AMPK signaling pathway, and is a putative therapeutic target for CRC. - Highlights: • Par3L knockout using the CRISPR/Cas system induces apoptosis in colorectal cancer cells. • Par3L interacts with Lkb1 and regulates the activity of AMPK signaling cascade. • Par3L knockout cells are more sensitive to treatment of different chemotherapy drugs and irradiation.« less

Overexpression of HOXA4 and HOXA9 genes promotes self-renewal and contributes to colon cancer stem cell overpopulation.

PubMed

Bhatlekar, Seema; Viswanathan, Vignesh; Fields, Jeremy Z; Boman, Bruce M

2018-02-01

Because HOX genes encode master regulatory transcription factors that regulate stem cells (SCs) during development and aberrant expression of HOX genes occurs in various cancers, our goal was to determine if dysregulation of HOX genes is involved in the SC origin of colorectal cancer (CRC). We previously reported that HOXA4 and HOXD10 are expressed in the colonic SC niche and are overexpressed in CRC. HOX gene expression was studied in SCs from human colon tissue and CRC cells (CSCs) using qPCR and immunostaining. siRNA-mediated knockdown of HOX expression was used to evaluate the role of HOX genes in modulating cancer SC (CSC) phenotype at the level of proliferation, SC marker expression, and sphere formation. All-trans-retinoic-acid (ATRA), a differentiation-inducing agent was evaluated for its effects on HOX expression and CSC growth. We found that HOXA4 and HOXA9 are up-regulated in CRC SCs. siRNA knockdown of HOXA4 and HOXA9 reduced: (i) proliferation and sphere-formation and (ii) gene expression of known SC markers (ALDH1, CD166, LGR5). These results indicate that proliferation and self-renewal ability of CRC SCs are reduced in HOXA4 and HOXA9 knockdown cells. ATRA decreased HOXA4, HOXA9, and HOXD10 expression in parallel with reduction in ALDH1 expression, self-renewal, and proliferation. Overall, our findings indicate that overexpression of HOXA4 and HOXA9 contributes to self-renewal and overpopulation of SCs in CRC. Strategies designed to modulate HOX expression may provide ways to target malignant SCs and to develop more effective therapies for CRC. © 2017 Wiley Periodicals, Inc.

Combinatorial anticancer effects of curcumin and 5-fluorouracil loaded thiolated chitosan nanoparticles towards colon cancer treatment.

PubMed

Anitha, A; Deepa, N; Chennazhi, K P; Lakshmanan, Vinoth-Kumar; Jayakumar, R

2014-09-01

Evaluation of the combinatorial anticancer effects of curcumin/5-fluorouracil loaded thiolated chitosan nanoparticles (CRC-TCS-NPs/5-FU-TCS-NPs) on colon cancer cells and the analysis of pharmacokinetics and biodistribution of CRC-TCS-NPs/5-FU-TCS-NPs in a mouse model. CRC-TCS-NPs/5-FU-TCS-NPs were developed by ionic cross-linking. The in vitro combinatorial anticancer effect of the nanomedicine was proven by different assays. Further the pharmacokinetics and biodistribution analyses were performed in Swiss Albino mouse using HPLC. The 5-FU-TCS-NPs (size: 150±40nm, zeta potential: +48.2±5mV) and CRC-TCS-NPs (size: 150±20nm, zeta potential: +35.7±3mV) were proven to be compatible with blood. The in vitro drug release studies at pH4.5 and 7.4 showed a sustained release profile over a period of 4 days, where both the systems exhibited a higher release in acidic pH. The in vitro combinatorial anticancer effects in colon cancer (HT29) cells using MTT, live/dead, mitochondrial membrane potential and cell cycle analysis measurements confirmed the enhanced anticancer effects (2.5 to 3 fold). The pharmacokinetic studies confirmed the improved plasma concentrations of 5-FU and CRC up to 72h, unlike bare CRC and 5-FU. To conclude, the combination of 5-FU-TCS-NPs and CRC-TCS-NPs showed enhanced anticancer effects on colon cancer cells in vitro and improved the bioavailability of the drugs in vivo. The enhanced anticancer effects of combinatorial nanomedicine are advantageous in terms of reduction in the dosage of 5-FU, thereby improving the chemotherapeutic efficacy and patient compliance of colorectal cancer cases. Copyright © 2014 Elsevier B.V. All rights reserved.

Integrated analysis identifies microRNA-195 as a suppressor of Hippo-YAP pathway in colorectal cancer.

PubMed

Sun, Min; Song, Haibin; Wang, Shuyi; Zhang, Chunxiao; Zheng, Liang; Chen, Fangfang; Shi, Dongdong; Chen, Yuanyuan; Yang, Chaogang; Xiang, Zhenxian; Liu, Qing; Wei, Chen; Xiong, Bin

2017-03-29

With persistent inconsistencies in colorectal cancer (CRC) miRNAs expression data, it is crucial to shift toward inclusion of a "pre-laboratory" integrated analysis to expedite effective precision medicine and translational research. Aberrant expression of hsa-miRNA-195 (miR-195) which is distinguished as a clinically noteworthy miRNA has previously been observed in multiple cancers, yet its role in CRC remains unclear. In this study, we performed an integrated analysis of seven CRC miRNAs expression datasets. The expression of miR-195 was validated in The Cancer Genome Atlas (TCGA) datasets, and an independent validation sample cohort. Colon cancer cells were transfected with miR-195 mimic and inhibitor, after which cell proliferation, colony formation, migration, invasion, and dual luciferase reporter were assayed. Xenograft mouse models were used to determine the role of miR-195 in CRC tumorigenicity in vivo. Four downregulated miRNAs (hsa-let-7a, hsa-miR-125b, hsa-miR-145, and hsa-miR-195) were demonstrated to be potentially useful diagnostic markers in the clinical setting. CRC patients with a decreased level of miR-195-5p in tumor tissues had significantly shortened survival as revealed by the TCGA colon adenocarcinoma (COAD) dataset and our CRC cohort. Overexpression of miR-195-5p in DLD1 and HCT116 cells repressed cell growth, colony formation, invasion, and migration. Inhibition of miR-195-5p function contributed to aberrant cell proliferation, migration, invasion, and epithelial mesenchymal transition (EMT). We identified miR-195-5p binding sites within the 3'-untranslated region (3'-UTR) of the human yes-associated protein (YAP) mRNA. YAP1 expression was downregulated after miR-195-5p treatment by qRT-PCR analysis and western blot. Four downregulated miRNAs were shown to be prime candidates for a panel of biomarkers with sufficient diagnostic accuracy for CRC in a clinical setting. Our integrated microRNA profiling approach identified miR-195-5p independently associated with prognosis in CRC. Our results demonstrated that miR-195-5p was a potent suppressor of YAP1, and miR-195-5p-mediated downregulation of YAP1 significantly reduced tumor development in a mouse CRC xenograft model. In the clinic, miR-195-5p can serve as a prognostic marker to predict the outcome of the CRC patients.

Inhibition of p70S6K1 activation by Pdcd4 overcomes the resistance to an IGF-1R/IR inhibitor in colon carcinoma cells

PubMed Central

Zhang, Yan; Wang, Qing; Chen, Li; Yang, Hsin-Sheng

2015-01-01

Agents targeting insulin-like growth factor 1 receptor (IGF-1R) are being actively examined in clinical trials. Although there has been some initial success of single agent targeting IGF-1R, attempts in later studies failed due to resistance. This study aimed to understand the effects of programmed cell death 4 (Pdcd4) on the chemosensitivity of the IGF-1R inhibitor, OSI-906, in colorectal cancer (CRC) cells and the mechanism underlying this impact. Using OSI-906 resistant and sensitive CRC cells, we found that the Pdcd4 level directly correlates with cell chemosensitivity to OSI-906. In addition, tumors derived from Pdcd4 knockdown cells resist the growth inhibitory effect of OSI-906 in a CRC xenograft mouse model. Moreover, Pdcd4 enhances the antiproliferative effect of OSI-906 in resistant cells through suppression of p70S6K1 activation. Knockdown of p70S6K1, but not p70S6K2, significantly increases the chemosensitivity of OSI-906 in cultured CRC cells. Furthermore, the combination of OSI-906 and PF4708671, a p70S6K1 inhibitor, efficiently suppresses the growth of OSI-906 resistant colon tumor cells in vitro and in vivo. Taken together, activation of p70S6K1 that is inhibited by Pdcd4 is essential for resistance to IGF-1R inhibitor in colon tumor cells, and the combinational treatment of OSI-906 and PF-4708671 results in enhanced antiproliferation effects in CRC cells in vitro and in vivo, providing a novel venue to overcome the resistance to IGF-1R inhibitor in treating colorectal cancer. PMID:25573956

Circulating follicular helper T cells in Crohn's disease (CD) and CD-associated colorectal cancer.

PubMed

Wang, Zhenlong; Wang, Zhiming; Diao, Yanqing; Qian, Xiaoli; Zhu, Nan; Dong, Wen

2014-09-01

Follicular helper T cells (Tfh) represent a distinct subset of CD4+ T cells specialized in providing help to B lymphocytes. Studies have indicated that Tfh in circulating blood can act as a prognostic marker for diseases. In the current study, we investigated the percentages of circulating Tfh (CTfh) in Crohn's disease (CD) and CD-associated colorectal cancer (CRC). CTfh and it subtypes were determined by measuring CD3, CD4, CXCR5, CXCR3, and CCR6 using flow cytometry in 32 healthy controls and 78 CD patients, which included 16 CD-associated CRC. Data showed that proportion of CTfh in CD4+ T cells was significantly increased in CD patients (9.8 %) than in controls (5.1 %) (p < 0.01). Further analysis revealed that the upregulation of CTfh was contributed by CTfh-Th1 subtype and CTfh-Th17 subtype. Investigating the behavior of the patients demonstrated that prevalence of CTfh was significantly elevated in penetrating CD (20.9 %) than inflammatory CD (8.2 %) or stricturing CD (7.5 %). In addition, we analyzed CTfh in CD-associated CRC, and identified that patients with CRC had 1.59-fold higher percentage of CTfh than patients without CRC (p < 0.01). Furthermore, the distribution of CTfh subsets was significantly altered in patients with the cancer. This study suggests the involvement of CTfh in CD and CD-associated CRC, in which the effect of CTfh is partially different between these two diseases.

Genetic instability caused by loss of MutS homolog 3 in human colorectal cancer

PubMed Central

Haugen, Astrid C.; Goel, Ajay; Yamada, Kanae; Marra, Giancarlo; Nguyen, Thuy-Phuong; Nagasaka, Takeshi; Kanazawa, Shinsaku; Koike, Junichi; Kikuchi, Yoshinori; Zhong, Xiaoling; Arita, Michitsune; Shibuya, Kazutoshi; Oshimura, Mitsuo; Hemmi, Hiromichi; Boland, Clement Richard; Koi, Minoru

2008-01-01

Microsatellite instability (MSI) is a hallmark of mismatch repair deficiency. High levels of MSI at mono- and dinucleotide repeats in colorectal cancer (CRC) are attributed to inactivation of the mismatch repair genes, hMLH1 and hMSH2. CRC with low levels of MSI (MSI-L) exists; however its molecular basis is unclear. There is another type of MSI - “elevated microsatellite alterations at selected tetranucleotide repeats” - (EMAST) where loci containing [AAAG]n or [ATAG]n repeats are unstable. EMAST is frequent in non-colorectal cancers; however the incidence of EMAST and its cause in CRC is not known. Here, we report that MSH3-knock-down or MSH3-deficient cells exhibit the EMAST phenotype and low levels of mutations at dinucleotide repeats. About 60% of 117 sporadic CRC cases exhibit EMAST. All of the cases defined as MSI-H (16 cases) exhibited high levels of EMAST. Among 101 non-MSI-H cases, all 19 cases of MSI-L and 35 of 82 cases of MSS exhibited EMAST. Although non-MSI-H CRC tissues contained MSH3-negative tumor cells ranging from 2-50% of the total tumor cell population, the tissues exhibiting EMAST contained more MSH3-negative cells (average 31.5%) than did the tissues not exhibiting EMAST (8.4%). Taken together, our results support the idea that MSH3-deficiency causes EMAST or EMAST with low levels of MSI at the loci with dinucleotide repeats in CRC. PMID:18922920

Inflammatory and redox reactions in colorectal carcinogenesis.

PubMed

Guina, Tina; Biasi, Fiorella; Calfapietra, Simone; Nano, Mario; Poli, Giuseppe

2015-03-01

It has been established that there is a relationship between chronic inflammation and cancer development. The constant colonic inflammation typical of inflammatory bowel diseases is now considered a risk factor for colorectal carcinoma (CRC) development. The inflammatory network of signaling molecules is also required during the late phases of carcinogenesis, to enable cancer cells to survive and to metastasize. Oxidative reactions are an integral part of the inflammatory response, and are generally associated with CRC development. However, when the malignant phenotype is acquired, increased oxidative status induces antioxidant defenses in cancer cells, favoring their aggressiveness. This contradictory behavior of cancer cells toward redox status is of great significance for potential anticancer therapies. This paper summarizes the essential background information relating to the molecules involved in regulating oxidative stress and inflammation during carcinogenesis. Understanding more of their function in CRC stages might provide the foundation for future developments in CRC treatment. © 2015 New York Academy of Sciences.

Significance of the E3 ubiquitin protein UBR5 as an oncogene and a prognostic biomarker in colorectal cancer

PubMed Central

Xu, Xiaowen; Zhu, Yan; Guo, Aizhen; Shen, Xian; Cao, Fuao; Chang, Wenjun

2017-01-01

The E3 ubiquitin protein UBR5 has been implicated in the regulation of multiple biological functions and has recently emerged as a key regulator of the ubiquitin-proteasome system (UPS) in cancer. However, the clinical significance and biological function of UBR5 in colorectal cancer (CRC) are poorly understood. In this study, we compared the expression pattern of UBR5 between CRC and adjacent normal tissues and found that UBR5 expression was frequently elevated in CRC, possibly through chromosomal gains. Using three CRC patient cohorts, we found that patients with high UBR5 mRNA levels, UBR5 gene amplification, or high nuclear UBR5 protein levels had poor prognoses. Multivariate analysis showed that the alterations in UBR5 were independent predictors of CRC prognosis with the TNM stage as a confounding factor. Furthermore, knockdown of UBR5 prevented the proliferation, colony formation, migration, and invasion of CRC cells in cell culture models. An in vivo animal model further confirmed that UBR5 knockdown reduced the growth of CRC tumors. In conclusion, our study is the first to systematically investigate the clinical and biological significance of UBR5 and to conclude that an elevated UBR5 level plays an oncogenic role and may be a potential prognostic marker in CRC. PMID:29296225

The tumor suppressor miR-138-5p targets PD-L1 in colorectal cancer

PubMed Central

Zhao, Lian; Yu, Haibo; Yi, Shuijing; Peng, Xiaowei; Su, Peng; Xiao, Zhiming; Liu, Rui; Tang, Anliu; Li, Xiayu; Liu, Fen; Shen, Shourong

2016-01-01

microRNAs (miRNAs) play critical roles in cancer development and progression. This study investigated the effects of miR-138-5p in human colorectal cancer (CRC) development. miR-138-5p was frequently downregulated in CRC tissues and was associated with advanced clinical stage, lymph node metastasis and poor overall survival. We found that miR-138-5p decreased expression of programmed cell death ligand 1 (PD-L1) through interaction with its PD-L1 3′ untranslated region. miR-138-5p also dramatically suppressed CRC cell growth in vitro and inhibited tumorigenesis in vivo. PD-L1 and miR-138-5p levels were inversely correlated in human CRC tumors, and miR-138-5p inhibited PD-L1 expression in tumor models. These results suggest that miR-138-5p is a tumor suppressor in CRC, and its effects are exerted at least partially through PD-L1 downregulation. Low miR-138-5p and high PD-L1 levels correlated with shorter overall CRC patient survival, indicating that miR-138-5p and PD-L1 may serve as CRC biomarkers for risk group assignment, optimal therapy selection and clinical outcome prediction. Targeting PD-L1, possibly by administering miR-138-5p mimics, might be a clinically effective anti-CRC therapeutic strategy. PMID:27248318

The tumor suppressor miR-138-5p targets PD-L1 in colorectal cancer.

PubMed

Zhao, Lian; Yu, Haibo; Yi, Shuijing; Peng, Xiaowei; Su, Peng; Xiao, Zhiming; Liu, Rui; Tang, Anliu; Li, Xiayu; Liu, Fen; Shen, Shourong

2016-07-19

microRNAs (miRNAs) play critical roles in cancer development and progression. This study investigated the effects of miR-138-5p in human colorectal cancer (CRC) development. miR-138-5p was frequently downregulated in CRC tissues and was associated with advanced clinical stage, lymph node metastasis and poor overall survival. We found that miR-138-5p decreased expression of programmed cell death ligand 1 (PD-L1) through interaction with its PD-L1 3' untranslated region. miR-138-5p also dramatically suppressed CRC cell growth in vitro and inhibited tumorigenesis in vivo. PD-L1 and miR-138-5p levels were inversely correlated in human CRC tumors, and miR-138-5p inhibited PD-L1 expression in tumor models. These results suggest that miR-138-5p is a tumor suppressor in CRC, and its effects are exerted at least partially through PD-L1 downregulation. Low miR-138-5p and high PD-L1 levels correlated with shorter overall CRC patient survival, indicating that miR-138-5p and PD-L1 may serve as CRC biomarkers for risk group assignment, optimal therapy selection and clinical outcome prediction. Targeting PD-L1, possibly by administering miR-138-5p mimics, might be a clinically effective anti-CRC therapeutic strategy.

Bio-Imaging of Colorectal Cancer Models Using Near Infrared Labeled Epidermal Growth Factor

PubMed Central

Cohen, Gadi; Lecht, Shimon; Arien-Zakay, Hadar; Ettinger, Keren; Amsalem, Orit; Oron-Herman, Mor; Yavin, Eylon; Prus, Diana; Benita, Simon; Nissan, Aviram; Lazarovici, Philip

2012-01-01

Novel strategies that target the epidermal growth factor receptor (EGFR) have led to the clinical development of monoclonal antibodies, which treat metastatic colorectal cancer (mCRC) but only subgroups of patients with increased wild type KRAS and EGFR gene copy, respond to these agents. Furthermore, resistance to EGFR blockade inevitably occurred, making future therapy difficult. Novel bio-imaging (BOI) methods may assist in quantization of EGFR in mCRC tissue thus complementing the immunohistochemistry methodology, in guiding the future treatment of these patients. The aim of the present study was to explore the usefulness of near infrared-labeled EGF (EGF-NIR) for bio-imaging of CRC using in vitro and in vivo orthotopic tumor CRC models and ex vivo human CRC tissues. We describe the preparation and characterization of EGF-NIR and investigate binding, using BOI of a panel of CRC cell culture models resembling heterogeneity of human CRC tissues. EGF-NIR was specifically and selectively bound by EGFR expressing CRC cells, the intensity of EGF-NIR signal to background ratio (SBR) reflected EGFR levels, dose-response and time course imaging experiments provided optimal conditions for quantization of EGFR levels by BOI. EGF-NIR imaging of mice with HT-29 orthotopic CRC tumor indicated that EGF-NIR is more slowly cleared from the tumor and the highest SBR between tumor and normal adjacent tissue was achieved two days post-injection. Furthermore, images of dissected tissues demonstrated accumulation of EGF-NIR in the tumor and liver. EGF-NIR specifically and strongly labeled EGFR positive human CRC tissues while adjacent CRC tissue and EGFR negative tissues expressed weak NIR signals. This study emphasizes the use of EGF-NIR for preclinical studies. Combined with other methods, EGF-NIR could provide an additional bio-imaging specific tool in the standardization of measurements of EGFR expression in CRC tissues. PMID:23144978

Bio-imaging of colorectal cancer models using near infrared labeled epidermal growth factor.

PubMed

Cohen, Gadi; Lecht, Shimon; Arien-Zakay, Hadar; Ettinger, Keren; Amsalem, Orit; Oron-Herman, Mor; Yavin, Eylon; Prus, Diana; Benita, Simon; Nissan, Aviram; Lazarovici, Philip

2012-01-01

Novel strategies that target the epidermal growth factor receptor (EGFR) have led to the clinical development of monoclonal antibodies, which treat metastatic colorectal cancer (mCRC) but only subgroups of patients with increased wild type KRAS and EGFR gene copy, respond to these agents. Furthermore, resistance to EGFR blockade inevitably occurred, making future therapy difficult. Novel bio-imaging (BOI) methods may assist in quantization of EGFR in mCRC tissue thus complementing the immunohistochemistry methodology, in guiding the future treatment of these patients. The aim of the present study was to explore the usefulness of near infrared-labeled EGF (EGF-NIR) for bio-imaging of CRC using in vitro and in vivo orthotopic tumor CRC models and ex vivo human CRC tissues. We describe the preparation and characterization of EGF-NIR and investigate binding, using BOI of a panel of CRC cell culture models resembling heterogeneity of human CRC tissues. EGF-NIR was specifically and selectively bound by EGFR expressing CRC cells, the intensity of EGF-NIR signal to background ratio (SBR) reflected EGFR levels, dose-response and time course imaging experiments provided optimal conditions for quantization of EGFR levels by BOI. EGF-NIR imaging of mice with HT-29 orthotopic CRC tumor indicated that EGF-NIR is more slowly cleared from the tumor and the highest SBR between tumor and normal adjacent tissue was achieved two days post-injection. Furthermore, images of dissected tissues demonstrated accumulation of EGF-NIR in the tumor and liver. EGF-NIR specifically and strongly labeled EGFR positive human CRC tissues while adjacent CRC tissue and EGFR negative tissues expressed weak NIR signals. This study emphasizes the use of EGF-NIR for preclinical studies. Combined with other methods, EGF-NIR could provide an additional bio-imaging specific tool in the standardization of measurements of EGFR expression in CRC tissues.

Inhibition of CD147 expression promotes chemosensitivity in HNSCC cells by deactivating MAPK/ERK signaling pathway.

PubMed

Ma, Chao; Wang, Jianqi; Fan, Longkun; Guo, Yanjun

2017-02-01

Head and neck squamous cell carcinoma (HNSCC) is one of the most common cancers in the world. CD147, a transmembrane glycoprotein, has been reported to be correlated with cancer progression, metastasis, and chemoresistance in various cancers. In this study, we aimed to investigate the mechanism of CD147 in regulating drug resistance in HNSCC cells. qRT-PCR were used to evaluated the expression of CD147 in 57 HNSCC tumorous tissues and 2 cell lines. Increased expression of CD147 was found in most HNSCC samples, and the expression level of CD147 was correlated with multidrug resistance. CD147 RNA silencing decreased the chemoresistance of HNSCC cells by deactivating MAPK/ERK signaling pathway. Further investigation revealed that either rescue expression of CD147 or treatment of MAPK/ERK activator phorbol 12-myristate 13-acetate (PMA) in CD147 knockdown CRC cell line attenuated the decreased chemoresistance in CD147 knockdown cells. Taken together, our results suggest that CD147 promotes chemoresistance by activating MAPK/ERK signaling pathway in HNSCC. Copyright © 2017. Published by Elsevier Inc.

In vivo and ex vivo cetuximab sensitivity assay using three-dimensional primary culture system to stratify KRAS mutant colorectal cancer

PubMed Central

Tashiro, Takahiro; Okuyama, Hiroaki; Endo, Hiroko; Kawada, Kenji; Ashida, Yasuko; Ohue, Masayuki; Sakai, Yoshiharu; Inoue, Masahiro

2017-01-01

In clinic, cetuximab, an anti-EGFR antibody, improves treatment outcomes in colorectal cancer (CRC). KRAS-mutant CRC is generally resistant to cetuximab, although difference of the sensitivity among KRAS-mutants has not been studied in detail. We previously developed the cancer tissue-originated spheroid (CTOS) method, a primary culture method for cancer cells. We applied CTOS method to investigate whether ex vivo cetuximab sensitivity assays reflect the difference in sensitivity in the xenografts. Firstly, in vivo cetuximab treatment was performed with xenografts derived from 10 CTOS lines (3 KRAS-wildtype and 7 KRAS mutants). All two CTOS lines which exhibited tumor regression were KRAS-wildtype, meanwhile all KRAS-mutant CTOS lines grew more than the initial size: were resistant to cetuximab according to the clinical evaluation criteria, although the sensitivity was quite diverse. We divided KRAS-mutants into two groups; partially responsive group in which cetuximab had a substantial growth inhibitory effect, and resistant group which exhibited no effect. The ex vivo signaling assay with EGF stimulation revealed that the partially responsive group, but not the resistant group, exhibited suppressed ERK phosphorylation ex vivo. Furthermore, two lines from the partially responsive group, but none of the lines in the resistant group, exhibited a combinatory effect of cetuximab and trametinib, a MEK inhibitor, ex vivo and in vivo. Taken together, the results indicate that ex vivo signaling assay reflects the difference in sensitivity in vivo and stratifies KRAS mutant CTOS lines by sensitivity. Therefore, coupling the in vivo and ex vivo assays with CTOS can be a useful platform for understanding the mechanism of diversity in drug sensitivity. PMID:28301591

Silencing of the Wnt transcription factor TCF4 sensitizes colorectal cancer cells to (chemo-) radiotherapy

PubMed Central

Kendziorra, Emil; Ahlborn, Kerstin; Spitzner, Melanie; Rave-Fränk, Margret; Emons, Georg; Gaedcke, Jochen; Kramer, Frank; Wolff, Hendrik A.; Becker, Heinz; Beissbarth, Tim; Ebner, Reinhard; Ghadimi, B.Michael; Pukrop, Tobias; Ried, Thomas; Grade, Marian

2011-01-01

A considerable percentage of rectal cancers are resistant to standard preoperative chemoradiotherapy. Because patients with a priori-resistant tumors do not benefit from multimodal treatment, understanding and overcoming this resistance remains of utmost clinical importance. We recently reported overexpression of the Wnt transcription factor TCF4, also known as TCF7L2, in rectal cancers that were resistant to 5-fluorouracil-based chemoradiotherapy. Because Wnt signaling has not been associated with treatment response, we aimed to investigate whether TCF4 mediates chemoradioresistance. RNA interference-mediated silencing of TCF4 was employed in three colorectal cancer (CRC) cell lines, and sensitivity to (chemo-) radiotherapy was assessed using a standard colony formation assay. Silencing of TCF4 caused a significant sensitization of CRC cells to clinically relevant doses of X-rays. This effect was restricted to tumor cells with high T cell factor (TCF) reporter activity, presumably in a β-catenin-independent manner. Radiosensitization was the consequence of (i) a transcriptional deregulation of Wnt/TCF4 target genes, (ii) a silencing-induced G2/M phase arrest, (iii) an impaired ability to adequately halt cell cycle progression after radiation and (iv) a compromised DNA double strand break repair as assessed by γH2AX staining. Taken together, our results indicate a novel mechanism through which the Wnt transcription factor TCF4 mediates chemoradioresistance. Moreover, they suggest that TCF4 is a promising molecular target to sensitize resistant tumor cells to (chemo-) radiotherapy. PMID:21983179

The addition of celecoxib improves the antitumor effect of cetuximab in colorectal cancer: role of EGFR-RAS-FOXM1-β-catenin signaling axis

PubMed Central

Valverde, Araceli; Peñarando, Jon; Cañas, Amanda; López-Sánchez, Laura M.; Conde, Francisco; Guil-Luna, Silvia; Hernández, Vanessa; Villar, Carlos; Morales-Estévez, Cristina; de la Haba-Rodríguez, Juan; Arand o, Enrique; Rodríguez-Ariza, Antonio

2017-01-01

Here we showed that the addition of the COX-2 inhibitor celecoxib improved the antitumor efficacy in colorectal cancer (CRC) of the monoclonal anti-EGFR antibody cetuximab. The addition of celecoxib augmented the efficacy of cetuximab to inhibit cell proliferation and to induce apoptosis in CRC cells. Moreover, the combination of celecoxib and cetuximab was more effective than either treatment alone in reducing the tumor volume in a mouse xenograft model. The combined treatment enhanced the inhibition of EGFR signaling and altered the subcellular distribution of β-catenin. Moreover, knockdown of FOXM1 showed that this transcription factor participates in this enhanced antitumoral response. Besides, the combined treatment decreased β-catenin/FOXM1 interaction and reduced the cancer stem cell subpopulation in CRC cells, as indicated their diminished capacity to form colonospheres. Notably, the inmunodetection of FOXM1 in the nuclei of tumor cells in human colorectal adenocarcinomas was significantly associated with response of patients to cetuximab. In summary, our study shows that the addition of celecoxib enhances the antitumor efficacy of cetuximab in CRC due to impairment of EGFR-RAS-FOXM1-β-catenin signaling axis. Results also support that FOXM1 could be a predictive marker of response of mCRC patients to cetuximab therapy. PMID:28423516

Protein content and functional characteristics of serum-purified exosomes from patients with colorectal cancer revealed by quantitative proteomics.

PubMed

Chen, Yanyu; Xie, Yong; Xu, Lai; Zhan, Shaohua; Xiao, Yi; Gao, Yanpan; Wu, Bin; Ge, Wei

2017-02-15

Tumor cells of colorectal cancer (CRC) release exosomes into the circulation. These exosomes can mediate communication between cells and affect various tumor-related processes in their target cells. We present a quantitative proteomics analysis of the exosomes purified from serum of patients with CRC and normal volunteers; data are available via ProteomeXchange with identifier PXD003875. We identified 918 proteins with an overlap of 725 Gene IDs in the Exocarta proteins list. Compared with the serum-purified exosomes (SPEs) of normal volunteers, we found 36 proteins upregulated and 22 proteins downregulated in the SPEs of CRC patients. Bioinformatics analysis revealed that upregulated proteins are involved in processes that modulate the pretumorigenic microenvironment for metastasis. In contrast, differentially expressed proteins (DEPs) that play critical roles in tumor growth and cell survival were principally downregulated. Our study demonstrates that SPEs of CRC patients play a pivotal role in promoting the tumor invasiveness, but have minimal influence on putative alterations in tumor survival or proliferation. According to bioinformatics analysis, we speculate that the protein contents of exosomes might be associated with whether they are involved in premetastatic niche establishment or growth and survival of metastatic tumor cells. This information will be helpful in elucidating the pathophysiological functions of tumor-derived exosomes, and aid in the development of CRC diagnostics and therapeutics. © 2016 UICC.

Treatment of Colorectal Peritoneal Carcinomatosis With Systemic Chemotherapy: A Pooled Analysis of North Central Cancer Treatment Group Phase III Trials N9741 and N9841

PubMed Central

Franko, Jan; Shi, Qian; Goldman, Charles D.; Pockaj, Barbara A.; Nelson, Garth D.; Goldberg, Richard M.; Pitot, Henry C.; Grothey, Axel; Alberts, Steven R.; Sargent, Daniel J.

2012-01-01

Purpose Symptoms and complications of metastatic colorectal cancer (mCRC) differ by metastatic sites. There is a paucity of prospective survival data for patients with peritoneal carcinomatosis colorectal cancer (pcCRC). We characterized outcomes of patients with pcCRC enrolled onto two prospective randomized trials of chemotherapy and contrasted that with other manifestations of mCRC (non-pcCRC). Methods A total of 2,095 patients enrolled onto two prospective randomized trials were evaluated for overall survival (OS) and progression-free survival (PFS). A Cox proportional hazard model was used to assess the adjusted associations. Results The characteristics of the pcCRC group (n = 364) were similar to those of the non-pcCRC patients in median age (63 v 61 years, P = .23), sex (57% males v 61%, P = .23), and performance status (Eastern Cooperative Oncology Group performance status 0 or 1 94% v 96%, P = .06), but differed in frequency of liver (63% v 82%, P < .001) and lung metastases (27% v 34%, P = .01). Median OS (12.7 v 17.6 months, hazard ratio [HR] = 1.3; 95% CI, 1.2 to 1.5; P < .001) and PFS (5.8 v 7.2 months, HR = 1.2; 95% CI, 1.1 to 1.3; P = .001) were shorter for pcCRC versus non-pcCRC. The unfavorable prognostic influence of pcCRC remained after adjusting for age, PS, liver metastases, and other factors (OS: HR = 1.3, P < .001; PFS: HR = 1.1, P = .02). Infusional fluorouracil, leucovorin, and oxaliplatin was superior to irinotecan, leucovorin, and fluorouracil as a first-line treatment among pcCRC (HR for OS = 0.62, P = .005) and non-pcCRC patients (HR = 0.66, P < .001). Conclusion pcCRC is associated with a significantly shorter OS and PFS as compared with other manifestations of mCRC. Future trials for mCRC should consider stratifying on the basis of pcCRC status. PMID:22162570

Increased expression of fatty acid synthase provides a survival advantage to colorectal cancer cells via upregulation of cellular respiration

PubMed Central

Zaytseva, Yekaterina Y.; Harris, Jennifer W.; Mitov, Mihail I.; Kim, Ji Tae; Butterfield, D. Allan; Lee, Eun Y.; Weiss, Heidi L.; Gao, Tianyan; Evers, B. Mark

2015-01-01

Fatty acid synthase (FASN), a lipogenic enzyme, is upregulated in colorectal cancer (CRC). Increased de novo lipid synthesis is thought to be a metabolic adaptation of cancer cells that promotes survival and metastasis; however, the mechanisms for this phenomenon are not fully understood. We show that FASN plays a role in regulation of energy homeostasis by enhancing cellular respiration in CRC. We demonstrate that endogenously synthesized lipids fuel fatty acid oxidation, particularly during metabolic stress, and maintain energy homeostasis. Increased FASN expression is associated with a decrease in activation of energy-sensing pathways and accumulation of lipid droplets in CRC cells and orthotopic CRCs. Immunohistochemical evaluation demonstrated increased expression of FASN and p62, a marker of autophagy inhibition, in primary CRCs and liver metastases compared to matched normal colonic mucosa. Our findings indicate that overexpression of FASN plays a crucial role in maintaining energy homeostasis in CRC via increased oxidation of endogenously synthesized lipids. Importantly, activation of fatty acid oxidation and consequent downregulation of stress-response signaling pathways may be key adaptation mechanisms that mediate the effects of FASN on cancer cell survival and metastasis, providing a strong rationale for targeting this pathway in advanced CRC. PMID:25970773

Increased expression of fatty acid synthase provides a survival advantage to colorectal cancer cells via upregulation of cellular respiration.

PubMed

Zaytseva, Yekaterina Y; Harris, Jennifer W; Mitov, Mihail I; Kim, Ji Tae; Butterfield, D Allan; Lee, Eun Y; Weiss, Heidi L; Gao, Tianyan; Evers, B Mark

2015-08-07

Fatty acid synthase (FASN), a lipogenic enzyme, is upregulated in colorectal cancer (CRC). Increased de novo lipid synthesis is thought to be a metabolic adaptation of cancer cells that promotes survival and metastasis; however, the mechanisms for this phenomenon are not fully understood. We show that FASN plays a role in regulation of energy homeostasis by enhancing cellular respiration in CRC. We demonstrate that endogenously synthesized lipids fuel fatty acid oxidation, particularly during metabolic stress, and maintain energy homeostasis. Increased FASN expression is associated with a decrease in activation of energy-sensing pathways and accumulation of lipid droplets in CRC cells and orthotopic CRCs. Immunohistochemical evaluation demonstrated increased expression of FASN and p62, a marker of autophagy inhibition, in primary CRCs and liver metastases compared to matched normal colonic mucosa. Our findings indicate that overexpression of FASN plays a crucial role in maintaining energy homeostasis in CRC via increased oxidation of endogenously synthesized lipids. Importantly, activation of fatty acid oxidation and consequent downregulation of stress-response signaling pathways may be key adaptation mechanisms that mediate the effects of FASN on cancer cell survival and metastasis, providing a strong rationale for targeting this pathway in advanced CRC.

A genetic variant in Rassf1a predicts outcome in mCRC patients treated with cetuximab plus chemotherapy: results from FIRE-3 and JACCRO 05 and 06 trials.

PubMed

Sebio, A; Stintzing, S; Heinemann, V; Sunakawa, Y; Zhang, W; Ichikawa, W; Tsuji, A; Takahashi, T; Parek, A; Yang, D; Cao, S; Ning, Y; Stremitzer, S; Matsusaka, S; Okazaki, S; Barzi, A; Berger, M D; Lenz, H-J

2018-01-01

The Hippo pathway is involved in colorectal cancer (CRC) development and progression. The Hippo regulator Rassf1a is also involved in the Ras signaling cascade. In this work, we tested single nucleotide polymorphisms within Hippo components and their association with outcome in CRC patients treated with cetuximab. Two cohorts treated with cetuximab plus chemotherapy were evaluated (198 RAS wild-type (WT) patients treated with first-line FOLFIRI plus Cetuximab within the FIRE-3 trial and 67 Ras WT patients treated either with first-line mFOLFOX6 or SOX plus Cetuximab). In these two populations, Rassf1a rs2236947 was associated with overall survival (OS), as patients with a CC genotype had significantly longer OS compared with those with CA or AA genotypes. This association was stronger in patients with left-side CRC (hazard ratio (HR): 1.79 (1.01-3.14); P=0.044 and HR: 2.83 (1.14-7.03); P=0.025, for Fire 3 and JACCRO cohorts, respectively). Rassf1a rs2236947 is a promising biomarker for patients treated with cetuximab plus chemotherapy.

The global regulator Crc plays a multifaceted role in modulation of type III secretion system in Pseudomonas aeruginosa

PubMed Central

Dong, Yi-Hu; Zhang, Xi-Fen; Zhang, Lian-Hui

2013-01-01

The opportunistic pathogen Pseudomonas aeruginosa utilizes type III secretion system (T3SS) to translocate effector proteins into eukaryotic host cells that subvert normal host cell functions to the benefit of the pathogen, and results in serious infections. T3SS in P. aeruginosa is controlled by a complex system of regulatory mechanisms and signaling pathways. In this study, we described that Crc, an RNA-binding protein, exerts a positive impact on T3SS in P. aeruginosa, as evidenced by promoter activity assays of several key T3SS genes, transcriptomics, RT-PCR, and immunoblotting in crc mutant. We further demonstrated that the regulatory function of Crc on the T3SS was mediated through the T3SS master regulator ExsA and linked to the Cbr/Crc signaling system. Expression profiling of the crc mutant revealed a downregulation of flagship T3SS genes as well as 16 other genes known to regulate T3SS gene expression in P. aeruginosa. On the basis of these data, we proposed that Crc may exert multifaceted control on the T3SS through various pathways, which may serve to fine-tune this virulence mechanism in response to environmental changes and nutrient sources. PMID:23292701

Conserved region C functions to regulate PD-1 expression and subsequent CD8 T cell memory1

PubMed Central

Bally, Alexander P. R.; Tang, Yan; Lee, Joshua T.; Barwick, Benjamin G.; Martinez, Ryan; Evavold, Brian D.; Boss, Jeremy M.

2016-01-01

Expression of programmed death 1 (PD-1) on CD8 T cells promotes T cell exhaustion during chronic antigen exposure. During acute infections, PD-1 is transiently expressed and has the potential to modulate CD8 T cell memory formation. Conserved Region C (CR-C), a promoter proximal cis-regulatory element that is critical to PD-1 expression in vitro, responds to NFATc1, FoxO1, and/or NF-κB signaling pathways. Here, a CR-C knockout mouse (CRC−) was established to determine its role on PD-1 expression and corresponding effects on T cell function in vivo. Deletion of CR-C decreased PD-1 expression on CD4 T cells and antigen-specific CD8 T cells during acute and chronic lymphocytic choriomeningitis virus (LCMV) challenges, but did not affect the ability to clear an infection. Following acute LCMV infection, memory CD8 T cells in the CRC− mouse were formed in greater numbers, were more functional, and were more effective at responding to a melanoma tumor than wild-type memory cells. These data implicate a critical role for CR-C in governing PD-1 expression, and a subsequent role in guiding CD8 T cell differentiation. The data suggest the possibility that titrating PD-1 expression during CD8 T cell activation could have important ramifications in vaccine development and clinical care. PMID:27895178

CP-31398 prevents the growth of p53-mutated colorectal cancer cells in vitro and in vivo.

PubMed

He, Xingxing; Kong, Xinjuan; Yan, Junwei; Yan, Jingjun; Zhang, Yunan; Wu, Qian; Chang, Ying; Shang, Haitao; Dou, Qian; Song, Yuhu; Liu, Fang

2015-03-01

Rescuing the function of mutant p53 protein is an attractive cancer therapeutic strategy. Small molecule CP-31398 was shown to restore mutant p53 tumor suppressor functions in cancer cells. Here, we determined the effects of CP-31398 on the growth of p53-mutated colorectal cancer (CRC) cells in vitro and in vivo. CRC cells which carry p53 mutation in codon 273 were treated with CP-31398 and the control, and the effects of CP-31398 on cell cycle, cell apoptosis, and proliferation were determined. The expression of p53-responsive downstream genes was evaluated by quantitative reverse transcriptase PCR (RT-PCR) and Western blot. CP-31398 was administrated into xenograft tumors created by the inoculation of HT-29 cells, and then the effect of CP-31398 on the growth of xenograft tumors was examined. CP-31398 induced p53 downstream target molecules in cultured HT-29 cells, which resulted in the inhibition of CRC cell growth assessed by the determination of cell cycle, apoptosis, and cell proliferation. In xenograft tumors, CP-31398 modulated the expression of Bax, Bcl-2, caspase 3, cyclin D, and Mdm2 and then blocked the growth of xenograft tumors. CP-31398 would be developed as a therapeutic candidate for p53-mutated CRC due to the restoration of mutant p53 tumor suppressor functions.

Vicenin-2 inhibits Wnt/β-catenin signaling and induces apoptosis in HT-29 human colon cancer cell line.

PubMed

Yang, Dong; Zhang, Xiling; Zhang, Wencun; Rengarajan, Thamaraiselvan

2018-01-01

Colorectal cancer (CRC) is among highest prevailing cancers in the whole world, especially in western countries. For a diverse of reasons, patients prefer naturally occurring dietary substances over synthetic agents to prevent cancer. Vicenin-2 is largely available in a medicinal plant Ocimum sanctum and is an apigenin form, 6,8-di-C-glucoside, which has been reported to have a range of pharmacological values which includes antioxidant, hepatoprotective, anti-inflammatory and anti-cancer. This study was aimed to analyze the anti-proliferative effect of Vicenin-2 on human colon cancer cells via the Wnt/β-catenin signaling inhibition. MTT assay was used to assess the cell viability at different concentrations and time point. Vicenin-2 at a concentration of 50 µM (IC 50 ) decreased the phosphorylated (inactive) glycogen synthase kinase-3β, cyclin D1, and non-p-β-catenin expressions in HT-29 cells, which were evidenced through western blot analysis. Further, Vincenin-2 reduced the T-cell factor (TCF) / Leukocyte erythroid factor (LEF) reporter activity in HT-29 cells. Vicenin-2 also promoted substantial cell cycle arrest at the G 2 M phase of HT-29 cells, as well induced apoptosis in HT-29 cells, as revealed through flow cytometric analysis. Furthermore, immunoblot analysis showed that Vicenin-2 treatment enhanced the expression of Cytochrome C, Bax and caspase-3 whereas suppressed the Bcl-2 expression. Together, these results revealed that Vicenin-2 can act as a potent inhibitor of HT-29 cell proliferation and can be used as an agent against CRC.

PAC exhibits potent anti-colon cancer properties through targeting cyclin D1 and suppressing epithelial-to-mesenchymal transition.

PubMed

Al-Qasem, Abeer; Al-Howail, Huda A; Al-Swailem, Mashael; Al-Mazrou, Amer; Al-Otaibi, Basem; Al-Jammaz, Ibrahim; Al-Khalaf, Huda H; Aboussekhra, Abdelilah

2016-03-01

Colorectal cancer (CRC) is a major cause of cancer morbidity and mortality worldwide. Although response rates and overall survival have been improved in recent years, resistance to multiple drug combinations is inevitable. Therefore, the development of more efficient drugs, with fewer side effects is urgently needed. To this end, we have investigated in the present report the effect of PAC, a novel cucumin analogue, on CRC cells both in vitro and in vivo. We have shown that PAC induces apoptosis, mainly via the internal mitochondrial route, and inhibits cell proliferation through delaying the cell cycle at G2/M phase. Interestingly, the pro-apoptotic effect was mediated through STAT3-dependent down-regulation of cyclin D1 and its downstream target survivin. Indeed, change in the expression level of cyclin D1 modulated the expression of survivin and the response of CRC cells to PAC. Furthermore, using the ChIP assay, we have shown PAC-dependent reduction in the binding of STAT3 to the cyclin D1 promoter in vivo. Additionally, PAC suppressed the epithelial-to-mesenchymal process through down-regulating the mesenchymal markers (N-cadherin, vimentin and Twist1) and inhibiting the invasion/migration abilities of the CRC cells via repressing the pro-migration/invasion protein kinases AKT and ERK1/2. In addition, PAC inhibited tumor growth and repressed the JAK2/STAT3, AKT/mTOR and MEK/ERK pathways as well as their common downstream effectors cyclin D1 and survivin in humanized CRC xenografts. Collectively, these results indicate that PAC has potent anti-CRC effects, and therefore could constitute an effective alternative chemotherapeutic agent, which may consolidate the adjuvant treatment of colon cancer. © 2015 Wiley Periodicals, Inc.

Gut microbiota modulate T cell trafficking into human colorectal cancer.

PubMed

Cremonesi, Eleonora; Governa, Valeria; Garzon, Jesus Francisco Glaus; Mele, Valentina; Amicarella, Francesca; Muraro, Manuele Giuseppe; Trella, Emanuele; Galati-Fournier, Virginie; Oertli, Daniel; Däster, Silvio Raffael; Droeser, Raoul A; Weixler, Benjamin; Bolli, Martin; Rosso, Raffaele; Nitsche, Ulrich; Khanna, Nina; Egli, Adrian; Keck, Simone; Slotta-Huspenina, Julia; Terracciano, Luigi M; Zajac, Paul; Spagnoli, Giulio Cesare; Eppenberger-Castori, Serenella; Janssen, Klaus-Peter; Borsig, Lubor; Iezzi, Giandomenica

2018-02-06

Tumour-infiltrating lymphocytes (TILs) favour survival in human colorectal cancer (CRC). Chemotactic factors underlying their recruitment remain undefined. We investigated chemokines attracting T cells into human CRCs, their cellular sources and microenvironmental triggers. Expression of genes encoding immune cell markers, chemokines and bacterial 16S ribosomal RNA (16SrRNA) was assessed by quantitative reverse transcription-PCR in fresh CRC samples and corresponding tumour-free tissues. Chemokine receptor expression on TILs was evaluated by flow cytometry on cell suspensions from digested tissues. Chemokine production by CRC cells was evaluated in vitro and in vivo, on generation of intraperitoneal or intracecal tumour xenografts in immune-deficient mice. T cell trafficking was assessed on adoptive transfer of human TILs into tumour-bearing mice. Gut flora composition was analysed by 16SrRNA sequencing. CRC infiltration by distinct T cell subsets was associated with defined chemokine gene signatures, including CCL5, CXCL9 and CXCL10 for cytotoxic T lymphocytes and T-helper (Th)1 cells; CCL17, CCL22 and CXCL12 for Th1 and regulatory T cells; CXCL13 for follicular Th cells; and CCL20 and CCL17 for interleukin (IL)-17-producing Th cells. These chemokines were expressed by tumour cells on exposure to gut bacteria in vitro and in vivo. Their expression was significantly higher in intracecal than in intraperitoneal xenografts and was dramatically reduced by antibiotic treatment of tumour-bearing mice. In clinical samples, abundance of defined bacteria correlated with high chemokine expression, enhanced T cell infiltration and improved survival. Gut microbiota stimulate chemokine production by CRC cells, thus favouring recruitment of beneficial T cells into tumour tissues. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Focal adhesion molecule Kindlin-1 mediates activation of TGF-β signaling by interacting with TGF-βRI, SARA and Smad3 in colorectal cancer cells

PubMed Central

Wang, Yunling; Yang, Mingzi; Gao, Jianchao; Wei, Xiaofan; Fang, Weigang; Zhan, Jun; Zhang, Hongquan

2016-01-01

Kindlin-1, an integrin-interacting protein, has been implicated in TGF-β/Smad3 signaling. However, the molecular mechanism underlying Kindlin-1 regulation of TGF-β/Smad3 signaling remains elusive. Here, we reported that Kindlin-1 is an important mediator of TGF-β/Smad3 signaling by showing that Kindlin-1 physically interacts with TGF-β receptor I (TβRI), Smad anchor for receptor activation (SARA) and Smad3. Kindlin-1 is required for the interaction of Smad3 with TβRI, Smad3 phosphorylation, nuclear translocation, and finally the activation of TGF-β/Smad3 signaling pathway. Functionally, Kindlin-1 promoted colorectal cancer (CRC) cell proliferation in vitro and tumor growth in vivo, and was also required for CRC cell migration and invasion via an epithelial to mesenchymal transition. Kindlin-1 was found to be increased with the CRC progression from stages I to IV. Importantly, raised expression level of Kindlin-1 correlates with poor outcome in CRC patients. Taken together, we demonstrated that Kindlin-1 promotes CRC progression by recruiting SARA and Smad3 to TβRI and thereby activates TGF-β/Smad3 signaling. Thus, Kindlin-1 is a novel regulator of TGF-β/Smad3 signaling and may also be a potential target for CRC therapeutics. PMID:27776350

CD38+ M-MDSC expansion characterizes a subset of advanced colorectal cancer patients.

PubMed

Karakasheva, Tatiana A; Dominguez, George A; Hashimoto, Ayumi; Lin, Eric W; Chiu, Christopher; Sasser, Kate; Lee, Jae W; Beatty, Gregory L; Gabrilovich, Dmitry I; Rustgi, Anil K

2018-03-22

Myeloid-derived suppressor cells (MDSCs) are a population of immature immune cells with several protumorigenic functions. CD38 is a transmembrane receptor-ectoenzyme expressed by MDSCs in murine models of esophageal cancer. We hypothesized that CD38 could be expressed on MDSCs in human colorectal cancer (CRC), which might allow for a new perspective on therapeutic targeting of human MDSCs with anti-CD38 monoclonal antibodies in this cancer. Blood samples were collected from 41 CRC patients and 8 healthy donors, followed by peripheral blood mononuclear cell (PBMC) separation. Polymorphonuclear (PMN-) and monocytic (M-) MDSCs and CD38 expression levels were quantified by flow cytometry. The immunosuppressive capacity of M-MDSCs from 10 CRC patients was validated in a mixed lymphocyte reaction (MLR) assay. A significant expansion of CD38+ M-MDSCs and a trend of expansion of CD38+ PMN-MDSCs (accompanied by a trend of increased CD38 expression on both M- and PMN-MDSCs) were observed in PBMCs of CRC patients when compared with healthy donors. The CD38+ M-MDSCs from CRC patients were found to be immunosuppressive when compared with mature monocytes. CD38+ M- and PMN-MDSC frequencies were significantly higher in CRC patients who previously received treatment when compared with treatment-naive patients. This study provides a rationale for an attempt to target M-MDSCs with an anti-CD38 monoclonal antibody in metastatic CRC patients. NCI P01-CA14305603, the American Cancer Society, Scott and Suzi Lustgarten Family Colon Cancer Research Fund, Hansen Foundation, and Janssen Research and Development.

Marine ω-3 Polyunsaturated Fatty Acid Intake and Risk of Colorectal Cancer Characterized by Tumor-Infiltrating T Cells.

PubMed

Song, Mingyang; Nishihara, Reiko; Cao, Yin; Chun, Eunyoung; Qian, Zhi Rong; Mima, Kosuke; Inamura, Kentaro; Masugi, Yohei; Nowak, Jonathan A; Nosho, Katsuhiko; Wu, Kana; Wang, Molin; Giovannucci, Edward; Garrett, Wendy S; Fuchs, Charles S; Ogino, Shuji; Chan, Andrew T

2016-09-01

Marine ω-3 polyunsaturated fatty acids (PUFAs), including eicosapentaenoic acid, docosahexaenoic acid, and docosapentaenoic acid, possess potent immunomodulatory activity and may protect against cancer development. However, evidence relating marine ω-3 PUFAs to colorectal cancer (CRC) risk remains inconclusive. To test the hypothesis that marine ω-3 PUFA intake may be associated with lower risk of CRC subsets characterized by immune infiltrate. This prospective cohort study was conducted among participants in the Nurses' Health Study (1984-2010) and Health Professionals Follow-up Study (1986-2010). Intake of marine ω-3 PUFAs. Incidence of CRC characterized by CD3+, CD8+, CD45RO (PTPRC)+, or FOXP3+ T-cell densities in tumor tissue, measured by immunohistochemical and computer-assisted image analysis. Among 173 229 predominantly white participants, 125 172 with 2 895 704 person-years of follow-up provided data about marine ω-3 PUFA intake every 4 years through a validated food frequency questionnaire and followed up for incident CRC evaluation. Of 2504 CRC cases, we documented 614 (252 men, 362 women) from which we could assess T-cell infiltration in the tumor microenvironment. The inverse association of marine ω-3 PUFAs intake with CRC risk differed according to FOXP3+ T-cell infiltration: compared with intake of less than 0.15 g/d of marine ω-3 PUFAs, intake of at least 0.35 g/d was associated with a multivariable hazard ratio (HR) of 0.57 (95% CI, 0.40-0.81; P < .001 for trend) for FOXP3+ T-cell-high tumors. In contrast, the HR for FOXP3+ T-cell-low tumors was 1.14 (95% CI, 0.8-1.60) (P = .77 for trend; P = .01 for heterogeneity). No statistically significant differential association was found for high-density tumors (compared with low-density tumors) according to CD3+, CD8+, or CD45RO+ cell density (P ≥ .34 for heterogeneity for all comparisons). In functional assays, the suppressive activity of regulatory T cells was approximately 2-fold lower (T-effector-cell proliferation, ≥64% vs 38%) when preincubated with docosahexaenoic acid at 50μM, 100μM, and 200μM concentrations than without docosahexaenoic acid (P < .001 for all comparisons). High marine ω-3 PUFA intake was associated with lower risk of CRC with high-level, but not low-level, FOXP3+ T-cell density, suggesting a potential role of ω-3 PUFAs in cancer immunoprevention through modulation of regulatory T cells.

Meta-Analysis of Oxaliplatin-Based Chemotherapy Combined With Traditional Medicines for Colorectal Cancer

PubMed Central

Chen, Menghua; May, Brian H.; Zhou, Iris W.; Xue, Charlie C. L.; Zhang, Anthony L.

2015-01-01

This meta-analysis evaluates the clinical evidence for the addition of traditional medicines (TMs) to oxaliplatin-based regimens for colorectal cancer (CRC) in terms of tumor response rate (TRR). Eight electronic databases were searched for randomized controlled trials of oxaliplatin-based chemotherapy combined with TMs compared to the same oxaliplatin-based regimen. Data on TRR from 42 randomized controlled trials were analyzed using Review Manager 5.1. Studies were conducted in China or Japan. Publication bias was not evident. The meta-analyses suggest that the combination of the TMs with oxaliplatin-based regimens increased TRR in the palliative treatment of CRC (risk ratio [RR] 1.31 [1.20-1.42], I2 = 0%). Benefits were evident for both injection products (RR 1.36 [1.18-1.57], I2 = 0%) and orally administered TMs (RR 1.27 [1.15-1.41], I2 = 0%). Further sensitivity analysis of specific plant-based TMs found that Paeonia, Curcuma, and Sophora produced consistently higher contributions to the RR results. Compounds in each of these TMs have shown growth-inhibitory effects in CRC cell-line studies. Specific combinations of TMs appeared to produce higher contributions to TRR than the TMs individually. Notable among these was the combination of Hedyotis, Astragalus, and Scutellaria. PMID:26254190

Low Tumor Infiltrating Mast Cell Density Confers Prognostic Benefit and Reflects Immunoactivation in Colorectal Cancer.

PubMed

Mao, Yihao; Feng, Qingyang; Zheng, Peng; Yang, Liangliang; Zhu, Dexiang; Chang, Wenju; Ji, Meiling; He, Guodong; Xu, Jianmin

2018-06-06

The role of mast cells (MCs) in colorectal cancer (CRC) progression was controversial. Thus, this study was designed to evaluate the prognostic value of MCs as well as their correlation with immune microenvironment. A retrospective cohort of CRC patients of stage I-IV was enrolled in this study. 854 consecutive patients were divided into training set (427 patients) and validation set (427 patients) randomly. The findings were further validated in a GEO cohort, GSE39582 (556 patients). The mast cell density (MCD) was measured by immunohistochemical staining of tryptase or by CIBERSORT algorithm. Low MCD predicted prolonged overall survival (OS) in training and validation set. Moreover, MCD was identified as an independent prognostic indicator in both sets. Better stratification for CRC prognosis can be achieved by building a MCD based nomogram. The prognostic role of MCD was further validated in GSE39582. In addition, MCD predicted improved survival in stage II and III CRC patients receiving adjuvant chemotherapy (ACT). Multiple immune pathways were enriched in low MCD group while cytokines/chemokines promoting anti-tumor immunity were highly expressed in such group. Furthermore, MCD was negatively correlated with CD8+ T cells infiltration. In conclusion, MCD was identified as an independent prognostic factor, as well as a potential biomarker for ACT benefit in stage II and III CRC. Better stratification of CRC prognosis could be achieved by building a MCD based nomogram. Moreover, immunoactivation in low MCD tumors may contributed to improved prognosis. This article is protected by copyright. All rights reserved. © 2018 UICC.

pH-sensitive nanomicelles for controlled and efficient drug delivery to human colorectal carcinoma LoVo cells.

PubMed

Feng, Shi-Ting; Li, Jingguo; Luo, Yanji; Yin, Tinghui; Cai, Huasong; Wang, Yong; Dong, Zhi; Shuai, Xintao; Li, Zi-Ping

2014-01-01

The triblock copolymers PEG-P(Asp-DIP)-P(Lys-Ca) (PEALCa) of polyethylene glycol (PEG), poly(N-(N',N'-diisopropylaminoethyl) aspartamide) (P(Asp-DIP)), and poly (lysine-cholic acid) (P(Lys-Ca)) were synthesized as a pH-sensitive drug delivery system. In neutral aqueous environment such as physiological environment, PEALCa can self-assemble into stable vesicles with a size around 50-60 nm, avoid uptake by the reticuloendothelial system (RES), and encase the drug in the core. However, the PEALCa micelles disassemble and release drug rapidly in acidic environment that resembles lysosomal compartments. The anticancer drug Paclitaxel (PTX) and hydrophilic superparamagnetic iron oxide (SPIO) were encapsulated inside the core of the PEALCa micelles and used for potential cancer therapy. Drug release study revealed that PTX in the micelles was released faster at pH 5.0 than at pH 7.4. Cell culture studies showed that the PTX-SPIO-PEALCa micelle was effectively internalized by human colon carcinoma cell line (LoVo cells), and PTX could be embedded inside lysosomal compartments. Moreover, the human colorectal carcinoma (CRC) LoVo cells delivery effect was verified in vivo by magnetic resonance imaging (MRI) and histology analysis. Consequently effective suppression of CRC LoVo cell growth was evaluated. These results indicated that the PTX-SPION-loaded pH-sensitive micelles were a promising MRI-visible drug release system for colorectal cancer therapy.

Clinical value of the preoperative neutrophil-to-lymphocyte ratio and red blood cell distribution width in patients with colorectal carcinoma.

PubMed

Han, Fuyan; Shang, Xuming; Wan, Furong; Liu, Zhanfeng; Tian, Wenjun; Wang, Dan; Liu, Yiqing; Wang, Yong; Zhang, Bingchang; Ju, Ying

2018-03-01

The aim of the present study was to investigate the clinical value of the preoperative neutrophil-to-lymphocyte ratio (NLR) and red blood cell distribution width (RDW) in the peripheral blood of colorectal carcinoma (CRC) patients. Clinical data obtained from 240 patients with CRC undergoing radical surgical resection in Shandong Provincial Hospital Affiliated to Shandong University (Jinan, Shandong, China) between January 2011 and April 2015 were retrospectively analyzed. Data were also collected from 110 patients with colon polyps and 48 healthy volunteers to serve as controls for comparative analysis. The clinicopathological characteristics of the patients in the low and high NLR and RDW groups were compared. The NLR and RDW values were compared prior to and following surgery. Kaplan-Meier analyses and Cox regression modeling were performed to predict overall survival (OS) and disease-free survival (DFS). The NLR and RDW levels in the CRC patients were markedly higher than those in the colon polyp patients and the healthy controls. The optimum NLR and RDW cutoff points for CRC were 2.06 and 13.45%, respectively. Significant differences were detected in tumor location, diameter, degree of differentiation, tumor depth, carcinoembryonic antigen and carbohydrate antigen 199 when comparing the high and low NLR groups (P<0.05). A high RDW was significantly associated with distant metastasis and older age in CRC patients. No significant difference was detected in the NLR and RDW levels of CRC patients prior to and following surgery (P>0.05). CRC patients with an increased RDW had significantly worse OS and DFS rates, particularly those with metastatic CRC (P<0.05). Patients with a high NLR exhibited a reduced DFS time in CRC (P=0.053), although this difference was not significant, and a significantly worse DFS time in metastatic CRC (P=0.047). In conclusion, it is convenient to use preoperative NLR and RDW to predict prognosis following surgery for patients with CRC.

DNA methylome profiling identifies novel methylated genes in African American patients with colorectal neoplasia.

PubMed

Ashktorab, Hassan; Daremipouran, M; Goel, Ajay; Varma, Sudhir; Leavitt, R; Sun, Xueguang; Brim, Hassan

2014-04-01

The identification of genes that are differentially methylated in colorectal cancer (CRC) has potential value for both diagnostic and therapeutic interventions specifically in high-risk populations such as African Americans (AAs). However, DNA methylation patterns in CRC, especially in AAs, have not been systematically explored and remain poorly understood. Here, we performed DNA methylome profiling to identify the methylation status of CpG islands within candidate genes involved in critical pathways important in the initiation and development of CRC. We used reduced representation bisulfite sequencing (RRBS) in colorectal cancer and adenoma tissues that were compared with DNA methylome from a healthy AA subject's colon tissue and peripheral blood DNA. The identified methylation markers were validated in fresh frozen CRC tissues and corresponding normal tissues from AA patients diagnosed with CRC at Howard University Hospital. We identified and validated the methylation status of 355 CpG sites located within 16 gene promoter regions associated with CpG islands. Fifty CpG sites located within CpG islands-in genes ATXN7L1 (2), BMP3 (7), EID3 (15), GAS7 (1), GPR75 (24), and TNFAIP2 (1)-were significantly hypermethylated in tumor vs. normal tissues (P<0.05). The methylation status of BMP3, EID3, GAS7, and GPR75 was confirmed in an independent, validation cohort. Ingenuity pathway analysis mapped three of these markers (GAS7, BMP3 and GPR) in the insulin and TGF-β1 network-the two key pathways in CRC. In addition to hypermethylated genes, our analysis also revealed that LINE-1 repeat elements were progressively hypomethylated in the normal-adenoma-cancer sequence. We conclude that DNA methylome profiling based on RRBS is an effective method for screening aberrantly methylated genes in CRC. While previous studies focused on the limited identification of hypermethylated genes, ours is the first study to systematically and comprehensively identify novel hypermethylated genes, as well as hypomethylated LINE-1 sequences, which may serve as potential biomarkers for CRC in African Americans. Our discovered biomarkers were intimately linked to the insulin/TGF-B1 pathway, further strengthening the association of diabetic disorders with colon oncogenic transformation.

Digital detection of multiple minority mutants and expression levels of multiple colorectal cancer-related genes using digital-PCR coupled with bead-array.

PubMed

Huang, Huan; Li, Shuo; Sun, Lizhou; Zhou, Guohua

2015-01-01

To simultaneously analyze mutations and expression levels of multiple genes on one detection platform, we proposed a method termed "multiplex ligation-dependent probe amplification-digital amplification coupled with hydrogel bead-array" (MLPA-DABA) and applied it to diagnose colorectal cancer (CRC). CRC cells and tissues were sampled to extract nucleic acid, perform MLPA with sequence-tagged probes, perform digital emulsion polymerase chain reaction (PCR), and produce a hydrogel bead-array to immobilize beads and form a single bead layer on the array. After hybridization with fluorescent probes, the number of colored beads, which reflects the abundance of expressed genes and the mutation rate, was counted for diagnosis. Only red or green beads occurred on the chips in the mixed samples, indicating the success of single-molecule PCR. When a one-source sample was analyzed using mixed MLPA probes, beads of only one color occurred, suggesting the high specificity of the method in analyzing CRC mutation and gene expression. In gene expression analysis of a CRC tissue from one CRC patient, the mutant percentage was 3.1%, and the expression levels of CRC-related genes were much higher than those of normal tissue. The highly sensitive MLPA-DABA succeeds in the relative quantification of mutations and gene expressions of exfoliated cells in stool samples of CRC patients on the same chip platform. MLPA-DABA coupled with hydrogel bead-array is a promising method in the non-invasive diagnosis of CRC.

Parecoxib: an enhancer of radiation therapy for colorectal cancer.

PubMed

Xiong, Wei; Li, Wen-Hui; Jiang, Yong-Xin; Liu, Shan; Ai, Yi-Qin; Liu, Rong; Chang, Li; Zhang, Ming; Wang, Xiao-Li; Bai, Han; Wang, Hong; Zheng, Rui; Tan, Jing

2015-01-01

To study the effect of parecoxib, a novel cyclooxygenase-2 selective inhibitor, on the radiation response of colorectal cancer (CRC) cells and its underlying mechanisms. Both in vitro colony formation and apoptosis assays as well as in vivo mouse xenograft experiments were used to explore the radiosensitizing effects of parecoxib in human HCT116 and HT29 CRC cells. Parecoxib sensitized CRC cells to radiation in vitro with a sensitivity enhancement ratio of 1.32 for HCT116 cells and 1.15 for HT29 cells at a surviving fraction of 0.37. This effect was partially attributable to enhanced apoptosis induction by parecoxib combined with radiation, as illustrated using an in vitro apoptosis assays. Parecoxib augmented the tumor response of HCT116 xenografts to radiation, achieving growth delay more than 20 days and an enhancement factor of 1.53. In accordance with the in vitro results, parecoxib combined with radiation resulted in less proliferation and more apoptosis in tumors than radiation alone. Radiation monotherapy decreased microvessel density (MVD) and microvessel intensity (MVI), but increased the hypoxia level in xenografts. Parecoxib did not affect MVD, but it increased MVI and attenuated hypoxia. Parecoxib can effectively enhance radiation sensitivity in CRC cells through direct effects on tumor cells and indirect effects on tumor vasculature.

MicroRNA-93 inhibits tumor growth and early relapse of human colorectal cancer by affecting genes involved in the cell cycle.

PubMed

Yang, I-Ping; Tsai, Hsiang-Lin; Hou, Ming-Feng; Chen, Ku-Chung; Tsai, Pei-Chien; Huang, Szu-Wei; Chou, Wen-Wen; Wang, Jaw-Yuan; Juo, Suh-Hang Hank

2012-08-01

Colorectal cancer (CRC) is associated with high recurrence and mortality. Because deregulation of microRNAs is associated with CRC development and recurrence, the expression levels of microRNAs can be a simple and reliable biomarker to detect postoperative early relapse, thereby helping physicians to treat high-risk patients more efficiently. We used microRNA arrays and observed that microRNA-93 had substantially different expression levels in early (recurrence within 12 months after surgery) and non-early relapse CRC patients. The replication study, which included 35 early relapse and 42 non-early relapse subjects, further confirmed overexpression of microRNA-93 in non-early relapse samples. The in vitro and in vivo effects of microRNA-93 were investigated by examining cell proliferation, migration and invasion, as well as cell cycles, target-gene expression and xenograft in null mice. Cellular studies showed that the overexpression of microRNA-93 inhibited colon cancer cell proliferation and migration but not invasion. The cell cycle studies also revealed that microRNA-93 caused an accumulation of the G2 population. However, microRNA-93 could not induce cell apoptosis or necrosis. Functional studies showed that microRNA-93 could suppress CCNB1 protein expression leading to cell cycle arrest in the G2 phase. Moreover, microRNA-93 repressed expression of ERBB2, p21 and VEGF, all of which are involved in cell proliferation. MicroRNA-93 also suppressed tumor growth in null mice. This study showed that microRNA-93 can inhibit tumorigenesis and reduce the recurrence of CRC; these findings may have potential clinical applications for predicting the recurrence of CRC.

Chemotherapy usage patterns in a US-wide cohort of patients with metastatic colorectal cancer.

PubMed

Abrams, Thomas A; Meyer, Gary; Schrag, Deborah; Meyerhardt, Jeffrey A; Moloney, Julie; Fuchs, Charles S

2014-02-01

Since the introduction of biologic therapies for the treatment of metastatic colorectal cancer (mCRC), few studies have examined patterns of care or predictors of specific treatment approaches. We assessed 4877 mCRC patients who received chemotherapy between January 2004 and March 2011 at academic, private, and community-based oncology practices subscribing to a US-wide chemotherapy order entry (system capturing disease, patient, provider, and treatment data. Multivariable analyses of these prospectively recorded characteristics were used to identify independent predictors of specific therapeutic choices. All statistical tests were two-sided. Throughout the study period, fluoropyrimidine/oxaliplatin combination was the most commonly used first-line chemotherapy regimen, representing 71% of first-line therapy by 2007. First-line bevacizumab use averaged 51%, peaking at 55% in 2006. Of those who received first-line bevacizumab, 34% continued to receive bevacizumab in the second-line. Only 26% of patients in our cohort ever received an anti-EGFR monoclonal antibody (cetuximab = 22%; panitumumab = 6%) at some point in their treatment course. Patients treated at academic centers, with longer duration of first-line therapy, and at sites in the western United States were statistically more likely to receive an anti-EGFR antibody. Anti-EGFR antibody use fell by 18% after the US Food and Drug Administration limited its use to patients with KRAS wild-type tumors in June 2009. Analysis of this US-wide mCRC cohort demonstrates that bevacizumab has been more consistently integrated into treatment regimens than anti-EGFR antibody therapies, particularly in first-line therapy. However, treatment choices vary substantially according to specific patient, practice, and provider characteristics.

The Association Between Molecular Markers in Colorectal Sessile Serrated Polyps and Colorectal Cancer Risk

DTIC Science & Technology

2017-08-01

mouse and human colon epithelium; Aim 2.) Perform genome editing using CRISPR /Cas9 on immortalized human colon epithelial cells to introduce CRC...relevant gene mutations; Aim 3.) Use CRISPR /Cas9 genome editing in colon organoid cultures to introduce CRC relevant gene mutations into primary colon cells

Colitis-associated colon cancer: Is it in your genes?

PubMed Central

Van Der Kraak, Lauren; Gros, Philippe; Beauchemin, Nicole

2015-01-01

Colitis-associated colorectal cancer (CA-CRC) is the cause of death in 10%-15% of inflammatory bowel disease (IBD) patients. CA-CRC results from the accumulation of mutations in intestinal epithelial cells and progresses through a well-characterized inflammation to dysplasia to carcinoma sequence. Quantitative estimates of overall CA-CRC risks are highly variable ranging from 2% to 40% depending on IBD severity, duration and location, with IBD duration being the most significant risk factor associated with CA-CRC development. Recently, studies have identified IBD patients with similar patterns of colonic inflammation, but that differ with respect to CA-CRC development, suggesting a role for additional non-inflammatory risk factors in CA-CRC development. One suggestion is that select IBD patients carry polymorphisms in various low penetrance disease susceptibility genes, which pre-dispose them to CA-CRC development, although these loci have proven difficult to identify in human genome-wide association studies. Mouse models of CA-CRC have provided a viable alternative for the discovery, validation and study of individual genes in CA-CRC pathology. In this review, we summarize the current CA-CRC literature with a strong focus on genetic pre-disposition and highlight an emerging role for mouse models in the search for CA-CRC risk alleles. PMID:26556996

PIK3CA and PIK3CB silencing by RNAi reverse MDR and inhibit tumorigenic properties in human colorectal carcinoma.

PubMed

Wu, Shuhua; Wen, Feifei; Li, Yangyang; Gao, Xiangqian; He, Shuang; Liu, Mengyao; Zhang, Xiangzhi; Tian, Dong

2016-07-01

Colorectal carcinoma (CRC) is the second most common and frequent cause of cancer-related deaths for men and women in the world. PIK3CA and PIK3CB that reverse multidrug resistance (MDR) can serve as predictive and prognostic markers as well as therapeutic targets for CRC treatment. In the present study, we showed that PIK3CA and PIK3CB are upregulated in CRCs and positively correlated with MDR-1, LRP, and GST-π. Long-term monitoring of 316 CRC patients showed that PIK3CA and PIK3CB were associated with poor survival time as shown by Kaplan-Meier analysis. Furthermore, we found that the downregulation of PIK3CA and PIK3CB reversed MDR; inhibited the capability of proliferation, migration, and invasion of CRC cells; and slowed down the CRC tumor growth in nude mice. Consistent with clinical observations, PIK3CA and PIK3CB significantly increase multidrug resistance of CRC cells in vivo. Together, these results suggest that PIK3CA and PIK3CB may be used as potential therapeutic drug targets for colorectal cancer.

Long non-coding RNA FOXD2-AS1 contributes to colorectal cancer proliferation through its interaction with microRNA-185-5p.

PubMed

Zhu, Yanyan; Qiao, Liang; Zhou, Yun; Ma, Ning; Wang, Chaojie; Zhou, Jianwei

2018-05-08

Emerging evidence has indicated that long non-coding RNA plays an important role in the carcinogenesis at the transcriptional and post-translational levels. The regulation of carcinogenesis related effectors is potent in the determination of tumor initiation and progression. In current study, FOXD2-AS1 was found to interact with miR-185-5p to modulate proliferation, migration and invasion of colorectal cancer (CRC) cells. Interestingly, cell division control (CDC) 42 expression was significantly influenced by FOXD2-AS1 and miR-185-5p. In CRC patients, the expression level of FOXD2-AS1 in CRC tissue was closely associated with miR-185-5p and CDC42, and implicated in the overall survival rate of CRC patient. Therefore, our study suggests that lncRNA FOXD2-AS1 plays a positive role in the CRC and could be developed and used as a potential biomarker for the diagnosis and therapy of CRC. This will greatly improve to the prevention and treatment of this third most common cancer worldwide. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

DUSP5 is methylated in CIMP-high colorectal cancer but is not a major regulator of intestinal cell proliferation and tumorigenesis.

PubMed

Tögel, Lars; Nightingale, Rebecca; Wu, Rui; Chüeh, Anderly C; Al-Obaidi, Sheren; Luk, Ian; Dávalos-Salas, Mercedes; Chionh, Fiona; Murone, Carmel; Buchanan, Daniel D; Chatterton, Zac; Sieber, Oliver M; Arango, Diego; Tebbutt, Niall C; Williams, David; Dhillon, Amardeep S; Mariadason, John M

2018-01-29

The ERK signalling pathway regulates key cell fate decisions in the intestinal epithelium and is frequently dysregulated in colorectal cancers (CRCs). Variations in the dynamics of ERK activation can induce different biological outcomes and are regulated by multiple mechanisms, including activation of negative feedback loops involving transcriptional induction of dual-specificity phosphatases (DUSPs). We have found that the nuclear ERK-selective phosphatase DUSP5 is downregulated in colorectal tumours and cell lines, as previously observed in gastric and prostate cancer. The DUSP5 promoter is methylated in a subset of CRC cell lines and primary tumours, particularly those with a CpG island methylator phenotype (CIMP). However, this epigenetic change alone could not account for reduced DUSP5 expression in CRC cells. Functionally, DUSP5 depletion failed to alter ERK signalling or proliferation in CRC cell lines, and its transgenic overexpression in the mouse intestine had minimal impact on normal intestinal homeostasis or tumour development. Our results suggest that DUSP5 plays a limited role in regulating ERK signalling associated with the growth of colorectal tumours, but that methylation the DUSP5 gene promoter can serve as an additional means of identifying CIMP-high colorectal cancers.

IFN-α potentiates the direct and immune-mediated antitumor effects of epigenetic drugs on both metastatic and stem cells of colorectal cancer.

PubMed

Buoncervello, Maria; Romagnoli, Giulia; Buccarelli, Mariachiara; Fragale, Alessandra; Toschi, Elena; Parlato, Stefania; Lucchetti, Donatella; Macchia, Daniele; Spada, Massimo; Canini, Irene; Sanchez, Massimo; Falchi, Mario; Musella, Martina; Biffoni, Mauro; Belardelli, Filippo; Capone, Imerio; Sgambato, Alessandro; Vitiani, Lucia Ricci; Gabriele, Lucia

2016-05-03

Epigenetic alterations, including dysregulated DNA methylation and histone modifications, govern the progression of colorectal cancer (CRC). Cancer cells exploit epigenetic regulation to control cellular pathways, including apoptotic and metastatic signals. Since aberrations in epigenome can be pharmacologically reversed by DNA methyltransferase and histone deacetylase inhibitors, epigenetics in combination with standard agents are currently envisaged as a new therapeutic frontier in cancer, expected to overcome drug resistance associated with current treatments. In this study, we challenged this idea and demonstrated that the combination of azacitidine and romidepsin with IFN-α owns a high therapeutic potential, targeting the most aggressive cellular components of CRC, such as metastatic cells and cancer stem cells (CSCs), via tight control of key survival and death pathways. Moreover, the antitumor efficacy of this novel pharmacological approach is associated with induction of signals of immunogenic cell death. Of note, a previously undisclosed key role of IFN-α in inducing both antiproliferative and pro-apoptotic effects on CSCs of CRC was also found. Overall, these findings open a new frontier on the suitability of IFN-α in association with epigenetics as a novel and promising therapeutic approach for CRC management.

IFN-α potentiates the direct and immune-mediated antitumor effects of epigenetic drugs on both metastatic and stem cells of colorectal cancer

PubMed Central

Buoncervello, Maria; Fragale, Alessandra; Toschi, Elena; Parlato, Stefania; Lucchetti, Donatella; Macchia, Daniele; Spada, Massimo; Canini, Irene; Sanchez, Massimo; Falchi, Mario; Musella, Martina; Biffoni, Mauro; Belardelli, Filippo; Capone, Imerio; Sgambato, Alessandro; Vitiani, Lucia Ricci; Gabriele, Lucia

2016-01-01

Epigenetic alterations, including dysregulated DNA methylation and histone modifications, govern the progression of colorectal cancer (CRC). Cancer cells exploit epigenetic regulation to control cellular pathways, including apoptotic and metastatic signals. Since aberrations in epigenome can be pharmacologically reversed by DNA methyltransferase and histone deacetylase inhibitors, epigenetics in combination with standard agents are currently envisaged as a new therapeutic frontier in cancer, expected to overcome drug resistance associated with current treatments. In this study, we challenged this idea and demonstrated that the combination of azacitidine and romidepsin with IFN-α owns a high therapeutic potential, targeting the most aggressive cellular components of CRC, such as metastatic cells and cancer stem cells (CSCs), via tight control of key survival and death pathways. Moreover, the antitumor efficacy of this novel pharmacological approach is associated with induction of signals of immunogenic cell death. Of note, a previously undisclosed key role of IFN-α in inducing both antiproliferative and pro-apoptotic effects on CSCs of CRC was also found. Overall, these findings open a new frontier on the suitability of IFN-α in association with epigenetics as a novel and promising therapeutic approach for CRC management. PMID:27028869

Targeting ADAM17 inhibits human colorectal adenocarcinoma progression and tumor-initiating cell frequency.

PubMed

Dosch, Joseph; Ziemke, Elizabeth; Wan, Shanshan; Luker, Kathryn; Welling, Theodore; Hardiman, Karin; Fearon, Eric; Thomas, Suneetha; Flynn, Matthew; Rios-Doria, Jonathan; Hollingsworth, Robert; Herbst, Ronald; Hurt, Elaine; Sebolt-Leopold, Judith

2017-09-12

ADAM17 (a disintegrin and metalloproteinase 17)/TACE (TNFα converting enzyme) has emerged as a potential therapeutic target in colorectal cancer (CRC) and other cancers, due in part to its role in regulating various tumor cell surface proteins and growth factors and cytokines in the tumor microenvironment. The emergence of MEDI3622, a highly potent and specific antibody-based ADAM17 inhibitor, has allowed testing of the concept that targeting ADAM17 may be an important new therapeutic approach for CRC patients. We demonstrate that MEDI3622 is highly efficacious on tumor growth in multiple human CRC PDX models, resulting in improved survival of animals bearing tumor xenografts. MEDI3622 was further found to impact Notch pathway activity and tumor-initiating cells. The promising preclinical activity seen here supports further clinical investigation of this treatment approach to improve therapeutic outcome for patients diagnosed with metastatic CRC, including patients with KRAS-mutant tumors for whom other therapeutic options are currently limited.

Metformin and aspirin treatment could lead to an improved survival rate for Type 2 diabetic patients with stage II and III colorectal adenocarcinoma relative to non-diabetic patients.

PubMed

De Monte, Ariella; Brunetti, Davide; Cattin, Luigi; Lavanda, Francesca; Naibo, Erica; Malagoli, Maria; Stanta, Giorgio; Bonin, Serena

2018-03-01

Metformin, the drug of choice in the treatment of type 2 diabetes mellitus (DM2), in addition to aspirin (ASA), the drug prescribed for cardioprotection of diabetic and non-diabetic patients, have an inhibitory effect on cancer cell survival. The present population-based study conducted in the province of Trieste (Italy), aimed to investigate the prevalence of DM2 in patients with colorectal adenocarcinoma (CRC) and survival for CRC in diabetic and nondiabetic patients. All permanent residents diagnosed with a CRC between 2004 and 2007 were ascertained through the regional health information system. CRC-specific and relative survival probabilities were computed for each group of patients defined by CRC stage, presence or absence of DM2 treated with metformin, and presence or absence of daily ASA therapy. A total of 515 CRC patients without DM2 and 156 with DM2 treated with metformin were enrolled in the study. At the time of CRC diagnosis, 71 (14%) nondiabetic and 39 (25%) diabetic patients were taking ASA daily. The five-year relative survival for stage III CRC was 101% [95% confidence interval (CI)=76-126] in the 18 patients with DM2 treated with metformin and ASA, 55% (95% CI=31-78) in the 23 without DM2 treated with ASA, 55% (95% CI=45-65) in the 150 without DM2 not taking ASA, and 29% (95% CI=13-45) in the 43 with DM2 treated with metformin, however not with ASA. The findings support the hypothesis of a possible inhibitory effect of metformin and ASA on CRC cells. Randomized controlled trials are required to verify this hypothesis.

Enrichment of colorectal cancer associations in functional regions: Insight for using epigenomics data in the analysis of whole genome sequence-imputed GWAS data.

PubMed

Bien, Stephanie A; Auer, Paul L; Harrison, Tabitha A; Qu, Conghui; Connolly, Charles M; Greenside, Peyton G; Chen, Sai; Berndt, Sonja I; Bézieau, Stéphane; Kang, Hyun M; Huyghe, Jeroen; Brenner, Hermann; Casey, Graham; Chan, Andrew T; Hopper, John L; Banbury, Barbara L; Chang-Claude, Jenny; Chanock, Stephen J; Haile, Robert W; Hoffmeister, Michael; Fuchsberger, Christian; Jenkins, Mark A; Leal, Suzanne M; Lemire, Mathieu; Newcomb, Polly A; Gallinger, Steven; Potter, John D; Schoen, Robert E; Slattery, Martha L; Smith, Joshua D; Le Marchand, Loic; White, Emily; Zanke, Brent W; Abeçasis, Goncalo R; Carlson, Christopher S; Peters, Ulrike; Nickerson, Deborah A; Kundaje, Anshul; Hsu, Li

2017-01-01

The evaluation of less frequent genetic variants and their effect on complex disease pose new challenges for genomic research. To investigate whether epigenetic data can be used to inform aggregate rare-variant association methods (RVAM), we assessed whether variants more significantly associated with colorectal cancer (CRC) were preferentially located in non-coding regulatory regions, and whether enrichment was specific to colorectal tissues. Active regulatory elements (ARE) were mapped using data from 127 tissues and cell-types from NIH Roadmap Epigenomics and Encyclopedia of DNA Elements (ENCODE) projects. We investigated whether CRC association p-values were more significant for common variants inside versus outside AREs, or 2) inside colorectal (CR) AREs versus AREs of other tissues and cell-types. We employed an integrative epigenomic RVAM for variants with allele frequency <1%. Gene sets were defined as ARE variants within 200 kilobases of a transcription start site (TSS) using either CR ARE or ARE from non-digestive tissues. CRC-set association p-values were used to evaluate enrichment of less frequent variant associations in CR ARE versus non-digestive ARE. ARE from 126/127 tissues and cell-types were significantly enriched for stronger CRC-variant associations. Strongest enrichment was observed for digestive tissues and immune cell types. CR-specific ARE were also enriched for stronger CRC-variant associations compared to ARE combined across non-digestive tissues (p-value = 9.6 × 10-4). Additionally, we found enrichment of stronger CRC association p-values for rare variant sets of CR ARE compared to non-digestive ARE (p-value = 0.029). Integrative epigenomic RVAM may enable discovery of less frequent variants associated with CRC, and ARE of digestive and immune tissues are most informative. Although distance-based aggregation of less frequent variants in CR ARE surrounding TSS showed modest enrichment, future association studies would likely benefit from joint analysis of transcriptomes and epigenomes to better link regulatory variation with target genes.

Overexpression of the S100A2 protein as a prognostic marker for patients with stage II and III colorectal cancer

PubMed Central

MASUDA, TAIKI; ISHIKAWA, TOSHIAKI; MOGUSHI, KAORU; OKAZAKI, SATOSHI; ISHIGURO, MEGUMI; IIDA, SATORU; MIZUSHIMA, HIROSHI; TANAKA, HIROSHI; UETAKE, HIROYUKI; SUGIHARA, KENICHI

2016-01-01

We aimed to identify a novel prognostic biomarker related to recurrence in stage II and III colorectal cancer (CRC) patients. Stage II and III CRC tissue mRNA expression was profiled using an Affymetrix Gene Chip, and copy number profiles of 125 patients were generated using an Affymetrix 250K Sty array. Genes showing both upregulated expression and copy number gains in cases involving recurrence were extracted as candidate biomarkers. The protein expression of the candidate gene was assessed using immunohistochemical staining of tissue from 161 patients. The relationship between protein expression and clinicopathological features was also examined. We identified 9 candidate genes related to recurrence of stage II and III CRC, whose mRNA expression was significantly higher in CRC than in normal tissue. Of these proteins, the S100 calcium-binding protein A2 (S100A2) has been observed in several human cancers. S100A2 protein overexpression in CRC cells was associated with significantly worse overall survival and relapse-free survival, indicating that S100A2 is an independent risk factor for stage II and III CRC recurrence. S100A2 overexpression in cancer cells could be a biomarker of poor prognosis in stage II and III CRC recurrence and a target for treatment of this disease. PMID:26783118

CD133+CD54+CD44+ circulating tumor cells as a biomarker of treatment selection and liver metastasis in patients with colorectal cancer

PubMed Central

Wang, Cun; Huang, Qiaorong; Meng, Wentong; Yu, Yongyang; Yang, Lie; Peng, Zhihai; Hu, Jiankun; Li, Yuan; Mo, Xianming; Zhou, Zongguang

2016-01-01

Introduction Liver is the most common site of distant metastasis in colorectal cancer (CRC). Early diagnosis and appropriate treatment selection decides overall prognosis of patients. However, current diagnostic measures were basically imaging but not functional. Circulating tumor cells (CTCs) known as hold the key to understand the biology of metastatic mechanism provide a novel and auxiliary diagnostic strategy for CRC with liver metastasis (CRC-LM). Results The expression of CD133+ and CD133+CD54+CD44+ cellular subpopulations were higher in the peripheral blood of CRC-LM patients when compared with those without metastasis (P<0.001). Multivariate analysis proved the association between the expression of CD133+CD44+CD54+ cellular subpopulation and the existence of CRC-LM (P<0.001). The combination of abdominal CT/MRI, CEA and the CD133+CD44+CD54+ cellular subpopulation showed increased detection and discrimination rate for liver metastasis, with a sensitivity of 88.2% and a specificity of 92.4%. Meanwhile, it also show accurate predictive value for liver metastasis (OR=2.898, 95% C.I.1.374–6.110). Materials and Method Flow cytometry and multivariate analysis was performed to detect the expression of cancer initiating cells the correlation between cellular subpopulations and liver metastasis in patients with CRC. The receiver operating characteristic curves combined with the area under the curve were generated to compare the predictive ability of the cellular subpopulation for liver metastasis with current CT and MRI images. Conclusions The identification, expression and application of CTC subpopulations will provide an ideal cellular predictive marker for CRC liver metastasis and a potential marker for further investigation. PMID:27764803

Expression of Oct-4 is significantly associated with the development and prognosis of colorectal cancer

PubMed Central

ZHOU, HUAN; HU, YU; WANG, WEIPENG; MAO, YONG; ZHU, JINGJIE; ZHOU, BIN; SUN, JING; ZHANG, XUEGUANG

2015-01-01

Octamer-binding transcription factor 4 (Oct-4), is an essential transcription factor, which is required for pluripotency and self-renewal in embryonic stem cells and germ cells. It is also involved in maintaining cancer stem-like properties in certain types of tumor, and is an important biomarker for cancer stem cells. The present study investigated whether Oct-4 expression was associated with colorectal cancer (CRC). In order to achieve this, primary CRC tissues, matched non-tumor tissues and benign polyp tissues, representing different stages of carcinogenesis, were obtained, and Oct-4 expression was analyzed using reverse transcription-quantitative polymerase chain reaction, flow cytometry analysis and immunohistochemistry. Furthermore, the medical records of patients with CRC were reviewed, and clinicopathological analysis was performed in order to assess the association between Oct-4 expression and certain clinicopathological parameters. It was shown that the transcription and translation of Oct-4 increased in a stepwise manner, from non-tumor to benign polyp tissues, and from benign polyps to CRC tissues. Oct-4 expression in CRC was significantly correlated with histological grade (P=0.007), lymph node metastasis (P=0.027), distant metastasis (P=0.017) and TNM stage (P=0.041). Kaplan-Meier survival curve analysis demonstrated that Oct-4+ cases had a shorter median survival time (37.0 months) compared with Oct-4− cases (76.0 months; P=0.001). These results indicated that aberrant expression of Oct-4 may be involved in the development of CRC. Thus, Oct-4 may be a biomarker for the prediction, diagnosis or assessment of prognosis in CRC, in addition to a potential target for the treatment of this disease. PMID:26622555

Concise Review: Aggressive Colorectal Cancer: Role of Epithelial Cell Adhesion Molecule in Cancer Stem Cells and Epithelial-to-Mesenchymal Transition.

PubMed

Boesch, Maximilian; Spizzo, Gilbert; Seeber, Andreas

2018-06-01

Colorectal cancer (CRC) is one of the most common malignancies worldwide. In spite of various attempts to ameliorate outcome by escalating treatment, significant improvement is lacking particularly in the adjuvant setting. It has been proposed that cancer stem cells (CSCs) and the epithelial-to-mesenchymal transition (EMT) are at least partially responsible for therapy resistance in CRC. The epithelial cell adhesion molecule (EpCAM) was one of the first CSC antigens to be described. Furthermore, an EpCAM-specific antibody (edrecolomab) has the merit of having launched the era of monoclonal antibody treatment in oncology in the 1990s. However, despite great initial enthusiasm, monoclonal antibody treatment has not proven successful in the adjuvant treatment of CRC patients. In the meantime, new insights into the function of EpCAM in CRC have emerged and new drugs targeting various epitopes have been developed. In this review article, we provide an update on the role of EpCAM in CSCs and EMT, and emphasize the potential predictive selection criteria for novel treatment strategies and refined clinical trial design. Stem Cells Translational Medicine 2018;7:495-501. © 2018 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Treatment of peritoneal carcinomatosis with hyperthermic intraperitoneal chemotherapy in colorectal cancer.

PubMed

Behrenbruch, Corina; Hollande, Frédéric; Thomson, Benjamin; Michael, Michael; Warrier, Satish K; Lynch, Craig; Heriot, Alexander

2017-09-01

The peritoneum is the second most common site of metastasis after the liver and the only site of metastatic disease in approximately 25% of patients with colorectal cancer (CRC). In the past, peritoneal carcinomatosis in CRC was thought to be equivalent to distant metastasis; however, the transcoelomic spread of malignant cells is an acknowledged alternative pathway. Metastasectomy with curative intent is well accepted in patients with liver metastasis in CRC despite the paucity of randomized trials. Therefore, there is rationale for local treatment with peritonectomy to eliminate macroscopic disease, followed by hyperthermic intraperitoneal chemotherapy to destroy any residual free tumour cells within the peritoneal cavity. The aim of this paper is to summarize the current evidence for cytoreduction and hyperthermic intraperitoneal chemotherapy in the treatment of peritoneal carcinomatosis in CRC. © 2017 Royal Australasian College of Surgeons.

Conserved mechanisms of tumorigenesis in the Drosophila adult midgut.

PubMed

Martorell, Òscar; Merlos-Suárez, Anna; Campbell, Kyra; Barriga, Francisco M; Christov, Christo P; Miguel-Aliaga, Irene; Batlle, Eduard; Casanova, Jordi; Casali, Andreu

2014-01-01

Whereas the series of genetic events leading to colorectal cancer (CRC) have been well established, the precise functions that these alterations play in tumor progression and how they disrupt intestinal homeostasis remain poorly characterized. Activation of the Wnt/Wg signaling pathway by a mutation in the gene APC is the most common trigger for CRC, inducing benign lesions that progress to carcinomas due to the accumulation of other genetic alterations. Among those, Ras mutations drive tumour progression in CRC, as well as in most epithelial cancers. As mammalian and Drosophila's intestines share many similarities, we decided to explore the alterations induced in the Drosophila midgut by the combined activation of the Wnt signaling pathway with gain of function of Ras signaling in the intestinal stem cells. Here we show that compound Apc-Ras clones, but not clones bearing the individual mutations, expand as aggressive intestinal tumor-like outgrowths. These lesions reproduce many of the human CRC hallmarks such as increased proliferation, blockade of cell differentiation and cell polarity and disrupted organ architecture. This process is followed by expression of tumoral markers present in human lesions. Finally, a metabolic behavioral assay shows that these flies suffer a progressive deterioration in intestinal homeostasis, providing a simple readout that could be used in screens for tumor modifiers or therapeutic compounds. Taken together, our results illustrate the conservation of the mechanisms of CRC tumorigenesis in Drosophila, providing an excellent model system to unravel the events that, upon mutation in Apc and Ras, lead to CRC initiation and progression.

Isolation of Circulating Tumor Cells in an Orthotopic Mouse Model of Colorectal Cancer.

PubMed

Kochall, Susan; Thepkaysone, May-Linn; García, Sebastián A; Betzler, Alexander M; Weitz, Jürgen; Reissfelder, Christoph; Schölch, Sebastian

2017-07-18

Despite the advantages of easy applicability and cost-effectiveness, subcutaneous mouse models have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Orthotopic mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in hollow organs such as the large bowel. In order to produce uniform tumors which reliably grow and metastasize, standardized techniques of tumor cell preparation and injection are critical. We have developed an orthotopic mouse model of colorectal cancer (CRC) which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor cells from either primary tumors, 2-dimensional (2D) cell lines or 3-dimensional (3D) organoids are injected into the cecum and, depending on the metastatic potential of the injected tumor cells, form highly metastatic tumors. In addition, CTCs can be found regularly. We here describe the technique of tumor cell preparation from both 2D cell lines and 3D organoids as well as primary tumor tissue, the surgical and injection techniques as well as the isolation of CTCs from the tumor-bearing mice, and present tips for troubleshooting.

A Rapid Filter Insert-based 3D Culture System for Primary Prostate Cell Differentiation

PubMed Central

Tricoli, Lucas; Berry, Deborah L.; Albanese, Chris

2018-01-01

Conditionally reprogrammed cells (CRCs) provide a sustainable method for primary cell culture and the ability to develop extensive “living biobanks” of patient derived cell lines. For many types of epithelial cells, various three dimensional (3D) culture approaches have been described that support an improved differentiated state. While CRCs retain their lineage commitment to the tissue from which they are isolated, they fail to express many of the differentiation markers associated with the tissue of origin when grown under normal two dimensional (2D) culture conditions. To enhance the application of patient-derived CRCs for prostate cancer research, a 3D culture format has been defined that enables a rapid (2 weeks total) luminal cell differentiation in both normal and tumor-derived prostate epithelial cells. Herein, a filter insert-based format is described for the culturing and differentiation of both normal and malignant prostate CRCs. A detailed description of the procedures required for cell collection and processing for immunohistochemical and immunofluorescent staining are provided. Collectively the 3D culture format described, combined with the primary CRC lines, provides an important medium- to high- throughput model system for biospecimen-based prostate research. PMID:28287583

Target Identification of Grape Seed Extract in Colorectal Cancer using Drug Affinity Responsive Target Stability (DARTS) Technique: Role of Endoplasmic Reticulum Stress Response Proteins

PubMed Central

Derry, Molly M.; Somasagara, Ranganatha; Raina, Komal; Kumar, Sushil; Gomez, Joe; Patel, Manisha; Agarwal, Rajesh; Agarwal, Chapla

2014-01-01

Various natural agents, including grape seed extract (GSE), have shown considerable chemopreventive and anti-cancer efficacy against different cancers in pre-clinical studies; however, their specific protein targets are largely unknown and thus, their clinical usefulness is marred by limited scientific evidences about their direct cellular targets. Accordingly, herein, employing, for the first time, the recently developed drug affinity responsive target stability (DARTS) technique, we aimed to profile the potential protein targets of GSE in human colorectal cancer (CRC) cells. Unlike other methods, which can cause chemical alteration of the drug components to allow for detection, this approach relies on the fact that a drug bound protein may become less susceptible to proteolysis and hence the enriched proteins can be detected by Mass Spectroscopy methods. Our results, utilizing the DARTS technique followed by examination of the spectral output by LC/MS and the MASCOT data, revealed that GSE targets endoplasmic reticulum (ER) stress response proteins resulting in overall down regulation of proteins involved in translation and that GSE also causes oxidative protein modifications, specifically on methionine amino acids residues on its protein targets. Corroborating these findings, mechanistic studies revealed that GSE indeed caused ER stress and strongly inhibited PI3k-Akt–mTOR pathway for its biological effects in CRC cells. Furthermore, bioenergetics studies indicated that GSE also interferes with glycolysis and mitochondrial metabolism in CRC cells. Together, the present study identifying GSE molecular targets in CRC cells, combined with its efficacy in vast pre-clinical CRC models, further supports its usefulness for CRC prevention and treatment. PMID:24724981

Circulating tumor cell levels are elevated in colorectal cancer patients with high tumor burden in the liver.

PubMed

Kaifi, Jussuf T; Kunkel, Miriam; Dicker, David T; Joude, Jamal; Allen, Joshua E; Das, Avisnata; Zhu, Junjia; Yang, Zhaohai; Sarwani, Nabeel E; Li, Guangfu; Staveley-O'Carroll, Kevin F; El-Deiry, Wafik S

2015-01-01

Metastatic spread is the most common cause of cancer-related death in colorectal cancer (CRC) patients, with the liver being the mostly affected organ. Circulating tumor cells (CTCs) are a prognostic marker in stage IV CRC. We hypothesized that tumor burden in the liver correlates with CTC quantity. Blood (7.5 ml) was prospectively collected from 24 patients with novel stage IV CRC diagnosis. Baseline EpCAM+ CTCs were analyzed with the FDA-approved CellSearch® system. Clinicopathological data were collected, and hepatic tumor burden was determined by radiographic liver volumetry with contrast-enhanced CT scans. CRC primary tumors were immunohistochemically stained for EpCAM expression with BerEP4 monoclonal antibody. Statistical analyses were performed using 2-sample T-test, non-parametric Wilcoxon Rank-Sum test, and Fisher's exact test. CTCs were detected n 17 (71%) of 24 patients. The overall mean CTC number as determined by EpCAM-based CellSearch® detection was 6.3 (SEM 2.9). High baseline CTC numbers (≥3) correlated significantly with a high tumor/liver ratio (≥30%), and with high serum CEA levels, as determined by two-sample T-test on log-transformed data and by Fisher's Exact test on categorical data analysis (P < 0.05). The CRC primary tumors were consistently expressing EpCAM by immunostaining. High tumor burden in the liver and high baseline serum CEA levels are associated with high number of baseline CTCs in stage IV CRC patients. Future studies should further investigate the biological role and expression patterns of single CTCs in cancer patients to further improve personalized treatment strategies.

High blood sugar levels significantly impact the prognosis of colorectal cancer patients through down-regulation of microRNA-16 by targeting Myb and VEGFR2

PubMed Central

Huang, Ching-Wen; Lu, Chien-Yu; Miao, Zhi-Feng; Chang, Se-Fen; Juo, Suh-Hang Hank; Wang, Jaw-Yuan

2016-01-01

The high prevalence of type 2 diabetes mellitus in colorectal cancer patients is a crucial public health issue worldwide. The deregulation of microRNAs has been shown to be associated with the progression of CRC; however, the effects of high blood sugar levels on miR deregulation and, in turn, CRC remain unexplored. In this study, 520 CRC patients were classified into two groups according to their blood sugar levels (≧110 or <110 mg/dL). Clinicopathologic features, clinical outcomes, and serum miR-16 levels of the two groups were then analyzed, while cell cycles, cell proliferation, migration, and cellular miR-16 expression were investigated via D-(+)-glucose administration. Additionally, the target genes of miR-16 were identified. Through multivariate analysis, both the disease-free survival and overall survival of the CRC patients were found to be associated with the UICC stage, perineural invasion, and blood glucose levels (P < 0.05). Serum miR-16 levels were significantly lower in the high blood glucose patients than in the normal blood glucose patients (P = 0.0329). With D-(+)-glucose administration, the proliferation and migration of CRC cells in vitro increased remarkably (P < 0.05), while their accumulation in the G1 phase decreased significantly. Cellular miR-16 expression was suppressed by D-(+)-glucose administration. The expression levels of two target genes, Myb and VEGFR2, were affected significantly by miR-16, while glucose administration inhibited miR-16 expression and enhanced tumor cell proliferation and migration. Hyperglycemia can impact the clinical outcomes of CRC patients, likely by inhibiting miR-16 expression and the expression of its downstream genes Myb and VEGFR2. PMID:26934556

Molecular mechanisms for inhibition of colon cancer cells by combined epigenetic-modulating epigallocatechin gallate and sodium butyrate.

PubMed

Saldanha, Sabita N; Kala, Rishabh; Tollefsbol, Trygve O

2014-05-15

Bioactive compounds are considered safe and have been shown to alter genetic and epigenetic profiles of tumor cells. However, many of these changes have been reported at molecular concentrations higher than physiologically achievable levels. We investigated the role of the combinatorial effects of epigallocatechin gallate (EGCG), a predominant polyphenol in green tea, and sodium butyrate (NaB), a dietary microbial fermentation product of fiber, in the regulation of survivin, which is an overexpressed anti-apoptotic protein in colon cancer cells. For the first time, our study showed that the combination treatment induced apoptosis and cell cycle arrest in RKO, HCT-116 and HT-29 colorectal cancer cells. This was found to be regulated by the decrease in HDAC1, DNMT1, survivin and HDAC activity in all three cell lines. A G2/M arrest was observed for RKO and HCT-116 cells, and G1 arrest for HT-29 colorectal cancer cells for combinatorial treatment. Further experimentation of the molecular mechanisms in RKO colorectal cancer (CRC) cells revealed a p53-dependent induction of p21 and an increase in nuclear factor kappa B (NF-κB)-p65. An increase in double strand breaks as determined by gamma-H2A histone family member X (γ-H2AX) protein levels and induction of histone H3 hyperacetylation was also observed with the combination treatment. Further, we observed a decrease in global CpG methylation. Taken together, these findings suggest that at low and physiologically achievable concentrations, combinatorial EGCG and NaB are effective in promoting apoptosis, inducing cell cycle arrest and DNA-damage in CRC cells. Copyright © 2014 Elsevier Inc. All rights reserved.

Digital Detection of Multiple Minority Mutants and Expression Levels of Multiple Colorectal Cancer-Related Genes Using Digital-PCR Coupled with Bead-Array

PubMed Central

Huang, Huan; Li, Shuo; Sun, Lizhou; Zhou, Guohua

2015-01-01

To simultaneously analyze mutations and expression levels of multiple genes on one detection platform, we proposed a method termed “multiplex ligation-dependent probe amplification–digital amplification coupled with hydrogel bead-array” (MLPA–DABA) and applied it to diagnose colorectal cancer (CRC). CRC cells and tissues were sampled to extract nucleic acid, perform MLPA with sequence-tagged probes, perform digital emulsion polymerase chain reaction (PCR), and produce a hydrogel bead-array to immobilize beads and form a single bead layer on the array. After hybridization with fluorescent probes, the number of colored beads, which reflects the abundance of expressed genes and the mutation rate, was counted for diagnosis. Only red or green beads occurred on the chips in the mixed samples, indicating the success of single-molecule PCR. When a one-source sample was analyzed using mixed MLPA probes, beads of only one color occurred, suggesting the high specificity of the method in analyzing CRC mutation and gene expression. In gene expression analysis of a CRC tissue from one CRC patient, the mutant percentage was 3.1%, and the expression levels of CRC-related genes were much higher than those of normal tissue. The highly sensitive MLPA–DABA succeeds in the relative quantification of mutations and gene expressions of exfoliated cells in stool samples of CRC patients on the same chip platform. MLPA–DABA coupled with hydrogel bead-array is a promising method in the non-invasive diagnosis of CRC. PMID:25880764

Sulfonamide derivative targeting carbonic anhydrase IX as a nuclear imaging probe for colorectal cancer detection in vivo

PubMed Central

Guan, Siao-Syun; Cheng, Chun-Chia; Ho, Ai-Sheng; Wang, Chia-Chi; Luo, Tsai-Yueh; Liao, Tse-Zung; Chang, Jungshan; Wu, Cheng-Tien; Liu, Shing-Hwa

2015-01-01

Hypoxic microenvironment is a common situation in solid tumors. Carbonic anhydrase IX (CA9) is one of the reliable cellular biomarkers of hypoxia. The role of CA9 in colorectal cancer (CRC) remains to be clarified. CA9 inhibitor such as sulfonamides is known to block CA9 activation and reduce tumor growth consequently. Here, we aimed to investigate the CA9 expression in serum and tumor from different stages of CRC patients and utilize sulfonamide derivative with indium-111 labeling as a probe for CRC nuclear imaging detection in vivo. The serum CA9 was correlated with the tumor CA9 levels in different stages of CRC patients. Hypoxia increased cell viability and CA9 expression in colorectal cancer HCT-15 cells. Sulfonamide derivative 5-(2-aminoethyl)thiophene-2-sulfonamide (ATS) could bind with CA9 in vitro under hypoxia. Moreover, tumor tissues in HCT-15-induced xenograft mice possessed higher hypoxic fluorescence signal as compared with other organs. We also found that the radioisotope signal of indium-111 labeled ATS, which was utilized for CRC detection in HCT-15-induced xenograft mice, was markedly enhanced in tumors as compared with non-ATS control. Taken together, these findings suggest that CA9 is a potential hypoxic CRC biomarker and measurement of serum CA9 can be as a potential tool for diagnosing CA9 expressions in CRC clinical practice. The radioisotope-labeled sulfonamide derivative (ATS) may be useful to apply in CRC patients for nuclear medicine imaging. PMID:26447758

Combination of preoperative NLR, PLR and CEA could increase the diagnostic efficacy for I-III stage CRC.

PubMed

Peng, Hong-Xin; Yang, Lin; He, Bang-Shun; Pan, Yu-Qin; Ying, Hou-Qun; Sun, Hui-Ling; Lin, Kang; Hu, Xiu-Xiu; Xu, Tao; Wang, Shu-Kui

2017-09-01

Inflammation plays an important role in the development and progression of CRC. The members of inflammatory biomarkers, preoperative NLR and PLR, have been proved by numerous studies to be promising prognostic biomarkers for CRC. However, the diagnostic value of the two biomarkers in CRC remains unknown, and no study reported the combined diagnostic efficacy of NLR, PLR and CEA. Five hundred and fifty-nine patients with I-III stage CRC undergoing surgical resection and 559 gender- and age-matched healthy controls were enrolled in this retrospective study. NLR and PLR were calculated from preoperative peripheral blood cell count detected using white blood cell five classification by Sysmex XT-1800i Automated Hematology System and serum CEA were measured by electrochemiluminescence by ELECSYS 2010. The diagnostic performance of NLR, PLR and CEA for CRC was evaluated by ROC curve. Levels of NLR and PLR in the cases were significantly higher than them in the healthy controls. ROC curves comparison analyses showed that the diagnostic efficacy of NLR (AUC=.755, 95%CI=.728-.780) alone for CRC was significantly higher than PLR (AUC=.723, 95%CI=.696-.749, P=.037) and CEA (AUC=.690, 95%CI=.662-.717, P=.002) alone. In addition, the diagnostic efficacy of the combination of NLR, PLR and CEA(AUC=.831, 95%CI=.807-.852)for CRC was not only significantly higher than NLR alone but also higher than any combinations of the two of these three biomarkers (P<.05). Moreover, the NLR and PLR in the patients with TNM stage I/II was higher than that in the healthy controls, and patients with stage III had a higher NLR and PLR than those with stage I/II, but no significant difference was observed. Our study indicated that preoperative NLR could be a CRC diagnostic biomarker, even for early stage CRC, and the combination of NLR, PLR and CEA could significantly improve the diagnostic efficacy. © 2016 Wiley Periodicals, Inc.

Abundance of the Organic Anion-transporting Polypeptide OATP4A1 in Early-Stage Colorectal Cancer Patients: Association With Disease Relapse.

PubMed

Buxhofer-Ausch, Veronika; Sheikh, Maidah; Ausch, Christoph; Zotter, Simone; Bauer, Heike; Mollik, Marina; Reiner, Angelika; Gleiss, Andreas; Jäger, Walter; Sebesta, Christian; Kriwanek, Stephan; Thalhammer, Theresia

2018-05-03

The abundance of OATP4A1 in colorectal cancer (CRC) might be related to tumor progression. This was studied by immunohistochemistry on paraffin-embedded samples obtained from 178 patients (43 patients with a relapse within 5 y) with early-stage CRC. Positivity for OATP4A1 in tumor cells and noncancerous mucosal cells was proved by double-immunofluorescence staining with antibodies against OATP4A1 and keratin 8, whereas antibodies against appropriate CD markers were used to identify immune cells. Automated microscopic image analysis was used to measure the percentage of OATP4A1-positive cells and OATP4A1 staining intensity in tumor, immune, and adjacent normal-looking mucosal cells separately, as well as in the mucosal and immune cells of 14 nonmalignant tissue samples. In CRC the percentage of OATP4A1-positive cells, but not staining intensity, was significantly higher in tumor and mucosal cells adjacent to the tumor compared to the mucosa of nonmalignant samples (P<0.001 each). No difference was registered between immune cells in malignant and nonmalignant samples. Importantly, high levels of OATP4A1 in immune (odds ratio, 0.73; confidence interval, 0.63-0.85; P<0.001), and tumor cells (odds ratio, 0.79; confidence interval, 0.69-0.91; P<0.001) are significantly associated with a low risk of recurrence and also significantly enhance the discriminative power of other clinical parameters [such as International Union Against Cancer (UICC), adjuvant therapy, localization of the primary tumor] of the risk of relapse (receiver operating characteristics analysis; P=0.002). Using an advanced digital microscopic quantification procedure, we showed that OATP4A1 abundance is negatively associated with tumor recurrence in early-stage CRC. This digital scoring procedure may serve as a novel tool for the assessment of potential prognostic markers in early-stage CRC.

[Cost-effectiveness Analysis of Panitumumab Plus mFOLFOX6 Compared to Bevacizumab Plus mFOLFOX6 for First-line Treatment of Patients with Wild-type RAS Metastatic Colorectal Cancer--Czech Republic Model Adaptation].

PubMed

Fínek, J; Skoupá, J; Jandová, P

2015-01-01

Pharmacoeconomic assessments are a part of the decision process not only during reimbursement setting, but in clinical practice as well. The presented cost-effectiveness analysis assesses panitumumab+mFOLFOX6 vs. bevacizumab+mFOLFOX6 in 1st line treatment of patients with wildtype RAS metastatic colorectal cancer (mCRC) in the Czech environment. The adaptation of a Markov model considers the healthcare perspective; clinical data (efficacy, healthcare utilization and adverse events) are derived from a head-to-head comparison (PEAK study). Health states included in the model: progression free on treatment, progression (with/ without active treatment), resection of metastases, disease-free after successful resection and death. Actual reimbursement levels were used to estimate costs, published literature to estimate duration of 2nd line treatment. The analysis assumes a lifetime horizon; uncertainty was limited by performing one-way and probabilistic sensitivity analyses. Analysis outcomes are life-years gained (LYG) and quality-adjusted life-years (QALYs). Panitumumab+mFOLFOX6 is more effective and more costly in 1st line patients with wildtype RAS mCRC. Incremental costs per QALY are 837,270 CZK, per LYG 615,022 CZK; however, below the willingness-to-pay threshold applied in the Czech Republic. Panitumumab+mFOLFOX6 is cost-effective in 1st line treatment of patients with wildtype RAS mCRC compared to bevacizumab+mFOLFOX6 in the Czech setting.

Epigenetics and colorectal cancer pathogenesis.

PubMed

Bardhan, Kankana; Liu, Kebin

2013-06-05

Colorectal cancer (CRC) develops through a multistage process that results from the progressive accumulation of genetic mutations, and frequently as a result of mutations in the Wnt signaling pathway. However, it has become evident over the past two decades that epigenetic alterations of the chromatin, particularly the chromatin components in the promoter regions of tumor suppressors and oncogenes, play key roles in CRC pathogenesis. Epigenetic regulation is organized at multiple levels, involving primarily DNA methylation and selective histone modifications in cancer cells. Assessment of the CRC epigenome has revealed that virtually all CRCs have aberrantly methylated genes and that the average CRC methylome has thousands of abnormally methylated genes. Although relatively less is known about the patterns of specific histone modifications in CRC, selective histone modifications and resultant chromatin conformation have been shown to act, in concert with DNA methylation, to regulate gene expression to mediate CRC pathogenesis. Moreover, it is now clear that not only DNA methylation but also histone modifications are reversible processes. The increased understanding of epigenetic regulation of gene expression in the context of CRC pathogenesis has led to development of epigenetic biomarkers for CRC diagnosis and epigenetic drugs for CRC therapy.

Epigenetics and Colorectal Cancer Pathogenesis

PubMed Central

Bardhan, Kankana; Liu, Kebin

2013-01-01

Colorectal cancer (CRC) develops through a multistage process that results from the progressive accumulation of genetic mutations, and frequently as a result of mutations in the Wnt signaling pathway. However, it has become evident over the past two decades that epigenetic alterations of the chromatin, particularly the chromatin components in the promoter regions of tumor suppressors and oncogenes, play key roles in CRC pathogenesis. Epigenetic regulation is organized at multiple levels, involving primarily DNA methylation and selective histone modifications in cancer cells. Assessment of the CRC epigenome has revealed that virtually all CRCs have aberrantly methylated genes and that the average CRC methylome has thousands of abnormally methylated genes. Although relatively less is known about the patterns of specific histone modifications in CRC, selective histone modifications and resultant chromatin conformation have been shown to act, in concert with DNA methylation, to regulate gene expression to mediate CRC pathogenesis. Moreover, it is now clear that not only DNA methylation but also histone modifications are reversible processes. The increased understanding of epigenetic regulation of gene expression in the context of CRC pathogenesis has led to development of epigenetic biomarkers for CRC diagnosis and epigenetic drugs for CRC therapy. PMID:24216997

Exosomes from human colorectal cancer induce a tumor-like behavior in colonic mesenchymal stromal cells

PubMed Central

Lugini, Luana; Valtieri, Mauro; Federici, Cristina; Cecchetti, Serena; Meschini, Stefania; Condello, Maria; Signore, Michele; Fais, Stefano

2016-01-01

Background Cancer cells, including colorectal cancer ones (CRC), release high amounts of nanovesicles (exosomes), delivering biochemical messages for paracrine or systemic crosstalk. Mesenchymal stromal cells (MSCs) have been shown to play contradicting roles in tumor progression. Results CRC exosomes induce in cMSCs: i) atypical morphology, higher proliferation, migration and invasion; ii) formation of spheroids; iii) an acidic extracellular environment associated with iv) a plasma membrane redistribution of vacuolar H+-ATPase and increased expression of CEA. Colon cancer derived MSCs, which were isolated from tumor masses, produce umbilicated spheroids, a future frequently observed in the inner core of rapidly growing tumors and recapitulate the changes observed in normal colonic MSCs exposed to CRC exosomes. Materials and Methods Tissue specific colonic (c)MSCs were exposed to primary or metastatic CRC exosomes and analysed by light and electron microscopy, proliferation in 2D and 3D cultures, migration and invasion assays, Western blot and confocal microscopy for vacuolar H+-ATPase expression. Conclusions CRC exosomes are able to induce morphological and functional changes in colonic MSCs, which may favour tumor growth and its malignant progression. Our results suggest that exosomes are actively involved in cancer progression and that inhibiting tumor exosome release may represent a way to interfere with cancer. PMID:27418137

Exosomes from human colorectal cancer induce a tumor-like behavior in colonic mesenchymal stromal cells.

PubMed

Lugini, Luana; Valtieri, Mauro; Federici, Cristina; Cecchetti, Serena; Meschini, Stefania; Condello, Maria; Signore, Michele; Fais, Stefano

2016-08-02

Cancer cells, including colorectal cancer ones (CRC), release high amounts of nanovesicles (exosomes), delivering biochemical messages for paracrine or systemic crosstalk. Mesenchymal stromal cells (MSCs) have been shown to play contradicting roles in tumor progression. CRC exosomes induce in cMSCs: i) atypical morphology, higher proliferation, migration and invasion; ii) formation of spheroids; iii) an acidic extracellular environment associated with iv) a plasma membrane redistribution of vacuolar H+-ATPase and increased expression of CEA. Colon cancer derived MSCs, which were isolated from tumor masses, produce umbilicated spheroids, a future frequently observed in the inner core of rapidly growing tumors and recapitulate the changes observed in normal colonic MSCs exposed to CRC exosomes. Tissue specific colonic (c)MSCs were exposed to primary or metastatic CRC exosomes and analysed by light and electron microscopy, proliferation in 2D and 3D cultures, migration and invasion assays, Western blot and confocal microscopy for vacuolar H+-ATPase expression. CRC exosomes are able to induce morphological and functional changes in colonic MSCs, which may favour tumor growth and its malignant progression. Our results suggest that exosomes are actively involved in cancer progression and that inhibiting tumor exosome release may represent a way to interfere with cancer.

Co-expression of LAG3 and TIM3 identifies a potent Treg population that suppresses macrophage functions in colorectal cancer patients.

PubMed

Ma, Qiang; Liu, Junning; Wu, Guoliang; Teng, Mujian; Wang, Shaoxuan; Cui, Meng; Li, Yuantao

2018-06-15

Regulatory T (Treg) cells are critical suppressors of inflammation and are thought to exert mainly deleterious effects in cancers. In colorectal cancer (CRC), Foxp3 +  Treg accumulation in the tumor was associated with poor prognosis. Hence, we examined the circulating Treg cells in CRC patients. Compared to controls, CRC patients presented mild upregulations in CD4 + CD25 +/hi T cells and in the more canonical CD4 + CD25 +/hi Foxp3 + Treg cells in peripheral blood mononuclear cells. Both of these Treg populations could be roughly divided into LAG3 - TIM3 - and LAG3 + TIM3 + subsets. In CRC patients, the LAG3 + TIM3 + subset represented approximately half of CD4 + CD25 +/hi T cells and greater than 60% of CD4 + CD25 +/hi Foxp3 + Treg cells, which was significantly more frequent than in healthy controls. Compared to the LAG3 - TIM3 - CD4 + CD25 +/hi T cells, the LAG3 + TIM3 + CD4 + CD25 +/hi T cells presented considerably higher transforming growth factor (TGF)-β and slightly higher interleukin (IL)-10 secretion, together with higher CTLA-4 and Foxp3 expression levels. Notably, macrophages following incubation with LAG3 - TIM3 - CD4 + CD25 +/hi T cells and LAG3 + TIM3 + CD4 + CD25 +/hi T cells displayed different characteristics. Macrophages incubated with LAG3 + TIM3 + CD4 + CD25 +/hi T cells presented lower expression of MHC class II, CD80, CD86, and tumor necrosis factor alpha (TNFα) but higher expression of IL-10, than macrophages incubated with LAG3 - TIM3 - CD4 + CD25 +/hi T cells. Together, our investigations demonstrated that CRC patients presented an enrichment of circulating Treg cells, in which the LAG3 + TIM3 + subset exhibited more potent expression of inhibitory molecules, and furthermore, the LAG3 + TIM3 + Treg cells could suppress the proinflammatory activation of macrophages more potently than the LAG3 - TIM3 - Treg cells. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Metformin enhances TRAIL-induced apoptosis by Mcl-1 degradation via Mule in colorectal cancer cells.

PubMed

Park, Seong Hye; Lee, Dae-Hee; Kim, Jung Lim; Kim, Bo Ram; Na, Yoo Jin; Jo, Min Jee; Jeong, Yoon A; Lee, Suk-Young; Lee, Sun Il; Lee, Yong Yook; Oh, Sang Cheul

2016-09-13

Metformin is an anti-diabetic drug with a promising anti-cancer potential. In this study, we show that subtoxic doses of metformin effectively sensitize human colorectal cancer (CRC) cells to tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), which induces apoptosis. Metformin alone did not induce apoptosis, but significantly potentiated TRAIL-induced apoptosis in CRC cells. CRC cells treated with metformin and TRAIL showed activation of the intrinsic and extrinsic pathways of caspase activation. We attempted to elucidate the underlying mechanism, and found that metformin significantly reduced the protein levels of myeloid cell leukemia 1 (Mcl-1) in CRC cells and, the overexpression of Mcl-1 inhibited cell death induced by metformin and/or TRAIL. Further experiments revealed that metformin did not affect mRNA levels, but increased proteasomal degradation and protein stability of Mcl-1. Knockdown of Mule triggered a significant decrease of Mcl-1 polyubiquitination. Metformin caused the dissociation of Noxa from Mcl-1, which allowed the binding of the BH3-containing ubiquitin ligase Mule followed by Mcl-1ubiquitination and degradation. The metformin-induced degradation of Mcl-1 required E3 ligase Mule, which is responsible for the polyubiquitination of Mcl-1. Our study is the first report indicating that metformin enhances TRAIL-induced apoptosis through Noxa and favors the interaction between Mcl-1 and Mule, which consequently affects Mcl-1 ubiquitination.

Rechallenge of oxaliplatin-containing regimens in the third- or later-line therapy for patients with heavily treated metastatic colorectal cancer

PubMed Central

Jiang, Zhimin; Wang, Huizhong; Li, Weiyu; Zhang, Bei; Xie, Derong

2018-01-01

Purpose The third- or later-line therapy available often yield poor survival benefit in patients metastatic colorectal cancer (mCRC). The retrospective study aimed to evaluate efficacy of rechallenge of oxaliplatin-containing regimens. Patients and methods Patients with mCRC who progressed from fluoropyrimidine, oxaliplatin, and irinotecan in the first- and second-line chemotherapy, were treated by reexposure to oxaliplatin-containing regimen. Patients treated by anti-epidermal growth factor receptor (EGFR) antibodies with irinotecan were included in the control arm. Results Ninety-five and 29 patients were treated with either oxaliplatin reexposure or anti-EGFR antibodies with irinotecan, respectively, as the third- or later-line therapy. The median time to treatment failure (TTF) and overall survival (OS) was 3.77 and 12.17 months in the oxaliplatin arm, with 4.77 months of TTF and 11.37 months of OS in the control arm; there was no significance between the 2 arms (p>0.05). Oxaliplatin reexposure resulted in 6.3% objective response rate with no complete response, 6 partial response, 39 stable disease, and 37 progressive disease. The disease control rate was 47.4% (45/95). The multivariate analysis found that patients who achieved disease control by oxaliplatin reexposure had a superior TTF (6.13 vs 1.7 months, p<0.001) and OS (15.73 vs 6.27 months, p<0.001) compared with those presenting with progressive disease. Conclusion This study showed that rechallenge of oxaliplatin-containing chemotherapy in the third- or later-line therapy may lead to tumor control and improved survival in mCRC patients, which was equivalent to that of anti-EGFR antibodies with irinotecan. Clinical significance Rechallenge of oxaliplatin-containing regimens in the third- or later-line of therapy is a common practice, despite few evidence available. The present study found that rechallenge of oxaliplatin-containing regimens produced equivalent tumor control and survival benefit to that of anti-EGFR antibodies with irinotecan in mCRC. PMID:29760556

Aminopeptidase A initiates tumorigenesis and enhances tumor cell stemness via TWIST1 upregulation in colorectal cancer

PubMed Central

Chuang, Hui-Yu; Jiang, Jeng-Kae; Yang, Muh-Hwa; Wang, Hsei-Wei; Li, Ming-Chun; Tsai, Chan-Yen; Jhang, Yau-Yun; Huang, Jason C.

2017-01-01

Metastasis accounts for the high mortality rate associated with colorectal cancer (CRC), but metastasis regulators are not fully understood. To identify a novel gene involved in tumor metastasis, we used oligonucleotide microarrays, transcriptome distance analyses, and machine learning algorithms to determine links between primary and metastatic colorectal cancers. Aminopeptidase A (APA; also known as ENPEP) was selected as our focus because its relationship with colorectal cancer requires clarification. Higher APA mRNA levels were observed in patients in advanced stages of cancer, suggesting a correlation between ENPEP and degree of malignancy. Our data also indicate that APA overexpression in CRC cells induced cell migration, invasion, anchorage-independent capability, and mesenchyme-like characteristics (e.g., EMT markers). We also observed TWIST induction in APA-overexpressing SW480 cells and TWIST down-regulation in HT29 cells knocked down with APA. Both APA silencing and impaired APA activity were found to reduce migratory capacity, cancer anchorage, stemness properties, and drug resistance in vitro and in vivo. We therefore suggest that APA enzymatic activity affects tumor initiation and cancer malignancy in a TWIST-dependent manner. Results from RT-qPCR and the immunohistochemical staining of specimens taken from CRC patients indicate a significant correlation between APA and TWIST. According to data from SurvExpress analyses of TWIST1 and APA mRNA expression profiles, high APA and TWIST expression are positively correlated with poor CRC prognosis. APA may act as a prognostic factor and/or therapeutic target for CRC metastasis and recurrence. PMID:28177885

A High RORγT/CD3 Ratio is a Strong Prognostic Factor for Postoperative Survival in Advanced Colorectal Cancer: Analysis of Helper T Cell Lymphocytes (Th1, Th2, Th17 and Regulatory T Cells).

PubMed

Yoshida, Naohiro; Kinugasa, Tetsushi; Miyoshi, Hiroaki; Sato, Kensaku; Yuge, Kotaro; Ohchi, Takafumi; Fujino, Shinya; Shiraiwa, Sachiko; Katagiri, Mitsuhiro; Akagi, Yoshito; Ohshima, Koichi

2016-03-01

Tumor-infiltrating lymphocytes (TILs), part of the host immune response, have been widely reported as influential factors in the tumor microenvironment for the clinical outcome of colorectal cancer (CRC). However, the network of helper T cells is very complex, and which T-cell subtypes affect the progression of CRC and postoperative prognosis remains unclear. This study investigated the expression of several subtypes of TILs including T helper type 1 (Th1), Th2, Th17, and regulatory T (Treg) cells to determine their correlation with clinicopathologic features and postoperative prognosis. The study investigated the expression of TILs using immunohistochemistry of tissue microarray samples for 199 CRC patients. The number of each T-cell subtype infiltrating tumors was counted using ImageJ software. The relationship between TIL marker expression, clinicopathologic features, and prognosis was analyzed. A high RORγT/CD3 ratio (Th17 ratio) was significantly correlated with lymph node metastasis (p = 0.002), and a high of Foxp3/CD3 ratio (Treg ratio) was correlated with tumor location in the colon (p = 0.04), as shown by the Chi square test. In multivariate analysis, a high RORγT/CD3 ratio was the only independent prognostic factor for overall survival (p = 0.04; hazard ratio [HR], 1.84; 95% confidence interval [CI] 1.02-3.45). This study confirmed a high RORγT/CD3 ratio as a strong prognostic marker for postoperative survival. The immunohistochemistry results suggest that Th17 may affect lymph node metastasis in CRC. If new immunotherapies reducing Th17 expression are established, they may improve the efficiency of cancer treatment and prolong the survival of patients with CRC.

Feasibility of a novel one-stop ISET device to capture CTCs and its clinical application

PubMed Central

Zheng, Liang; Zhi, Xuan; Cheng, Boran; Chen, Yuanyuan; Zhang, Chunxiao; Shi, Dongdong; Song, Haibin; Cai, Congli; Zhou, Pengfei; Xiong, Bin

2017-01-01

Introduction Circulating tumor cells (CTCs) play a crucial role in cancer metastasis. In this study, we introduced a novel isolation method by size of epithelial tumor cells (ISET) device with automatic isolation and staining procedure, named one-stop ISET (osISET) and validated its feasibility to capture CTCs from cancer patients. Moreover, we aim to investigate the correlation between clinicopathologic features and CTCs in colorectal cancer (CRC) in order to explore its clinical application. Results The capture efficiency ranged from 80.3% to 88% with tumor cells spiked into medium while 67% to 78.3% with tumor cells spiked into healthy donors’ blood. In detection blood samples of 72 CRC patients, CTCs and clusters of circulating tumor cells (CTC-clusters) were detected with a positive rate of 52.8% (38/72) and 18.1% (13/72) respectively. Moreover, CTC positive rate was associated with factors of lymphatic or venous invasion, tumor depth, lymph node metastasis and TNM stage in CRC patients (p < 0.01). Lymphocyte count and neutrophil to lymphocyte ratio (NLR) were significantly different between CTC positive and negative groups (p < 0.01). Materials and Methods The capture efficiency of the device was tested by spiking cancer cells (MCF-7, A549, SW480, Hela) into medium or blood samples of healthy donors. Blood samples of 72 CRC patients were detected by osISET device. The clinicopathologic characteristics of 72 CRC patients were collected and the association with CTC positive rate or CTC count were analyzed. Conclusions Our osISET device was feasible to capture and identify CTCs and CTC-clusters from cancer patients. In addition, our device holds a potential for application in cancer management. PMID:27935872

Epigenetic changes in histone acetylation underpin resistance to the topoisomerase I inhibitor irinotecan

PubMed Central

Meisenberg, Cornelia; Ashour, Mohamed E.; El-Shafie, Lamia; Liao, Chunyan; Hodgson, Adam; Pilborough, Alice; Khurram, Syed A.; Downs, Jessica A.; Ward, Simon E.

2017-01-01

Abstract The topoisomerase I (TOP1) inhibitor irinotecan triggers cell death by trapping TOP1 on DNA, generating cytotoxic protein-linked DNA breaks (PDBs). Despite its wide application in a variety of solid tumors, the mechanisms of cancer cell resistance to irinotecan remains poorly understood. Here, we generated colorectal cancer (CRC) cell models for irinotecan resistance and report that resistance is neither due to downregulation of the main cellular target of irinotecan TOP1 nor upregulation of the key TOP1 PDB repair factor TDP1. Instead, the faster repair of PDBs underlies resistance, which is associated with perturbed histone H4K16 acetylation. Subsequent treatment of irinotecan-resistant, but not parental, CRC cells with histone deacetylase (HDAC) inhibitors can effectively overcome resistance. Immunohistochemical analyses of CRC tissues further corroborate the importance of histone H4K16 acetylation in CRC. Finally, the resistant clones exhibit cross-resistance with oxaliplatin but not with ionising radiation or 5-fluoruracil, suggesting that the latter two could be employed following loss of irinotecan response. These findings identify perturbed chromatin acetylation in irinotecan resistance and establish HDAC inhibitors as potential therapeutic means to overcome resistance. PMID:28180300

Epigenetic silencing of ADAMTS5 is associated with increased invasiveness and poor survival in patients with colorectal cancer.

PubMed

Li, Jizhen; Liao, Yi; Huang, Jintuan; Sun, Yi; Chen, Hao; Chen, Chunyu; Li, Senmao; Yang, Zuli

2018-02-01

A disintegrin and metalloprotease with motif 5(ADAMTS5) has been involved in colorectal cancer (CRC) with hypermethylation in the promoter. However, its role in CRC remains unclear. The aim of this study was to explore the clinical significance and biological effect of ADAMTS5 on colorectal carcinogenesis. Through MSP, qRT-PCR, WB and IHC analysis, followed by a variety of in vitro assays, we report the function of ADAMTS5 in CRC. ADAMTS5 was markedly hypermethylaed and downregulated in tumor tissues compared with non-tumor tissues (p < 0.001). Negative expression of ADAMTS5 was much more common in tumor tissues than that in normal tissues (p < 0.001) and correlated with histologic types (p = 0.002), poor OS (p = 0.029) and DFS (p = 0.018). In vitro assay revealed that overexpression of ADAMTS5 inhibited the capabilities of migration and invasion of CRC cells, and no effect on cell growth, cell cycle and apoptosis. ADAMTS5 is hypermethylated and inhibits cancer cells invasion and migration in colorectal cancer, and correlates with OS and DFS, indicating that ADAMTS5 might be a useful biomarker in colorectal cancer therapy.

Dragon (RGMb) induces oxaliplatin resistance in colon cancer cells.

PubMed

Shi, Ying; Huang, Xiao-Xiao; Chen, Guo-Bin; Wang, Ying; Zhi, Qiang; Liu, Yuan-Sheng; Wu, Xiao-Ling; Wang, Li-Fen; Yang, Bing; Xiao, Chuan-Xing; Xing, Hui-Qin; Ren, Jian-Lin; Xia, Yin; Guleng, Bayasi

2016-07-26

Colorectal cancer (CRC) is one of the most commonly diagnosed cancers and a major cause of cancer mortality. Chemotherapy resistance remains a major challenge for treating advanced CRC. Therefore, the identification of targets that induce drug resistance is a priority for the development of novel agents to overcome resistance. Dragon (also known as RGMb) is a member of the repulsive guidance molecule (RGM) family. We previously showed that Dragon expression increases with CRC progression in human patients. In the present study, we found that Dragon inhibited apoptosis and increased viability of CMT93 and HCT116 cells in the presence of oxaliplatin. Dragon induced resistance of xenograft tumor to oxaliplatinin treatment in mice. Mechanistically, Dragon inhibited oxaliplatin-induced JNK and p38 MAPK activation, and caspase-3 and PARP cleavages. Our results indicate that Dragon may be a novel target that induces drug resistance in CRC.

Dragon (RGMb) induces oxaliplatin resistance in colon cancer cells

PubMed Central

Wang, Ying; Zhi, Qiang; Liu, Yuan-Sheng; Wu, Xiao-Ling; Wang, Li-Fen; Yang, Bing; Xiao, Chuan-Xing; Xing, Hui-Qin; Ren, Jian-Lin; Xia, Yin; Guleng, Bayasi

2016-01-01

Colorectal cancer (CRC) is one of the most commonly diagnosed cancers and a major cause of cancer mortality. Chemotherapy resistance remains a major challenge for treating advanced CRC. Therefore, the identification of targets that induce drug resistance is a priority for the development of novel agents to overcome resistance. Dragon (also known as RGMb) is a member of the repulsive guidance molecule (RGM) family. We previously showed that Dragon expression increases with CRC progression in human patients. In the present study, we found that Dragon inhibited apoptosis and increased viability of CMT93 and HCT116 cells in the presence of oxaliplatin. Dragon induced resistance of xenograft tumor to oxaliplatinin treatment in mice. Mechanistically, Dragon inhibited oxaliplatin-induced JNK and p38 MAPK activation, and caspase-3 and PARP cleavages. Our results indicate that Dragon may be a novel target that induces drug resistance in CRC. PMID:27384995

The regulation of host cellular and gut microbial metabolism in the development and prevention of colorectal cancer.

PubMed

Zhou, Cheng-Bei; Fang, Jing-Yuan

2018-01-23

Metabolism regulation is crucial in colorectal cancer (CRC) and has emerged as a remarkable field currently. The cellular metabolism of glucose, amino acids and lipids in CRC are all reprogrammed. Each of them changes tumour microenvironment, modulates bacterial composition and activity, and eventually promotes CRC development. Metabolites such as short chain fatty acids, secondary bile acids, N-nitroso compounds, hydrogen sulphide, polyphenols and toxins like fragilysin, FadA, cytolethal distending toxin and colibactin play a dual role in CRC. The relationship of gut microbe-metabolite is essential in remodelling intestinal microbial ecology composition and metabolic activity. It regulates the metabolism of colonic epithelial cells and changes the tumour microenvironment in CRC. Microbial metabolism manipulation has been considered to be potentially preventive in CRC, but more large-scale clinical trials are required before their application in clinical practice in the near future.

Preparation of corrosion-resistant and conductive trivalent Cr-C coatings on 304 stainless steel for use as bipolar plates in proton exchange membrane fuel cells by electrodeposition

NASA Astrophysics Data System (ADS)

Wang, Hsiang-Cheng; Sheu, Hung-Hua; Lu, Chen-En; Hou, Kung-Hsu; Ger, Ming-Der

2015-10-01

In this study, Cr-C-coated bipolar plates are produced by electroplating on the SS304 plates with a machined flow channel. The resulting plates were tested using potentiodynamic and potentiostatic measurements in simulated PEMFC environments, which show that the bipolar plate coated with Cr-C exhibited good anticorrosion performance. The corrosive current density of the Cr-C coating formed for a plating time of 10 min for 10 h exhibits a low stable value of 1.51 × 10-10 A/cm2 during the potentiostatic test in a 0.5 M H2SO4 + 2 ppm HF solution at 70 °C with an air purge, indicating that the Cr-C coating plated for 10 min is stable in a cathode environment. The interfacial contact resistance (ICR) of the bipolar plate with the Cr-C coating clearly improved, presenting an ICR of 19.52 mΩ cm2 at a pressure of 138 N/cm2. The results from scanning electron microscopy (SEM) and ICR before and after the corrosion tests indicate that the bipolar plate with the Cr-C coating is electrochemically stable. In this study, the maximum power density (212.41 mW/cm2) is obtained at a cell temperature of 80 °C and a gas flow rate of 300 standard cubic centimeters per minute (sccm) when Cr-C coated SS304 bipolar plates were used.

Cancer testis antigen OY-TES-1 expression and serum immunogenicity in colorectal cancer: its relationship to clinicopathological parameters.

PubMed

Luo, Bin; Yun, Xiang; Fan, Rong; Lin, Yong-Da; He, Shu-Jia; Zhang, Qing-Mei; Mo, Fa-Rong; Chen, Fang; Xiao, Shao-Wen; Xie, Xiao-Xun

2013-01-01

Cancer testis (CT) antigens are attractive targets for cancer immunotherapy because their expression is restricted in normal germ line tissues but frequently detected in variety of tumors. OY-TES-1 is identified as a member of CT antigens. Current knowledge about OY-TES-1 expression in colorectal cancer (CRC) is solely based on mRNA analysis. None of previous researches has studied OY-TES-1 at protein level. In this study, OY-TES-1 polyclonal antibody was generated. The expression of OY-TES-1 mRNA and protein was detected by RT-PCR and immunohistochemistry in 60 CRC and paired adjacent non-tumor tissues, 24 colorectal adenoma and 3 normal colon tissues, respectively. Sera from 73 CRC patients were also tested for OY-TES-1 antibody by ELISA. Our results showed that the frequency of OY-TES-1 mRNA expression was statistically higher in CRC (73.3%, 44/60) than that in adjacent non-tumor tissue (55.0%, 33/60) and colorectal adenoma (45.8%, 11/24). For the first time, OY-TES-1 protein expression was found in (43.3%, 26/60) of CRC tissues, but absent in any of adjacent non-tumor and colorectal adenoma tissues. No OY-TES-1 expression was found in normal colon by either RT-PCR or immunohistochemistry. Furthermore, OY-TES-1 protein expression was correlated with tumor invasion stage (P=0.004) and histological grade (P=0.040). Anti-OY-TES-1 antibody was detected in (9.6%, 7/73) of CRC patients' sera but not in 76 healthy donors. This finding demonstrates that OY-TES-1 is frequently expressed in CRC and is able to induce humoral immune response spontaneously in CRC patients, suggesting that it might be a promising immunotherapy target for CRC.

Cyclooxygenase-2/carbonic anhydrase-IX up-regulation promotes invasive potential and hypoxia survival in colorectal cancer cells

PubMed Central

Sansone, Pasquale; Piazzi, Giulia; Paterini, Paola; Strillacci, Antonio; Ceccarelli, Claudio; Minni, Francesco; Biasco, Guido; Chieco, Pasquale; Bonafè, Massimiliano

2009-01-01

Inflammation promotes colorectal carcinogenesis. Tumour growth often generates a hypoxic environment in the inner tumour mass. We here report that, in colon cancer cells, the expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2) associates with that of the hypoxia response gene carbonic anhydrase-IX (CA-IX). The COX-2 knockdown, achieved by the stable infection of a COX-2 specific short harpin RNA interference (shCOX-2), down-regulates CA-IX gene expression. In colorectal cancer (CRC) cells, PGE2, the main COX-2 gene products, promotes CA-IX gene expression by ERK1/2 activation. In normoxic environment, shCOX-2 infected/CA-IX siRNA transfected CRC cells show a reduced level of active metalloproteinase-2 (MMP-2) that associates with a decreased extracellular matrix invasion capacity. In presence of hypoxia, COX-2 gene expression and PGE2 production increase. The knockdown of COX-2/CA-IX blunts the survival capability of CRC cells in hypoxia. At a high cell density, a culture condition that creates a mild pericellular hypoxic environment, the expression of COX-2/CA-IX genes is increased and triggers the invasive potential of colon cancer cells. In human colon cancer tissues, COX-2/CA-IX protein expression levels, assessed by Western blot and immunohistochemistry, correlate each other and increase with tumour stage. In conclusion, these data indicate that COX-2/CA-IX interplay promotes the aggressive behaviour of CRC cells. PMID:19017360

Combination epigenetic therapy in metastatic colorectal cancer (mCRC) with subcutaneous 5-azacitidine and entinostat: a phase 2 consortium/stand up 2 cancer study.

PubMed

Azad, Nilofer S; El-Khoueiry, Anthony; Yin, Jun; Oberg, Ann L; Flynn, Patrick; Adkins, Douglas; Sharma, Anup; Weisenberger, Daniel J; Brown, Thomas; Medvari, Prakriti; Jones, Peter A; Easwaran, Hariharan; Kamel, Ihab; Bahary, Nathan; Kim, George; Picus, Joel; Pitot, Henry C; Erlichman, Charles; Donehower, Ross; Shen, Hui; Laird, Peter W; Piekarz, Richard; Baylin, Stephen; Ahuja, Nita

2017-05-23

Therapy with demethylating agent 5-azacitidine and histone deacetylase inhibitor entinostat shows synergistic re-expression of tumor-suppressor genes and growth inhibition in colorectal (CRC) cell lines and in vivo studies. We conducted a phase II, multi-institutional study of the combination in metastatic CRC patients. Subcutaneous azacitidine was administered at 40 mg/m2 days 1-5 and 8-10 and entinostat was given 7 mg orally on days 3 and 10. An interim analysis indicated toxicity crossed the pre-specified safety boundary but was secondary to disease. A 2nd cohort with added eligibility restrictions was accrued: prior therapies were limited to no more than 2 or 3 (KRAS-mutated and KRAS-wildtype cancers, respectively) and <30% of liver involvement. The primary endpoint was RECIST response. Serial biopsies were performed at baseline and after 2 cycles of therapy. Forty-seven patients were enrolled (24:Cohort 1, 23:Cohort 2). Patients were heavily pre-treated (median prior therapies 4: Cohort 1 and 2.5: cohort 2). No responses were observed. Median progression-free survival was 1.9 months; overall survival was 5.6 and 8.3 months in Cohorts 1 and 2, respectively. Toxicity was tolerable and as expected. Unsupervised cluster analysis of serial tumor biopsies suggested greater DNA demethylation in patients with PFS above the median. In this first trial of CRC patients with combination epigenetic therapy, we show tolerable therapy without significant clinical activity as determined by RECIST responses. Reversal of hypermethylation was seen in a subset of patients and correlated with improved PFS.

Combination epigenetic therapy in metastatic colorectal cancer (mCRC) with subcutaneous 5-azacitidine and entinostat: a phase 2 consortium/stand Up 2 cancer study

PubMed Central

Azad, Nilofer S.; el-Khoueiry, Anthony; Yin, Jun; Oberg, Ann L.; Flynn, Patrick; Adkins, Douglas; Sharma, Anup; Weisenberger, Daniel J.; Brown, Thomas; Medvari, Prakriti; Jones, Peter A.; Easwaran, Hariharan; Kamel, Ihab; Bahary, Nathan; Kim, George; Picus, Joel; Pitot, Henry C.; Erlichman, Charles; Donehower, Ross; Shen, Hui; Laird, Peter W.; Piekarz, Richard; Baylin, Stephen; Ahuja, Nita

2017-01-01

Purpose Therapy with demethylating agent 5-azacitidine and histone deacetylase inhibitor entinostat shows synergistic re-expression of tumor-suppressor genes and growth inhibition in colorectal (CRC) cell lines and in vivo studies. Experimental Design We conducted a phase II, multi-institutional study of the combination in metastatic CRC patients. Subcutaneous azacitidine was administered at 40 mg/m2 days 1-5 and 8-10 and entinostat was given 7 mg orally on days 3 and 10. An interim analysis indicated toxicity crossed the pre-specified safety boundary but was secondary to disease. A 2nd cohort with added eligibility restrictions was accrued: prior therapies were limited to no more than 2 or 3 (KRAS-mutated and KRAS-wildtype cancers, respectively) and <30% of liver involvement. The primary endpoint was RECIST response. Serial biopsies were performed at baseline and after 2 cycles of therapy. Results Forty-seven patients were enrolled (24:Cohort 1, 23:Cohort 2). Patients were heavily pre-treated (median prior therapies 4: Cohort 1 and 2.5: cohort 2). No responses were observed. Median progression-free survival was 1.9 months; overall survival was 5.6 and 8.3 months in Cohorts 1 and 2, respectively. Toxicity was tolerable and as expected. Unsupervised cluster analysis of serial tumor biopsies suggested greater DNA demethylation in patients with PFS above the median. Conclusion In this first trial of CRC patients with combination epigenetic therapy, we show tolerable therapy without significant clinical activity as determined by RECIST responses. Reversal of hypermethylation was seen in a subset of patients and correlated with improved PFS. PMID:28186961

MicroRNA-409-3p suppresses colorectal cancer invasion and metastasis partly by targeting GAB1 expression.

PubMed

Bai, Rongpan; Weng, Chunhua; Dong, Haojie; Li, Siqi; Chen, Guangdi; Xu, Zhengping

2015-11-15

Colorectal cancer (CRC) is one of the most common cancers worldwide and its metastasis accounts for the majority of deaths. However, the molecular mechanisms underlying CRC progression are not well characterized. In this study, we identified miR-409-3p as a tumor suppressor of CRC. MiR-409-3p expression was significantly downregulated in CRC tissue compared to adjacent non-tumor tissue, and reduced miR-409-3p expression was correlated with CRC metastasis. In vitro and in vivo studies revealed that miR-409-3p negatively regulated CRC metastatic capacities, including suppressing cancer cell migration, invasion and metastasis. To explore the mechanism of action of miR-409-3p, we adopted a pathway and pathophysiological event-based target screening and validation approach, and found nine known metastasis-related genes as potential targets. The 3'-UTR binding assays between the candidates and miR-409-3p suggested that only GAB1, NR4A2 and LMO4 were directly regulated by the miRNA. However, endogenous expression analysis revealed that only GAB1 was modulated by miR-409-3p in CRC cells at both the mRNA and protein levels. Furthermore, we provided evidence to conclude that GAB1 was partially responsible for miR-409-3p-mediated metastasis. Taken together, our data demonstrate that miR-409-3p is a metastatic suppressor, and post-transcriptional inhibition of the oncoprotein GAB1 is one of the mechanisms of action of this miRNA. Our finding suggests miR-409-3p might be a novel target for CRC metastasis treatment. © 2015 UICC.

Global-scale profiling of differential expressed lysine acetylated proteins in colorectal cancer tumors and paired liver metastases.

PubMed

Shen, Zhanlong; Wang, Bo; Luo, Jianyuan; Jiang, Kewei; Zhang, Hui; Mustonen, Harri; Puolakkainen, Pauli; Zhu, Jun; Ye, Yingjiang; Wang, Shan

2016-06-16

Lysine acetylated modification was indicated to impact colorectal cancer (CRC)'s distant metastasis. However, the global acetylated proteins in CRC and the differential expressed acetylated proteins and acetylated sites between CRC primary and distant metastatic tumor remains unclear. Our aim was to construct a complete atlas of acetylome in CRC and paired liver metastases. Combining high affinity enrichment of acetylated peptides with high sensitive mass spectrometry, we identified 603 acetylation sites from 316 proteins, among which 462 acetylation sites corresponding to 243 proteins were quantified. We further classified them into groups according to cell component, molecular function and biological process and analyzed the metabolic pathways, domain structures and protein interaction networks. Finally, we evaluated the differentially expressed lysine acetylation sites and revealed that 31 acetylated sites of 22 proteins were downregulated in CRC liver metastases compared to that in primary CRC while 40 acetylated sites of 32 proteins were upregulated, of which HIST2H3AK19Ac and H2BLK121Ac were the acetylated histones most changed, while TPM2 K152Ac and ADH1B K331Ac were the acetylated non-histones most altered. These results provide an expanded understanding of acetylome in CRC and its distant metastasis, and might prove applicable in the molecular targeted therapy of metastatic CRC. This study described provides, for the first time, that full-scale profiling of lysine acetylated proteins were identified and quantified in colorectal cancer (CRC) and paired liver metastases. The novelty of the study is that we constructed a complete atlas of acetylome in CRC and paired liver metastases. Moreover, we analyzed these differentially expressed acetylated proteins in cell component, molecular function and biological process. In addition, metabolic pathways, domain structures and protein interaction networks of acetylated proteins were also investigated. Our approaches shows that of the differentially expressed proteins, HIST2H3AK19Ac and H2BLK121Ac were the acetylated histones most changed, while TPM2 K152Ac and ADH1B K331Ac were the acetylated non-histones most altered. Our findings provide an expanded understanding of acetylome in CRC and its distant metastasis, and might prove applicable in the molecular targeted therapy of metastatic CRC. Copyright © 2016 Elsevier B.V. All rights reserved.

Geographic distribution of the incidence of colorectal cancer in Iran: a population-based study.

PubMed

Khosravi Shadmani, Fatemeh; Ayubi, Erfan; Khazaei, Salman; Sani, Mohadeseh; Mansouri Hanis, Shiva; Khazaei, Somayeh; Soheylizad, Mokhtar; Mansori, Kamyar

2017-01-01

Colorectal cancer (CRC) is the third most common cancer and the fourth most common cause of cancer death in the world. The aim of this study was to investigate the provincial distribution of the incidence of CRC across Iran. This epidemiologic study used data from the National Cancer Registry of Iran and the Center for Disease Control and Prevention of the Ministry of Health and Medical Education of Iran. The average annual age-standardized rate (ASR) for the incidence of CRC was calculated for each province. We found that adenocarcinoma (not otherwise specified) was the most common histological subtype of CRC in males and females, accounting for 81.91 and 81.95% of CRC cases, respectively. Signet ring cell carcinoma was the least prevalent subtype of CRC in males and females and accounted for 1.5 and 0.94% of CRC cases, respectively. In patients aged 45 years or older, there was a steady upward trend in the incidence of CRC, and the highest ASR of CRC incidence among both males and females was in the age group of 80-84 years, with an ASR of 144.69 per 100,000 person-years for males and 119.18 per 100,000 person-years for females. The highest incidence rates of CRC in Iran were found in the central, northern, and western provinces. Provinces in the southeast of Iran had the lowest incidence rates of CRC. Wide geographical variation was found in the incidence of CRC across the 31 provinces of Iran. These variations must be considered for prevention and control programs for CRC, as well as for resource allocation purposes.

Antitumor efficacy of triple monoclonal antibody inhibition of epidermal growth factor receptor (EGFR) with MM151 in EGFR-dependent and in cetuximab-resistant human colorectal cancer cells

PubMed Central

Napolitano, Stefania; Martini, Giulia; Martinelli, Erika; Della Corte, Carminia Maria; Morgillo, Floriana; Belli, Valentina; Cardone, Claudia; Matrone, Nunzia; Ciardiello, Fortunato; Troiani, Teresa

2017-01-01

Purpose We investigated the effect of triple monoclonal antibody inhibition of EGFR to overcome acquired resistance to first generation of anti-EGFR inhibitors. Experimental design MM151 is a mixture of three different monoclonal IgG1 antibodies directed toward three different, non-overlapping, epitopes of the EGFR. We performed an in vivo study by using human CRC cell lines (SW48, LIM 1215 and CACO2) which are sensitive to EGFR inhibitors, in order to evaluate the activity of MM151 as compared to standard anti-EGFR mAbs, such as cetuximab, as single agent or in a sequential strategy of combination MM151 with irinotecan (induction therapy) followed by MM151 with a selective MEK1/2 inhibitor (MEKi) (maintenance therapy). Furthermore, the ability of MM151 to overcome acquired resistance to cetuximab has been also evaluated in cetuximab-refractory CRC models. Results MM151 shown stronger antitumor activity as compared to cetuximab. The maintenance treatment with MM151 plus MEKi resulted the most effective therapeutic modality. In fact, this combination caused an almost complete suppression of tumor growth in SW48, LIM 1215 and CACO2 xenografts model at 30 week. Moreover, in this treatment group, mice with no evidence of tumor were more than double as compared to single agent treated mice. Its superior activity has also been demonstrated, in cetuximab-refractory CRC models. Conclusions These results provide experimental evidence that more efficient and complete EGFR blockade may determine better antitumor activity and could contribute to prevent and/or overcome acquired resistance to EGFR inhibitors. PMID:29137301

Precision Medicine for CRC Patients in the Veteran Population: State-of-the-Art, Challenges and Research Directions.

PubMed

Mohapatra, Shyam S; Batra, Surinder K; Bharadwaj, Srinivas; Bouvet, Michael; Cosman, Bard; Goel, Ajay; Jogunoori, Wilma; Kelley, Michael J; Mishra, Lopa; Mishra, Bibhuti; Mohapatra, Subhra; Patel, Bhaumik; Pisegna, Joseph R; Raufman, Jean-Pierre; Rao, Shuyun; Roy, Hemant; Scheuner, Maren; Singh, Satish; Vidyarthi, Gitanjali; White, Jon

2018-05-01

Colorectal cancer (CRC) accounts for ~9% of all cancers in the Veteran population, a fact which has focused a great deal of the attention of the VA's research and development efforts. A field-based meeting of CRC experts was convened to discuss both challenges and opportunities in precision medicine for CRC. This group, designated as the VA Colorectal Cancer Cell-genomics Consortium (VA4C), discussed advances in CRC biology, biomarkers, and imaging for early detection and prevention. There was also a discussion of precision treatment involving fluorescence-guided surgery, targeted chemotherapies and immunotherapies, and personalized cancer treatment approaches. The overarching goal was to identify modalities that might ultimately lead to personalized cancer diagnosis and treatment. This review summarizes the findings of this VA field-based meeting, in which much of the current knowledge on CRC prescreening and treatment was discussed. It was concluded that there is a need and an opportunity to identify new targets for both the prevention of CRC and the development of effective therapies for advanced disease. Also, developing methods integrating genomic testing with tumoroid-based clinical drug response might lead to more accurate diagnosis and prognostication and more effective personalized treatment of CRC.

Ginsenoside 20(S)-Rh2 exerts anti-cancer activity through targeting IL-6-induced JAK2/STAT3 pathway in human colorectal cancer cells.

PubMed

Han, Songhee; Jeong, Ae Jin; Yang, Heejung; Bin Kang, Kyo; Lee, Haeri; Yi, Eun Hee; Kim, Byung-Hak; Cho, Chung-Hyun; Chung, Jin Woong; Sung, Sang Hyun; Ye, Sang-Kyu

2016-12-24

Panax ginseng is one of the most well-known medicinal herbs in Korea and China, which has been used for treatment and prevention of cancer, obesity, diabetes, and cardiovascular diseases. Ginsenosides are the major components of P. ginseng, having a wide range of pharmacological activities. Among the ginsenosides, protopanaxadiol (PPD)-types reportedly have potent anti-cancer effects. Rh2 is PPD-type ginsenoside, and two stereoisomeric forms of Rh2 as 20(S)- and 20(R)-Rh2 were selectively isolated recently. The biological activities of Rh2 ginsenosides are known to depend on their differences in stereochemistry. Colorectal cancer (CRC) is one of the most lethal neoplasm, and cancer-related death is usually associated with metastasis to other organs. We aimed this study to investigate whether 20(S)- and 20(R)-Rh2 can suppress tumor invasion in human CRC cells. 20(S)- and 20(R)-Rh2 were isolated from the roots of ginseng. Human CRC cells were incubated with 20(S)- or 20(R)-Rh2 in the presence or absence of interleukin-6. An MTT assay was used to measure cell viability. Western blot and quantitative real-time PCR analyses were performed to determine levels of expression and phosphorylation. An invasion assay was performed using a Boyden chamber system with the Matrigel-coated membrane to measure cancer cell invasion. 20(S)- and 20(R)-Rh2 showed differential cytotoxic activity. Only 20(S)-Rh2 decreased cancer cell viability. Additionally, 20(S)-Rh2 effectively inhibited IL-6-induced signal transducer and activator of transcription 3 (STAT3) phosphorylation and the expression of matrix metalloproteinases (MMPs), including MMP-1, -2, and -9, resulting in inhibition of cancer cell invasion. Interestingly, these pharmacological activities of 20(S)-Rh2 were more potent than those of 20(R)-Rh2. Furthermore, combination treatment showed that 20(S)-Rh2 enhanced the sensitization of doxorubicin-treated anti-cancer activities in CRC cells. Our results demonstrated that ginsenoside 20(S)-Rh2 has therapeutic potential for the treatment with CRC and may be valuable as a combination partner with more classic chemotherapeutic agents, such as doxorubicin, to treat CRC. Copyright © 2016. Published by Elsevier Ireland Ltd.

Decreased expression of miR‑490‑3p in colorectal cancer predicts poor prognosis and promotes cell proliferation and invasion by targeting RAB14.

PubMed

Wang, Bo; Yin, Mujun; Cheng, Cheng; Jiang, Hongpeng; Jiang, Kewei; Shen, Zhanlong; Ye, Yingjiang; Wang, Shan

2018-06-19

Growing evidence indicates a potential role for miR‑490‑3p in tumorigenesis. However, its function in colorectal carcinoma (CRC) remains undefined. In this study, miR‑490‑3p was markedly downregulated in fifty colorectal cancer tissue samples compared with the corresponding adjacent non‑cancerous specimens, by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The expression levels of miR‑490‑3p were closely associated with tumor differentiation and distant metastasis. In addition, both Kaplan-Meier and multivariate analyses indicated CRC patients with elevated miR‑490‑3p amounts had prolonged overall survival. Overexpression of miR‑490‑3p markedly reduced proliferation, colony formation and invasion in CRC cells by enhancing apoptosis and promoting G2/M phase arrest. Furthermore, ectopic expression of miR‑490‑3p resulted in decreased expression of RAB14, which was directly targeted by miR‑490‑3p, as shown by the dual-luciferase reporter gene assay. Finally, in a nude mouse model, miR‑490‑3p overexpression significantly suppressed the growth of CRC cells. The above results indicated that miR‑490‑3p might constitute a prognostic indicator and a novel molecular target for miRNA-based CRC therapy.

BDNF/TrkB signaling protects HT-29 human colon cancer cells from EGFR inhibition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brunetto de Farias, Caroline; Children's Cancer Institute, 90420-140 Porto Alegre, RS; Laboratory of Neuropharmacology and Neural Tumor Biology, Department of Pharmacology, Institute for Basic Health Sciences, Federal University of Rio Grande do Sul, 90050-170 Porto Alegre, RS

2012-08-24

Highlights: Black-Right-Pointing-Pointer BDNF protected HT-29 colorectal cancer cells from the antitumor effect of cetuximab. Black-Right-Pointing-Pointer TrkB inhibition potentiated the antitumor effect of cetuximab. Black-Right-Pointing-Pointer BDNF/TrkB signaling might be involved in resistance to anti-EGFR therapy. -- Abstract: The clinical success of targeted treatment of colorectal cancer (CRC) is often limited by resistance to anti-epidermal growth factor receptor (EGFR) therapy. The neurotrophin brain-derived neurotrophic factor (BDNF) and its receptor TrkB have recently emerged as anticancer targets, and we have previously shown increased BDNF levels in CRC tumor samples. Here we report the findings from in vitro experiments suggesting that BDNF/TrkB signaling canmore » protect CRC cells from the antitumor effects of EGFR blockade. The anti-EGFR monoclonal antibody cetuximab reduced both cell proliferation and the mRNA expression of BDNF and TrkB in human HT-29 CRC cells. The inhibitory effect of cetuximab on cell proliferation and survival was counteracted by the addition of human recombinant BDNF. Finally, the Trk inhibitor K252a synergistically enhanced the effect of cetuximab on cell proliferation, and this effect was blocked by BDNF. These results provide the first evidence that increased BDNF/TrkB signaling might play a role in resistance to EGFR blockade. Moreover, it is possible that targeting TrkB could potentiate the anticancer effects of anti-EGFR therapy.« less

Cost-minimization analysis of panitumumab compared with cetuximab for first-line treatment of patients with wild-type RAS metastatic colorectal cancer.

PubMed

Graham, Christopher N; Hechmati, Guy; Fakih, Marwan G; Knox, Hediyyih N; Maglinte, Gregory A; Hjelmgren, Jonas; Barber, Beth; Schwartzberg, Lee S

2015-01-01

To compare the costs of first-line treatment with panitumumab + FOLFOX in comparison to cetuximab + FOLFIRI among patients with wild-type (WT) RAS metastatic colorectal cancer (mCRC) in the US. A cost-minimization model was developed assuming similar treatment efficacy between both regimens. The model estimated the costs associated with drug acquisition, treatment administration frequency (every 2 weeks for panitumumab, weekly for cetuximab), and incidence of infusion reactions. Average anti-EGFR doses were calculated from the ASPECCT clinical trial, and average doses of chemotherapy regimens were based on product labels. Using the medical component of the consumer price index, adverse event costs were inflated to 2014 US dollars, and all other costs were reported in 2014 US dollars. The time horizon for the model was based on average first-line progression-free survival of a WT RAS patient, estimated from parametric survival analyses of PRIME clinical trial data. Relative to cetuximab + FOLFIRI in the first-line treatment of WT RAS mCRC, the cost-minimization model demonstrated lower projected drug acquisition, administration, and adverse event costs for patients who received panitumumab + FOLFOX. The overall cost per patient for first-line treatment was $179,219 for panitumumab + FOLFOX vs $202,344 for cetuximab + FOLFIRI, resulting in a per-patient saving of $23,125 (11.4%) in favor of panitumumab + FOLFOX. From a value perspective, the cost-minimization model supports panitumumab + FOLFOX instead of cetuximab + FOLFIRI as the preferred first-line treatment of WT RAS mCRC patients requiring systemic therapy.

Sodium Butyrate Inhibits Colorectal Cancer Cell Migration by Downregulating Bmi-1 Through Enhanced miR-200c Expression.

PubMed

Xu, Zhiyao; Tao, Jingjing; Chen, Ping; Chen, Long; Sharma, Sherven; Wang, Guanyu; Dong, Qinghua

2018-03-01

Short-chain fatty acid sodium butyrate (NaB) is the byproduct of bacterial anaerobic fermentation of dietary fiber in the colon, and has been shown to have an antitumor effect on colorectal cancer (CRC). The miR-200 family is a key regulator of the epithelial-mesenchymal transition (EMT). We investigate the role of miR-200s expression on cell migration in NaB-treated CRC cells. HCT116 and LOVO CRC cells treated with NaB depicted reduced cell proliferation, enhanced apoptosis, and cell cycle arrest. NaB inhibited cell migration in the wound healing and transwell assays, and in spheriod cultures while regulating EMT-related protein expression. NaB reciprocally increased miR-200s but reduced expression of their target genes (Bmi-1, Zeb1, EZH2). Cells transfected with miR-200c shRNA displayed a significant blockade of NaB-induced anti-invasive activity. Upregulation of Bmi-1 expression partially reversed the effect of NaB. In addition to inhibition of tumor growth in vivo, qRT-PCR results showed that NaB increased miR-200c/200b/492 expression in the tumor tissues. Immunohistochemistry and Western blotting results demonstrated that NaB decreased Bmi-1 expression in vivo. NaB inhibits CRC cell migration by enhancing miR-200c expression-mediated downregulation of Bmi-1. These findings support the utility of NaB in colorectal cancer therapy. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A Pilot Study of Apatinib as Third-Line Treatment in Patients With Heavily Treated Metastatic Colorectal Cancer.

PubMed

Liang, Lijun; Wang, Lei; Zhu, Panrong; Xia, Youyou; Qiao, Yun; Wu, Jiang; Zhuang, Wei; Fei, Jiayan; Wen, Yixuan; Jiang, Xiaodong

2018-03-02

Antiangiogenic therapy has shown improved clinical outcome in metastatic colorectal cancer (mCRC). After the failure of standard treatments, regorafenib and TAS-102 would be recommended for patients with mCRC, however, they have not been approved in China during this study period. This pilot study aimed to assess the efficacy and safety of apatinib, a novel oral inhibitor targeting vascular endothelial growth factor receptor 2, as third-line treatment for patients with mCRC refractory to standard therapies. In this retrospective study, all patients received apatinib treatment until progressive disease (PD), death, unacceptable toxicity, and curative surgery. The dose or treatment schedule was modified according to the physician's discretion according to the toxicity profiles. Between March 2015 and June 2017, 36 patients were enrolled and eligible for evaluation of the safety and efficacy. One patient (2.8%) achieved complete response, 3 (8.3%) achieved partial response, 24 (66.7%) achieved stable disease, and 8 (22.2%) PD. The objective response rate and the disease control rate were 11.1% (4 of 36), and 77.8% (28 of 36), respectively. Moreover, the median overall survival (OS) since the initiation of first-line treatment was 33.2 months. The median progression-free survival (PFS) and median OS from apatinib treatment were 4.8 and 10.1 months, respectively. Intergroup analysis showed that there was no significant difference in median PFS and median OS between patients who were previously treated with and without bevacizumab. The most common Grade 3 to 4 adverse reactions were hand-foot syndrome, hypertension, and proteinuria. Our results suggested that apatinib was active as a third-line treatment of refractory mCRC with a manageable tolerability profile. In addition, preliminary data suggested that the efficacy of apatinib would not be affected by previous bevacizumab treatment. Further prospective randomized controlled clinical trials are urgently needed. Copyright © 2018 Elsevier Inc. All rights reserved.

S100A8 facilitates the migration of colorectal cancer cells through regulating macrophages in the inflammatory microenvironment.

PubMed

Zha, He; Sun, Hui; Li, Xueru; Duan, Liang; Li, Aifang; Gu, Yue; Zeng, Zongyue; Zhao, Jiali; Xie, Jiaqing; Yuan, Shimei; Li, Huan; Zhou, Lan

2016-07-01

Previous studies have shown that S100 calcium-binding protein A8 (S100A8) contributes to the survival and migration of colorectal cancer (CRC) cells. However, whether S100A8 participates in the progression and metastasis of CRC via the regulation of macrophages in the tumor inflammatory microenvironment remains unknown. In this study, phorbol myristate acetate (PMA) was used to induce the differentiation of THP-1 monocytes to macrophages. MTT assay, western blot analysis, immunofluorescence staining, semi-quantitative RT-PCR (semi-PCR), quantitative real-time PCR (qPCR), Gaussia luciferase activity assay and ELISA were performed to analyze the roles and molecular mechanisms of S100A8 in the modulation of macrophages. MTT assay, flow cytometric analysis, Hoechst staining, wound healing and Transwell migration assay were used to test the effect of S100A8 on the viability and migration of CRC cells co-cultured with macrophages in the inflammatory microenvironment. We found that THP-1 monocytes were induced by PMA and differentiated to macrophages. S100A8 activated the NF-κB pathway in the macrophages and promoted the expression of miR-155 and inflammatory cytokines IL-1β and TNF-α in the inflammatory microenvironment mimicked by lipopolysaccharides (LPS). Furthermore, S100A8 contributed to augment the migration but not the viability of the CRC cells co-cultured with the macrophages in the inflammatory microenvironment. Altogether, our study demonstrated that S100A8 facilitated the migration of CRC cells in the inflammatory microenvironment, and the underlying molecular mechanisms may be partially attributed to the overexpression of miR-155, IL-1β and TNF-α through activation of the NF-κB pathway in macrophages.

FBXW7-mutated colorectal cancer cells exhibit aberrant expression of phosphorylated-p53 at Serine-15

PubMed Central

Normatova, Makhliyo; Babaei-Jadidi, Roya; Tomlinson, Ian; Nateri, Abdolrahman S.

2015-01-01

FBXW7 mutations occur in a variety of human cancers including colorectal cancer (CRC). Elucidating its mechanism of action has become crucial for cancer therapy; however, it is also complicated by the fact that FBXW7 can influence many pathways due to its role as an E3-ubiquitin ligase in proteasome degradation. FBXW7 and TP53 are tumour suppressors intensively implicated in colorectal carcinogenesis. Deletion mutations in these two genes in animal models mark the progression from adenoma to carcinoma. Although still largely unknown, the last defense mechanism against CRC at the molecular level could be through a synergistic effect of the two genes. The underlying mechanism requires further investigation. In our laboratory, we have used a phospho-kinase profiler array to illustrate a potential molecular link between FBXW7 and p53 in CRC cells. In vitro and in vivo assessments demonstrated aberrant induction of phosphorylated p53 at Serine 15 [phospho-p53(Ser15)] in human FBXW7-deficient CRC cells as compared to their FBXW7-wild-type counterparts. FBXW7 loss in HCT116 cells promoted resistance to oxaliplatin. Immunoblotting data further confirmed that reduction of phospho-p53(Ser15) may contribute to the decreased efficacy of therapy in FBXW7-mutated CRC cells. The findings may suggest the applicability of phospho-p53(Ser15) as an indicative marker of FBXW7-mutations. Phospho-p53(Ser15) regulation by FBXW7 E3-ligase activity could provide important clues for understanding FBXW7 behavior in tumour progression and grounds for its clinical applicability thereafter. PMID:25860929

DEAD-box helicase 27 promotes colorectal cancer growth and metastasis and predicts poor survival in CRC patients.

PubMed

Tang, Jieting; Chen, Huarong; Wong, Chi-Chun; Liu, Dabin; Li, Tong; Wang, Xiaohong; Ji, Jiafu; Sung, Joseph Jy; Fang, Jing-Yuan; Yu, Jun

2018-03-14

Copy number alterations (CNAs) are crucial for colorectal cancer (CRC) development. In this study, DEAD box polypeptide 27 (DDX27) was identified to be highly amplified in both TCGA CRC (474/615) and primary CRC (47/103), which was positively correlated with its mRNA overexpression. High DDX27 mRNA (N = 199) and protein expression (N = 260) predicted poor survival in CRC patients. Ectopic expression of DDX27 increased CRC cells proliferation, migration and invasion, but suppressed apoptosis. Conversely, silencing of DDX27 exerted opposite effects in vitro and significantly inhibited murine xenograft tumor growth and lung metastasis in vivo. Up-regulation of DDX27 enhanced and prolonged TNF-α-mediated NF-κB signaling. Nucleophosmin (NPM1) was identified as a binding partner of DDX27. DDX27 increased nuclear NPM1 and NF-κB-p65 interaction to enhance DNA binding activity of NF-κB. Silencing NPM1 abrogated DDX27-activating NF-κB signaling and its tumor-promoting function. Together, DDX27 is overexpressed and plays a pivotal oncogenic role in CRC.

Protein kinase C zeta suppresses low‐ or high‐grade colorectal cancer (CRC) phenotypes by interphase centrosome anchoring

PubMed Central

Deevi, Ravi Kiran; Javadi, Arman; McClements, Jane; Vohhodina, Jekaterina; Savage, Kienan; Loughrey, Maurice Bernard; Evergren, Emma

2018-01-01

Abstract Histological grading provides prognostic stratification of colorectal cancer (CRC) by scoring heterogeneous phenotypes. Features of aggressiveness include aberrant mitotic spindle configurations, chromosomal breakage, and bizarre multicellular morphology, but pathobiology is poorly understood. Protein kinase C zeta (PKCz) controls mitotic spindle dynamics, chromosome segregation, and multicellular patterns, but its role in CRC phenotype evolution remains unclear. Here, we show that PKCz couples genome segregation to multicellular morphology through control of interphase centrosome anchoring. PKCz regulates interdependent processes that control centrosome positioning. Among these, interaction between the cytoskeletal linker protein ezrin and its binding partner NHERF1 promotes the formation of a localized cue for anchoring interphase centrosomes to the cell cortex. Perturbation of these phenomena induced different outcomes in cells with single or extra centrosomes. Defective anchoring of a single centrosome promoted bipolar spindle misorientation, multi‐lumen formation, and aberrant epithelial stratification. Collectively, these disturbances induce cribriform multicellular morphology that is typical of some categories of low‐grade CRC. By contrast, defective anchoring of extra centrosomes promoted multipolar spindle formation, chromosomal instability (CIN), disruption of glandular morphology, and cell outgrowth across the extracellular matrix interface characteristic of aggressive, high‐grade CRC. Because PKCz enhances apical NHERF1 intensity in 3D epithelial cultures, we used an immunohistochemical (IHC) assay of apical NHERF1 intensity as an indirect readout of PKCz activity in translational studies. We show that apical NHERF1 IHC intensity is inversely associated with multipolar spindle frequency and high‐grade morphology in formalin‐fixed human CRC samples. To conclude, defective PKCz control of interphase centrosome anchoring may underlie distinct categories of mitotic slippage that shape the development of low‐ or high‐grade CRC phenotypes. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. PMID:29520890

Protein kinase C zeta suppresses low- or high-grade colorectal cancer (CRC) phenotypes by interphase centrosome anchoring.

PubMed

Deevi, Ravi Kiran; Javadi, Arman; McClements, Jane; Vohhodina, Jekaterina; Savage, Kienan; Loughrey, Maurice Bernard; Evergren, Emma; Campbell, Frederick Charles

2018-04-01

Histological grading provides prognostic stratification of colorectal cancer (CRC) by scoring heterogeneous phenotypes. Features of aggressiveness include aberrant mitotic spindle configurations, chromosomal breakage, and bizarre multicellular morphology, but pathobiology is poorly understood. Protein kinase C zeta (PKCz) controls mitotic spindle dynamics, chromosome segregation, and multicellular patterns, but its role in CRC phenotype evolution remains unclear. Here, we show that PKCz couples genome segregation to multicellular morphology through control of interphase centrosome anchoring. PKCz regulates interdependent processes that control centrosome positioning. Among these, interaction between the cytoskeletal linker protein ezrin and its binding partner NHERF1 promotes the formation of a localized cue for anchoring interphase centrosomes to the cell cortex. Perturbation of these phenomena induced different outcomes in cells with single or extra centrosomes. Defective anchoring of a single centrosome promoted bipolar spindle misorientation, multi-lumen formation, and aberrant epithelial stratification. Collectively, these disturbances induce cribriform multicellular morphology that is typical of some categories of low-grade CRC. By contrast, defective anchoring of extra centrosomes promoted multipolar spindle formation, chromosomal instability (CIN), disruption of glandular morphology, and cell outgrowth across the extracellular matrix interface characteristic of aggressive, high-grade CRC. Because PKCz enhances apical NHERF1 intensity in 3D epithelial cultures, we used an immunohistochemical (IHC) assay of apical NHERF1 intensity as an indirect readout of PKCz activity in translational studies. We show that apical NHERF1 IHC intensity is inversely associated with multipolar spindle frequency and high-grade morphology in formalin-fixed human CRC samples. To conclude, defective PKCz control of interphase centrosome anchoring may underlie distinct categories of mitotic slippage that shape the development of low- or high-grade CRC phenotypes. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Optimization of routine KRAS mutation PCR-based testing procedure for rational individualized first-line-targeted therapy selection in metastatic colorectal cancer.

PubMed

Chretien, Anne-Sophie; Harlé, Alexandre; Meyer-Lefebvre, Magali; Rouyer, Marie; Husson, Marie; Ramacci, Carole; Harter, Valentin; Genin, Pascal; Leroux, Agnès; Merlin, Jean-Louis

2013-02-01

KRAS mutation detection represents a crucial issue in metastatic colorectal cancer (mCRC). The optimization of KRAS mutation detection delay enabling rational prescription of first-line treatment in mCRC including anti-EGFR-targeted therapy requires robust and rapid molecular biology techniques. Routine analysis of mutations in codons 12 and 13 on 674 paraffin-embedded tissue specimens of mCRC has been performed for KRAS mutations detection using three molecular biology techniques, that is, high-resolution melting (HRM), polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP), and allelic discrimination PCR (TaqMan PCR). Discordant cases were assessed with COBAS 4800 KRAS CE-IVD assay. Among the 674 tumor specimens, 1.5% (10/674) had excessive DNA degradation and could not be analyzed. KRAS mutations were detected in 38.0% (256/674) of the analysable specimens (82.4% in codon 12 and 17.6% in codon 13). Among 613 specimens in whom all three techniques were used, 12 (2.0%) cases of discordance between the three techniques were observed. 83.3% (10/12) of the discordances were due to PCR-RFLP as confirmed by COBAS 4800 retrospective analysis. The three techniques were statistically comparable (κ > 0.9; P < 0.001). From these results, optimization of the routine procedure consisted of proceeding to systematic KRAS detection using HRM and TaqMan and PCR-RFLP in case of discordance and allowed significant decrease in delays. The results showed an excellent correlation between the three techniques. Using HRM and TaqMan warrants high-quality and rapid-routine KRAS mutation detection in paraffin-embedded tumor specimens. The new procedure allowed a significant decrease in delays for reporting results, enabling rational prescription of first-line-targeted therapy in mCRC.

The effects of selected drugs and dietary compounds on proliferation and apoptosis in colorectal carcinoma.

PubMed

Kiedrowski, Miroslaw; Mroz, Andrzej

2014-01-01

Like many malignancies, the development of colorectal carcinoma (CRC) can be considered as an imbalance between the compromised process of programmed cell death (apoptosis) and excessive, uncontrolled proliferation. Several mutations and epigenetic alterations are acquired during colorectal carcinogenesis. These are responsible for the cell cycle regulation, cellular sensitivity to pro- and antiapoptotic factors, cell proliferation, angiogenesis, invasiveness, as well as metastatic potential. The molecular alterations, along with their morphological expressions, have been recognised in detail, and most of the CRC cases can be attributed to either adenoma-carcinoma or serrated neoplasia pathways: in the first, the antiapoptotic features prevail; while in the second, the proliferative activity is of the utmost importance. The aim of the work is to discuss the influence of selected drugs and dietary compounds on the proliferation and apoptosis in CRC.

The global regulator Crc plays a multifaceted role in modulation of type III secretion system in Pseudomonas aeruginosa.

PubMed

Dong, Yi-Hu; Zhang, Xi-Fen; Zhang, Lian-Hui

2013-02-01

The opportunistic pathogen Pseudomonas aeruginosa utilizes type III secretion system (T3SS) to translocate effector proteins into eukaryotic host cells that subvert normal host cell functions to the benefit of the pathogen, and results in serious infections. T3SS in P. aeruginosa is controlled by a complex system of regulatory mechanisms and signaling pathways. In this study, we described that Crc, an RNA-binding protein, exerts a positive impact on T3SS in P. aeruginosa, as evidenced by promoter activity assays of several key T3SS genes, transcriptomics, RT-PCR, and immunoblotting in crc mutant. We further demonstrated that the regulatory function of Crc on the T3SS was mediated through the T3SS master regulator ExsA and linked to the Cbr/Crc signaling system. Expression profiling of the crc mutant revealed a downregulation of flagship T3SS genes as well as 16 other genes known to regulate T3SS gene expression in P. aeruginosa. On the basis of these data, we proposed that Crc may exert multifaceted control on the T3SS through various pathways, which may serve to fine-tune this virulence mechanism in response to environmental changes and nutrient sources. © 2012 The Authors. Published by Blackwell Publishing Ltd. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

Prevention and Treatment of Colorectal Cancer by Natural Agents From Mother Nature

PubMed Central

Aggarwal, Bharat; Prasad, Sahdeo; Sung, Bokyung; Krishnan, Sunil; Guha, Sushovan

2013-01-01

Colorectal cancer (CRC) is the third most commonly diagnosed cancer in the United States after cancers of the lung and the breast/prostate. While the incidence of CRC in the United States is among the highest in the world (approximately 52/100,000), its incidence in countries in India is among the lowest (approximately 7/100,000), suggesting that lifestyle factors may play a role in development of the disease. Whereas obesity, excessive alcohol consumption, a high-calorie diet, and a lack of physical activity promote this cancer, evidence indicates that foods containing folates, selenium, Vitamin D, dietary fiber, garlic, milk, calcium, spices, vegetables, and fruits are protective against CRC in humans. Numerous agents from “mother nature” (also called “nutraceuticals,”) that have potential to both prevent and treat CRC have been identified. The most significant discoveries relate to compounds such as cardamonin, celastrol, curcumin, deguelin, diosgenin, thymoquinone, tocotrienol, ursolic acid, and zerumbone. Unlike pharmaceutical drugs, these agents modulate multiple targets, including transcription factors, growth factors, tumor cell survival factors, inflammatory pathways, and invasion and angiogenesis linked closely to CRC. We describe the potential of these dietary agents to suppress the growth of human CRC cells in culture and to inhibit tumor growth in animal models. We also describe clinical trials in which these agents have been tested for efficacy in humans. Because of their safety and affordability, these nutraceuticals provide a novel opportunity for treatment of CRC, an “old age” disease with an “age old” solution. PMID:23814530

Use of early tumor shrinkage to predict long-term outcome in metastatic colorectal cancer treated with cetuximab.

PubMed

Piessevaux, Hubert; Buyse, Marc; Schlichting, Michael; Van Cutsem, Eric; Bokemeyer, Carsten; Heeger, Steffen; Tejpar, Sabine

2013-10-20

Early tumor shrinkage (ETS) is associated with long-term outcome in patients with chemorefractory metastatic colorectal cancer (mCRC) receiving cetuximab. This association was investigated in the first-line setting in the randomized CRYSTAL and OPUS mCRC trials, after controlling for KRAS tumor mutation status. Radiologic assessments at week 8 were used to calculate the relative change in the sum of the longest diameters of the target lesions. Time-dependent receiver operating characteristics provided Cτ-indices (time-dependent c-index). Cox regression models and subpopulation treatment effect pattern plot analysis investigated associations between ETS (radiologic tumor size decrease at week 8) and survival and progression-free survival (PFS). In both trials, in patients with KRAS wild-type mCRC, Cτ values for PFS and survival were higher (P < .001) in those receiving chemotherapy plus cetuximab versus chemotherapy alone, indicating a stronger predictive value of ETS for long-term outcome in these patients. In the CRYSTAL and OPUS trials, respectively, the cutoff value of ETS ≥ 20% (v < 20%) identified patients with KRAS wild-type mCRC receiving chemotherapy plus cetuximab with longer PFS (medians 14.1 v 7.3 months, hazard ratio [HR] = 0.32; P < .001, and medians 11.9 v 5.7 months, HR = 0.22; P < .001) and survival (medians 30.0 v 18.6 months, HR = 0.53; P < .001 and medians 26.0 v 15.7 months, HR = 0.43; P = .006). ETS was significantly associated with long-term outcome in patients with KRAS wild-type mCRC treated first-line with chemotherapy plus cetuximab. Validation in prospective trials is required to assess the value of this on-treatment marker in the clinical decision-making process.

Effect of CRC::etr1-1 transgene expression on ethylene production, sex expression, fruit set and fruit ripening in transgenic melon (Cucumis melo L.).

PubMed

Switzenberg, Jessica A; Beaudry, Randy M; Grumet, Rebecca

2015-06-01

Ethylene is a key factor regulating sex expression in cucurbits. Commercial melons (Cucumis melo L.) are typically andromonoecious, producing male and bisexual flowers. Our prior greenhouse studies of transgenic melon plants expressing the dominant negative ethylene perception mutant gene, etr1-1, under control of the carpel- and nectary-primordia targeted CRAB'S CLAW (CRC) promoter showed increased number and earlier appearance of carpel-bearing flowers. To further investigate this phenomenon which could be potentially useful for earlier fruit production, we observed CRC::etr1-1 plants in the field for sex expression, fruit set, fruit development, and ripening. CRC::etr1-1 melon plants showed increased number of carpel-bearing open flowers on the main stem and earlier onset by 7-10 nodes. Additional phenotypes observed in the greenhouse and field were conversion of approximately 50% of bisexual buds to female, and elongated ovaries and fruits. Earlier and greater fruit set occurred on the transgenic plants. However, CRC::etr1-1 plants had greater abscission of young fruit, and smaller fruit, so that final yield (kg/plot) was equivalent to wild type. Earlier fruit set in line M5 was accompanied by earlier appearance of ripe fruit. Fruit from line M15 frequently did not exhibit external ripening processes of rind color change and abscission, but when cut open, the majority showed a ripe or overripe interior accompanied by elevated internal ethylene. The non-ripening external phenotype in M15 fruit corresponded with elevated etr1-1 transgene expression in the exocarp. These results provide insight into the role of ethylene perception in carpel-bearing flower production, fruit set, and ripening.

Body Mass Index Is Prognostic in Metastatic Colorectal Cancer: Pooled Analysis of Patients From First-Line Clinical Trials in the ARCAD Database

PubMed Central

Loupakis, Fotios; Adams, Richard A.; Seymour, Matthew T.; Heinemann, Volker; Schmoll, Hans-Joachim; Douillard, Jean-Yves; Hurwitz, Herbert; Fuchs, Charles S.; Diaz-Rubio, Eduardo; Porschen, Rainer; Tournigand, Christophe; Chibaudel, Benoist; Falcone, Alfredo; Tebbutt, Niall C.; Punt, Cornelis J.A.; Hecht, J. Randolph; Bokemeyer, Carsten; Van Cutsem, Eric; Goldberg, Richard M.; Saltz, Leonard B.; de Gramont, Aimery; Sargent, Daniel J.; Lenz, Heinz-Josef

2016-01-01

Purpose In recent retrospective analyses of early-stage colorectal cancer (CRC), low and high body mass index (BMI) scores were associated with worsened outcomes. Whether BMI is a prognostic or predictive factor in metastatic CRC (mCRC) is unclear. Patients and Methods Individual data from 21,149 patients enrolled onto 25 first-line mCRC trials during 1997 to 2012 were pooled. We assessed both prognostic and predictive effects of BMI on overall survival and progression-free survival, and we accounted for patient and tumor characteristics and therapy type (targeted v nontargeted). Results BMI was prognostic for overall survival (P < .001) and progression-free survival (P < .001), with an L-shaped pattern. That is, risk of progression and/or death was greatest for low BMI; risk decreased as BMI increased to approximately 28 kg/m2, and then it plateaued. Relative to obese patients, patients with a BMI of 18.5 kg/m2 had a 27% increased risk of having a PFS event (95% CI, 20% to 34%) and a 50% increased risk of death (95% CI, 43% to 56%). Low BMI was associated with poorer survival for men than women (interaction P < .001). BMI was not predictive of treatment effect. Conclusion Low BMI is associated with an increased risk of progression and death among the patients enrolled on the mCRC trials, with no increased risk for elevated BMI, in contrast to the adjuvant setting. Possible explanations include negative effects related to cancer cachexia in patients with low BMI, increased drug delivery or selection bias in patients with high BMI, and potential for an interaction between BMI and molecular signaling pathways. PMID:26503203

Addition of bevacizumab to fluorouracil-based first-line treatment of metastatic colorectal cancer: pooled analysis of cohorts of older patients from two randomized clinical trials.

PubMed

Kabbinavar, Fairooz F; Hurwitz, Herbert I; Yi, Jing; Sarkar, Somnath; Rosen, Oliver

2009-01-10

Colorectal cancer (CRC) occurs predominantly in older persons. To provide more statistical power to assess risk/benefit in older patients, we examined the clinical benefit of bevacizumab (BV) plus fluorouracil-based chemotherapy in first-line metastatic CRC (mCRC) treatment in patients aged > or = 65 years, using data pooled from two placebo-controlled studies. Pooled efficacy data for 439 patients > or = 65 years old randomized to BV plus chemotherapy (n = 218) or placebo plus chemotherapy (n = 221) in study 1 and study 2 were retrospectively analyzed on an intent-to-treat basis for overall survival (OS), progression-free survival (PFS), and objective response. Safety analysis was based on reports of targeted adverse events in treated patients. Median OS with BV plus chemotherapy was 19.3 v 14.3 months with placebo plus chemotherapy (hazard ratio [HR] = 0.70; 95% CI, 0.55 to 0.90; P = .006). Patients treated with BV plus chemotherapy had a median PFS of 9.2 v 6.2 months for placebo plus chemotherapy patients (HR = 0.52; 95% CI, 0.40 to 0.67; P < .0001). The objective response rate was 34.4% with BV plus chemotherapy versus 29.0% with placebo plus chemotherapy (difference not statistically significant). Rates of BV-associated adverse events in the pooled BV plus chemotherapy group were consistent with those reported in the overall populations for the two studies. Analysis of pooled patient cohorts age >/= 65 years from two similar trials in mCRC indicates that adding bevacizumab to fluorouracil-based chemotherapy improved OS and PFS, similar to the benefits in younger patients. Also, the risks of treatment do not seem to exceed those in younger patients with mCRC.

Cost-effectiveness analysis of panitumumab plus mFOLFOX6 compared with bevacizumab plus mFOLFOX6 for first-line treatment of patients with wild-type RAS metastatic colorectal cancer.

PubMed

Graham, Christopher N; Hechmati, Guy; Hjelmgren, Jonas; de Liège, Frédérique; Lanier, Julie; Knox, Hediyyih; Barber, Beth

2014-11-01

To investigate the cost-effectiveness of panitumumab plus mFOLFOX6 (oxaliplatin, 5-fluorouracil and leucovorin) compared with bevacizumab plus mFOLFOX6 in first-line treatment of patients with wild-type RAS metastatic colorectal cancer (mCRC). A semi-Markov model was constructed from a French health collective perspective, with health states related to first-line treatment (progression-free), disease progression with and without subsequent active treatment, resection of metastases, disease-free after successful resection and death. Parametric survival analyses of patient-level progression-free and overall survival data from the only head-to-head clinical trial of panitumumab and bevacizumab (PEAK) were performed to estimate transitions to disease progression and death. Additional data from PEAK informed the amount of each drug consumed, duration of therapy, subsequent therapy use, and toxicities related to mCRC treatment. Literature and French public data sources were used to estimate unit costs associated with treatment and duration of subsequent active therapies. Utility weights were calculated from patient-level data from panitumumab trials in the first-, second- and third-line settings. A life-time perspective was applied. Scenario, one-way, and probabilistic sensitivity analyses were performed. Based on a head-to-head clinical trial that demonstrates better efficacy outcomes for patients with wild-type RAS mCRC who receive panitumumab plus mFOLFOX6 versus bevacizumab plus mFOLFOX6, the incremental cost per life-year gained was estimated to be €26,918, and the incremental cost per quality-adjusted life year (QALY) gained was estimated to be €36,577. Sensitivity analyses indicate the model is robust to alternative parameters and assumptions. The incremental cost per QALY gained indicates that panitumumab plus mFOLFOX6 represents good value for money in comparison to bevacizumab plus mFOLFOX6 and, with a willingness-to-pay ranging from €40,000 to €60,000, can be considered cost-effective in first-line treatment of patients with wild-type RAS mCRC. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Early and Partial Reduction in CD4+Foxp3+ Regulatory T Cells during Colitis-Associated Colon Cancer Induces CD4+ and CD8+ T Cell Activation Inhibiting Tumorigenesis

PubMed Central

Olguín, Jonadab E.; Medina-Andrade, Itzel; Molina, Emmanuel; Vázquez, Armando; Pacheco-Fernández, Thalia; Saavedra, Rafael; Pérez-Plasencia, Carlos; Chirino, Yolanda I.; Vaca-Paniagua, Felipe; Arias-Romero, Luis E.; Gutierrez-Cirlos, Emma B.; León-Cabrera, Sonia A.; Rodriguez-Sosa, Miriam; Terrazas, Luis I.

2018-01-01

Colorectal cancer (CRC) is the second most commonly diagnosed cancer in women and the third in men in North America and Europe. CRC is associated with inflammatory responses in which intestinal pathology is caused by different cell populations including a T cell dysregulation that concludes in an imbalance between activated T (Tact) and regulatory T (Treg) cells. Treg cells are CD4+Foxp3+ cells that actively suppress pathological and physiological immune responses, contributing to the maintenance of immune homeostasis. A tumor-promoting function for Treg cells has been suggested in CRC, but the kinetics of Treg cells during CRC development are poorly known. Therefore, using a mouse model of colitis-associated colon cancer (CAC) induced by azoxymethane and dextran sodium sulfate, we observed the dynamic and differential kinetics of Treg cells in blood, spleen and mesenteric lymph nodes (MLNs) as CAC progresses, highlighting a significant reduction in Treg cells in blood and spleen during early CAC development, whereas increasing percentages of Treg cells were detected in late stages in MLNs. Interestingly, when Treg cells were decreased, Tact cells were increased and vice versa. Treg cells from late stages of CAC displayed an activated phenotype by expressing PD1, CD127 and Tim-3, suggesting an increased suppressive capacity. Suppression assays showed that T-CD4+ and T-CD8+ cells were suppressed more efficiently by MLN Treg cells from CAC animals. Finally, an antibody-mediated reduction in Treg cells during early CAC development resulted in a better prognostic value, because animals showed a reduction in tumor progression associated with an increased percentage of activated CD4+CD25+Foxp3- and CD8+CD25+ T cells in MLNs, suggesting that Treg cells suppress T cell activation at early steps during CAC development. PMID:29344269

Targeting LGR5 in Colorectal Cancer: therapeutic gold or too plastic?

PubMed

Morgan, R G; Mortensson, E; Williams, A C

2018-05-01

Leucine-rich repeat-containing G-protein coupled receptor (LGR5 or GPR49) potentiates canonical Wnt/β-catenin signalling and is a marker of normal stem cells in several tissues, including the intestine. Consistent with stem cell potential, single isolated LGR5 + cells from the gut generate self-organising crypt/villus structures in vitro termed organoids or 'mini-guts', which accurately model the parent tissue. The well characterised deregulation of Wnt/β-catenin signalling that occurs during the adenoma-carcinoma sequence in colorectal cancer (CRC) renders LGR5 an interesting therapeutic target. Furthermore, recent studies demonstrating that CRC tumours contain LGR5 + subsets and retain a degree of normal tissue architecture has heightened translational interest. Such reports fuel hope that specific subpopulations or molecules within a tumour may be therapeutically targeted to prevent relapse and induce long-term remissions. Despite these observations, many studies within this field have produced conflicting and confusing results with no clear consensus on the therapeutic value of LGR5. This review will recap the various oncogenic and tumour suppressive roles that have been described for the LGR5 molecule in CRC. It will further highlight recent studies indicating the plasticity or redundancy of LGR5 + cells in intestinal cancer progression and assess the overall merit of therapeutically targeting LGR5 in CRC.

miR-659-3p is involved in the regulation of the chemotherapy response of colorectal cancer via modulating the expression of SPHK1

PubMed Central

Li, Shuyuan; Fang, Ying; Qin, Hai; Fu, Wenzheng; Zhang, Xipeng

2016-01-01

Colorectal cancer (CRC) is one of most prevalent malignant diseases worldwide. Metastasis and chemo-resistance are the two prominent death-related factors of CRCs. Thus, it is urgent to understand the mechanism responsible for the chemo-resistant properties of CRC and develop new therapeutic methods. Here, we found that the expression of miR-659-3p was significantly reduced in cisplatin (CDDP)-resistant HT29 and LOVO colorectal cancer cells and in CDDP-resistant clinical colorectal cancer samples compared with respective CDDP-sensitive counterparts. Sphingosine kinase 1 (SPHK1) is a direct target of miR-659-3p in colorectal cancer cells, and it is negatively regulated by miR-659-3p. We found that anti-miR-659-3p could increase the IC50 of CDDP in parental HT29 and LOVO colorectal cancer cells; additionally, miR-659-3p mimics decreased the IC50 of CDDP in HT29/CDDP and LOVO/CDDP colorectal cancer cells. Furthermore, we showed that the miR-659-3p/SPHK1 pathway was involved in the regulation of chemotherapy responses of colorectal cancer cells in vivo. In all, our findings suggest a new mechanism involved in the regulation of the chemotherapy response of CRC and might provide new targets for CRC prevention and treatment. PMID:27725903

Celastrol Ameliorates Ulcerative Colitis-Related Colorectal Cancer in Mice via Suppressing Inflammatory Responses and Epithelial-Mesenchymal Transition

PubMed Central

Lin, Lianjie; Sun, Yan; Wang, Dongxu; Zheng, Shihang; Zhang, Jing; Zheng, Changqing

2016-01-01

Celastrol, also named as tripterine, is a pharmacologically active ingredient extracted from the root of traditional Chinese herb Tripterygium wilfordii Hook F with potent anti-inflammatory and anti-tumor activities. In the present study, we investigated the effects of celastrol on ulcerative colitis-related colorectal cancer (UC-CRC) as well as CRC in vivo and in vitro and explored its underlying mechanisms. UC-CRC model was induced in C57BL/6 mice by administration of azoxymethane (AOM) and dextran sodium sulfate (DSS). Colonic tumor xenograft models were developed in BALB/c-nu mice by subcutaneous injection with HCT116 and HT-29 cells. Intragastric administration of celastrol (2 mg/kg/d) for 14 weeks significantly increased the survival ratio and reduced the multiplicity of colonic neoplasms compared with AOM/DSS model mice. Mechanically, celastrol treatment significantly prevented AOM/DSS-induced up-regulation of expression levels of oncologic markers including mutated p53 and phospho-p53, β-catenin and proliferating cell nuclear antigen (PCNA). In addition, treatment with celastrol inhibited inflammatory responses, as indicated by the decrease of serum tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-6, down-regulation of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), and inactivation of nuclear factor κB (NF-κB). Moreover, celastrol obviously suppressed epithelial-mesenchymal transition (EMT) through up-regulating E-cadherin and down-regulating N-cadherin, Vimentin and Snail. Additionally, we also demonstrated that celastrol inhibited human CRC cell proliferation and attenuated colonic xenograft tumor growth via reversing EMT. Taken together, celastrol could effectively ameliorate UC-CRC by suppressing inflammatory responses and EMT, suggesting a potential drug candidate for UC-CRC therapy. PMID:26793111

Isolation and characterization of circulating micro(nano)vesicles in the plasma of colorectal cancer patients and their interactions with tumor cells.

PubMed

Stec, Małgorzata; Baj-Krzyworzeka, Monika; Baran, Jarosław; Węglarczyk, Kazimierz; Zembala, Maria; Barbasz, Jakub; Szczepanik, Antoni; Zembala, Marek

2015-11-01

Micro(nano)vesicles (MV) are regarded as important messengers in cell-to-cell communication. There is also evidence for their pivotal role in cancer progression. Circulating MV are of different body cells origin, including tumor cell‑derived MV (TMV) in cancer patients. Determination of circulating TMV is of importance because of their potential diagnostic and therapeutic applications. In the present study, an analysis of circulating MV in colorectal cancer (CRC) patients was undertaken. Plasma from healthy donors was used as the control. In order to define MV characteristics, two plasma fractions: obtained by sequential centrifugation at 15,000 x g (MV15) and 50,000 x g (MV50) were used for analysis. The two fractions possessed a large range of sizes: 70(80)-1,300(1,400) nm and the most common particles with sizes 70-90 nm, both in patients and controls. Atomic force microscopy images of MV50 revealed a heterogeneous population of particles with different shapes and sizes. MV15 contained an increased level of CD41+ and CD61+ particles, suggesting their platelet origin. No difference between patients and controls was observed. A more precise analysis of MV50 showed the increased level of particles expressing EGFR (HER-1/Erb B1), HER-2/neu and Mucin1 (MUC1), suggesting their tumor origin. The total level of MV50‑expressing EGFR, HER-2/neu and MUC1 was enhanced in CRC patients. MV50 both of patients and controls attached to a colon cancer cell line (SW480) and to isolated blood monocytes at 2 h and were engulfed at 24 h. This uptake showed the lack of specificity. Thus, apart from the direct delivery of MV to the tumor site by plasma, monocytes carrying MV may also be involved in their transportation. Taken together, the presented data indicate that MV15 contain mainly platelet‑derived particles, while MV50 from CRC patients are enriched in TMV. Interaction of MV with cancer cells may pin-point their role in communication between tumor cells, resulting in molecular cargo exchange between them.

Influence of the Hfq and Crc global regulators on the control of iron homeostasis in Pseudomonas putida.

PubMed

Sánchez-Hevia, Dione L; Yuste, Luis; Moreno, Renata; Rojo, Fernando

2018-04-30

Metabolically versatile bacteria use catabolite repression control to select their preferred carbon sources, thus optimizing carbon metabolism. In pseudomonads, this occurs through the combined action of the proteins Hfq and Crc, which form stable tripartite complexes at target mRNAs, inhibiting their translation. The activity of Hfq/Crc is antagonised by small RNAs of the CrcZ family, the amounts of which vary according to carbon availability. The present work examines the role of Pseudomonas putida Hfq protein under conditions of low-level catabolite repression, in which Crc protein would have a minor role since it is sequestered by CrcZ/CrcY. The results suggest that, under these conditions, Hfq remains operative and plays an important role in iron homeostasis. In this scenario, Crc appears to participate indirectly by helping CrcZ/CrcY to control the amount of free Hfq in the cell. Iron homeostasis in pseudomonads relies on regulatory elements such as the Fur protein, the PrrF1-F2 sRNAs, and several extracytoplasmic sigma factors. Our results show that the absence of Hfq is paralleled by a reduction in PrrF1-F2 small RNAs. Hfq thus provides a regulatory link between iron and carbon metabolism, coordinating the iron supply to meet the needs of the enzymes operational under particular nutritional regimes. This article is protected by copyright. All rights reserved. © 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.

Circulating tumor cell isolation during resection of colorectal cancer lung and liver metastases: a prospective trial with different detection techniques.

PubMed

Kaifi, Jussuf T; Kunkel, Miriam; Das, Avisnata; Harouaka, Ramdane A; Dicker, David T; Li, Guangfu; Zhu, Junjia; Clawson, Gary A; Yang, Zhaohai; Reed, Michael F; Gusani, Niraj J; Kimchi, Eric T; Staveley-O'Carroll, Kevin F; Zheng, Si-Yang; El-Deiry, Wafik S

2015-01-01

Colorectal cancer (CRC) metastasectomy improves survival, however most patient develop recurrences. Circulating tumor cells (CTCs) are an independent prognostic marker in stage IV CRC. We hypothesized that CTCs can be enriched during metastasectomy applying different isolation techniques. 25 CRC patients undergoing liver (16 (64%)) or lung (9 (36%)) metastasectomy were prospectively enrolled (clinicaltrial.gov identifier: NCT01722903). Central venous (liver) or radial artery (lung) tumor outflow blood (7.5 ml) was collected at incision, during resection, 30 min after resection, and on postoperative day (POD) 1. CTCs were quantified with 1. EpCAM-based CellSearch® system and 2. size-based isolation with a novel filter device (FMSA). CTCs were immunohistochemically identified using CellSearch®'s criteria (cytokeratin 8/18/19+, CD45- cells containing a nucleus (DAPI+)). CTCs were also enriched with a centrifugation technique (OncoQuick®). CTC numbers peaked during the resection with the FMSA in contrast to CellSearch® (mean CTC number during resection: FMSA: 22.56 (SEM 7.48) (p = 0.0281), CellSearch®: 0.87 (SEM ± 0.44) (p = 0.3018)). Comparing the 2 techniques, CTC quantity was significantly higher with the FMSA device (range 0-101) than CellSearch® (range 0-9) at each of the 4 time points examined (P < 0.05). Immunofluorescence staining of cultured CTCs revealed that CTCs have a combined epithelial (CK8/18/19) and macrophage (CD45/CD14) phenotype. Blood sampling during CRC metastasis resection is an opportunity to increase CTC capture efficiency. CTC isolation with the FMSA yields more CTCs than the CellSearch® system. Future studies should focus on characterization of single CTCs to identify targets for molecular therapy and immune escape mechanisms of cancer cells.

Hydrogen sulfide-releasing naproxen suppresses colon cancer cell growth and inhibits NF-κB signaling.

PubMed

Kodela, Ravinder; Nath, Niharika; Chattopadhyay, Mitali; Nesbitt, Diandra E; Velázquez-Martínez, Carlos A; Kashfi, Khosrow

2015-01-01

Colorectal cancer (CRC) is the second leading cause of death due to cancer and the third most common cancer in men and women in the USA. Nuclear factor kappa B (NF-κB) is known to be activated in CRC and is strongly implicated in its development and progression. Therefore, activated NF-κB constitutes a bona fide target for drug development in this type of malignancy. Many epidemiological and interventional studies have established nonsteroidal anti-inflammatory drugs (NSAIDs) as a viable chemopreventive strategy against CRC. Our previous studies have shown that several novel hydrogen sulfide-releasing NSAIDs are promising anticancer agents and are safer derivatives of NSAIDs. In this study, we examined the growth inhibitory effect of a novel H2S-releasing naproxen (HS-NAP), which has a repertoire as a cardiovascular-safe NSAID, for its effects on cell proliferation, cell cycle phase transitions, and apoptosis using HT-29 human colon cancer cells. We also investigated its effect as a chemo-preventive agent in a xenograft mouse model. HS-NAP suppressed the growth of HT-29 cells by induction of G0/G1 arrest and apoptosis and downregulated NF-κB. Tumor xenografts in mice were significantly reduced in volume. The decrease in tumor mass was associated with a reduction of cell proliferation, induction of apoptosis, and decreases in NF-κB levels in vivo. Therefore, HS-NAP demonstrates strong anticancer potential in CRC.

Systems-wide RNAi analysis of CASP8AP2/FLASH shows transcriptional deregulation of the replication-dependent histone genes and extensive effects on the transcriptome of colorectal cancer cells

PubMed Central

2012-01-01

Background Colorectal carcinomas (CRC) carry massive genetic and transcriptional alterations that influence multiple cellular pathways. The study of proteins whose loss-of-function (LOF) alters the growth of CRC cells can be used to further understand the cellular processes cancer cells depend upon for survival. Results A small-scale RNAi screen of ~400 genes conducted in SW480 CRC cells identified several candidate genes as required for the viability of CRC cells, most prominently CASP8AP2/FLASH. To understand the function of this gene in maintaining the viability of CRC cells in an unbiased manner, we generated gene specific expression profiles following RNAi. Silencing of CASP8AP2/FLASH resulted in altered expression of over 2500 genes enriched for genes associated with cellular growth and proliferation. Loss of CASP8AP2/FLASH function was significantly associated with altered transcription of the genes encoding the replication-dependent histone proteins as a result of the expression of the non-canonical polyA variants of these transcripts. Silencing of CASP8AP2/FLASH also mediated enrichment of changes in the expression of targets of the NFκB and MYC transcription factors. These findings were confirmed by whole transcriptome analysis of CASP8AP2/FLASH silenced cells at multiple time points. Finally, we identified and validated that CASP8AP2/FLASH LOF increases the expression of neurofilament heavy polypeptide (NEFH), a protein recently linked to regulation of the AKT1/ß-catenin pathway. Conclusions We have used unbiased RNAi based approaches to identify and characterize the function of CASP8AP2/FLASH, a protein not previously reported as required for cell survival. This study further defines the role CASP8AP2/FLASH plays in the regulating expression of the replication-dependent histones and shows that its LOF results in broad and reproducible effects on the transcriptome of colorectal cancer cells including the induction of expression of the recently described tumor suppressor gene NEFH. PMID:22216762

Systems-wide RNAi analysis of CASP8AP2/FLASH shows transcriptional deregulation of the replication-dependent histone genes and extensive effects on the transcriptome of colorectal cancer cells.

PubMed

Hummon, Amanda B; Pitt, Jason J; Camps, Jordi; Emons, Georg; Skube, Susan B; Huppi, Konrad; Jones, Tamara L; Beissbarth, Tim; Kramer, Frank; Grade, Marian; Difilippantonio, Michael J; Ried, Thomas; Caplen, Natasha J

2012-01-04

Colorectal carcinomas (CRC) carry massive genetic and transcriptional alterations that influence multiple cellular pathways. The study of proteins whose loss-of-function (LOF) alters the growth of CRC cells can be used to further understand the cellular processes cancer cells depend upon for survival. A small-scale RNAi screen of ~400 genes conducted in SW480 CRC cells identified several candidate genes as required for the viability of CRC cells, most prominently CASP8AP2/FLASH. To understand the function of this gene in maintaining the viability of CRC cells in an unbiased manner, we generated gene specific expression profiles following RNAi. Silencing of CASP8AP2/FLASH resulted in altered expression of over 2500 genes enriched for genes associated with cellular growth and proliferation. Loss of CASP8AP2/FLASH function was significantly associated with altered transcription of the genes encoding the replication-dependent histone proteins as a result of the expression of the non-canonical polyA variants of these transcripts. Silencing of CASP8AP2/FLASH also mediated enrichment of changes in the expression of targets of the NFκB and MYC transcription factors. These findings were confirmed by whole transcriptome analysis of CASP8AP2/FLASH silenced cells at multiple time points. Finally, we identified and validated that CASP8AP2/FLASH LOF increases the expression of neurofilament heavy polypeptide (NEFH), a protein recently linked to regulation of the AKT1/ß-catenin pathway. We have used unbiased RNAi based approaches to identify and characterize the function of CASP8AP2/FLASH, a protein not previously reported as required for cell survival. This study further defines the role CASP8AP2/FLASH plays in the regulating expression of the replication-dependent histones and shows that its LOF results in broad and reproducible effects on the transcriptome of colorectal cancer cells including the induction of expression of the recently described tumor suppressor gene NEFH.

Interplay between apoptosis and autophagy in colorectal cancer.

PubMed

Qian, Hao-Ran; Shi, Zhao-Qi; Zhu, He-Pan; Gu, Li-Hu; Wang, Xian-Fa; Yang, Yi

2017-09-22

Autophagy and apoptosis are two pivotal mechanisms in mediating cell survival and death. Cross-talk of autophagy and apoptosis has been documented in the tumorigenesis and progression of cancer, while the interplay between the two pathways in colorectal cancer (CRC) has not yet been comprehensively summarized. In this study, we outlined the basis of apoptosis and autophagy machinery firstly, and then reviewed the recent evidence in cellular settings or animal studies regarding the interplay between them in CRC. In addition, several key factors that modulate the cross-talk between autophagy and apoptosis as well as its significance in clinical practice were discussed. Understanding of the interplay between the cell death mechanisms may benefit the translation of CRC treatment from basic research to clinical use.

Interplay between apoptosis and autophagy in colorectal cancer

PubMed Central

Qian, Hao-Ran; Shi, Zhao-Qi; Zhu, He-Pan; Gu, Li-Hu; Wang, Xian-Fa; Yang, Yi

2017-01-01

Autophagy and apoptosis are two pivotal mechanisms in mediating cell survival and death. Cross-talk of autophagy and apoptosis has been documented in the tumorigenesis and progression of cancer, while the interplay between the two pathways in colorectal cancer (CRC) has not yet been comprehensively summarized. In this study, we outlined the basis of apoptosis and autophagy machinery firstly, and then reviewed the recent evidence in cellular settings or animal studies regarding the interplay between them in CRC. In addition, several key factors that modulate the cross-talk between autophagy and apoptosis as well as its significance in clinical practice were discussed. Understanding of the interplay between the cell death mechanisms may benefit the translation of CRC treatment from basic research to clinical use. PMID:28977986

Patients with colorectal cancer associated with Lynch syndrome and MLH1 promoter hypermethylation have similar prognoses.

PubMed

Haraldsdottir, Sigurdis; Hampel, Heather; Wu, Christina; Weng, Daniel Y; Shields, Peter G; Frankel, Wendy L; Pan, Xueliang; de la Chapelle, Albert; Goldberg, Richard M; Bekaii-Saab, Tanios

2016-09-01

Mismatch repair-deficient (dMMR) colorectal cancer (CRC) is caused by Lynch syndrome (LS) in 3% and sporadic inactivation of MLH1 by hypermethylation (MLH1-hm) in 12% of cases. It is not clear whether outcomes between LS-associated and MLH1-hm CRC differ. The objective of this study was to explore differences in clinical factors and outcomes in these two groups. Patients with dMMR CRC identified by immunohistochemistry staining and treated at a single institution from 1998 to 2012 were included. MLH1-hm was established with BRAF mutational analysis or hypermethylation testing. Patients' charts were accessed for information on pathology, germ-line MMR mutation testing, and clinical course. A total of 143 patients had CRC associated with LS (37 patients, 26%) or MLH1-hm (106 patients, 74%). Patients with LS were younger, more often male, presented more often with stage III disease, and had more metachronous disease than patients with MLH1-hm tumors. There was no difference in cancer-specific survival (CSS) between the groups; overall survival was longer in patients with LS, but this difference was minimal after adjusting for age and stage at diagnosis. CSS did not differ in LS-associated CRC compared with MLH1-hm CRC, suggesting that they carry a similar prognosis.Genet Med 18 9, 863-868.

The impact of computer self-efficacy, computer anxiety, and perceived usability and acceptability on the efficacy of a decision support tool for colorectal cancer screening

PubMed Central

Lindblom, Katrina; Gregory, Tess; Flight, Ingrid H K; Zajac, Ian

2011-01-01

Objective This study investigated the efficacy of an internet-based personalized decision support (PDS) tool designed to aid in the decision to screen for colorectal cancer (CRC) using a fecal occult blood test. We tested whether the efficacy of the tool in influencing attitudes to screening was mediated by perceived usability and acceptability, and considered the role of computer self-efficacy and computer anxiety in these relationships. Methods Eighty-one participants aged 50–76 years worked through the on-line PDS tool and completed questionnaires on computer self-efficacy, computer anxiety, attitudes to and beliefs about CRC screening before and after exposure to the PDS, and perceived usability and acceptability of the tool. Results Repeated measures ANOVA found that PDS exposure led to a significant increase in knowledge about CRC and screening, and more positive attitudes to CRC screening as measured by factors from the Preventive Health Model. Perceived usability and acceptability of the PDS mediated changes in attitudes toward CRC screening (but not CRC knowledge), and computer self-efficacy and computer anxiety were significant predictors of individuals' perceptions of the tool. Conclusion Interventions designed to decrease computer anxiety, such as computer courses and internet training, may improve the acceptability of new health information technologies including internet-based decision support tools, increasing their impact on behavior change. PMID:21857024

PTEN Overexpression Cooperates With Lithium to Reduce the Malignancy and to Increase Cell Death by Apoptosis via PI3K/Akt Suppression in Colorectal Cancer Cells.

PubMed

de Araujo, Wallace Martins; Robbs, Bruno Kaufmann; Bastos, Lilian G; de Souza, Waldemir F; Vidal, Flávia C B; Viola, João P B; Morgado-Diaz, Jose A

2016-02-01

Lithium is a well-established non-competitive inhibitor of glycogen synthase kinase-3β (GSK-3β), a kinase that is involved in several cellular processes related to cancer progression. GSK-3β is regulated upstream by PI3K/Akt, which is negatively modulated by PTEN. The role that lithium plays in cancer is controversial because lithium can activate or inhibit survival signaling pathways depending on the cell type. In this study, we analyzed the mechanisms by which lithium can modulate events related to colorectal cancer (CRC) progression and evaluated the role that survival signaling pathways such as PI3K/Akt and PTEN play in this context. We show that the administration of lithium decreased the proliferative potential of CRC cells in a GSK-3β-independent manner but induced the accumulation of cells in G2/M phase. Furthermore, high doses of lithium increased apoptosis, which was accompanied by decreased proteins levels of Akt and PTEN. Then, cells that were induced to overexpress PTEN were treated with lithium; we observed that low doses of lithium strongly increased apoptosis. Additionally, PTEN overexpression reduced proliferation, but this effect was minor compared with that in cells treated with lithium alone. Furthermore, we demonstrated that PTEN overexpression and lithium treatment separately reduced cell migration, colony formation, and invasion, and these effects were enhanced when lithium treatment and PTEN overexpression were combined. In conclusion, our findings indicate that PTEN overexpression and lithium treatment cooperate to reduce the malignancy of CRC cells and highlight lithium and PTEN as potential candidates for studies to identify new therapeutic approaches for CRC treatment. © 2015 Wiley Periodicals, Inc.

Resveratrol Regulates Colorectal Cancer Cell Invasion by Modulation of Focal Adhesion Molecules

PubMed Central

Buhrmann, Constanze; Shayan, Parviz; Goel, Ajay; Shakibaei, Mehdi

2017-01-01

Resveratrol, a safe and multi-targeted agent, has been associated with suppression of survival, proliferation and metastasis of cancer, however, the underlying mechanisms for its anti-cancer activity, particularly on cellular signaling during cancer cell migration still remain poorly understood. We investigated the invasion response of two human colorectal cancer (CRC) cells (HCT116 and SW480) to resveratrol and studied the effect of specific pharmacological inhibitors, cytochalasin D (CytD) and focal adhesion kinase-inhibitor (FAK-I) on FAK, cell viability and migration in CRC. We found that resveratrol altered cell phenotype of both CRC cells, reduced cell viability and the results were comparable to FAK-I and CytD. These effects of resveratrol were associated with marked Sirt1 up-regulation, FAK down-regulation, inhibition of focal adhesion and potentiation of effects by combinatorial treatment of resveratrol and inhibitors. Interestingly, inhibition of FAK with FAK-I or treatment with CytD suppressed resveratrol-induced Sirt1 up-regulation and markedly down-regulated FAK expression. Resveratrol or combination treatment with inhibitors significantly activated caspase-3 and potentiated apoptosis. Moreover, resveratrol suppressed invasion and colony forming capacity, cell proliferation, β1-Integrin expression and activation of FAK of cells in alginate tumor microenvironment, similar to FAK-I or CytD. Finally, we demonstrated that resveratrol, FAK-I or CytD inhibited activation of NF-κB, suppressed NF-κB-dependent gene end-products involved in invasion, metastasis, and apoptosis; and these effects of resveratrol were potentiated by combination treatment with FAK-I or CytD. Our data illustrated that the anti-invasion effect of resveratrol by inhibition of FAK activity has a potential beneficial role in disease prevention and therapeutic management of CRC. PMID:28953264

Resveratrol Regulates Colorectal Cancer Cell Invasion by Modulation of Focal Adhesion Molecules.

PubMed

Buhrmann, Constanze; Shayan, Parviz; Goel, Ajay; Shakibaei, Mehdi

2017-09-27

Resveratrol, a safe and multi-targeted agent, has been associated with suppression of survival, proliferation and metastasis of cancer, however, the underlying mechanisms for its anti-cancer activity, particularly on cellular signaling during cancer cell migration still remain poorly understood. We investigated the invasion response of two human colorectal cancer (CRC) cells (HCT116 and SW480) to resveratrol and studied the effect of specific pharmacological inhibitors, cytochalasin D (CytD) and focal adhesion kinase-inhibitor (FAK-I) on FAK, cell viability and migration in CRC. We found that resveratrol altered cell phenotype of both CRC cells, reduced cell viability and the results were comparable to FAK-I and CytD. These effects of resveratrol were associated with marked Sirt1 up-regulation, FAK down-regulation, inhibition of focal adhesion and potentiation of effects by combinatorial treatment of resveratrol and inhibitors. Interestingly, inhibition of FAK with FAK-I or treatment with CytD suppressed resveratrol-induced Sirt1 up-regulation and markedly down-regulated FAK expression. Resveratrol or combination treatment with inhibitors significantly activated caspase-3 and potentiated apoptosis. Moreover, resveratrol suppressed invasion and colony forming capacity, cell proliferation, β1-Integrin expression and activation of FAK of cells in alginate tumor microenvironment, similar to FAK-I or CytD. Finally, we demonstrated that resveratrol, FAK-I or CytD inhibited activation of NF-κB, suppressed NF-κB-dependent gene end-products involved in invasion, metastasis, and apoptosis; and these effects of resveratrol were potentiated by combination treatment with FAK-I or CytD. Our data illustrated that the anti-invasion effect of resveratrol by inhibition of FAK activity has a potential beneficial role in disease prevention and therapeutic management of CRC.

Cancer testis antigen OY-TES-1 expression and serum immunogenicity in colorectal cancer: its relationship to clinicopathological parameters

PubMed Central

Luo, Bin; Yun, Xiang; Fan, Rong; Lin, Yong-Da; He, Shu-Jia; Zhang, Qing-Mei; Mo, Fa-Rong; Chen, Fang; Xiao, Shao-Wen; Xie, Xiao-Xun

2013-01-01

Cancer testis (CT) antigens are attractive targets for cancer immunotherapy because their expression is restricted in normal germ line tissues but frequently detected in variety of tumors. OY-TES-1 is identified as a member of CT antigens. Current knowledge about OY-TES-1 expression in colorectal cancer (CRC) is solely based on mRNA analysis. None of previous researches has studied OY-TES-1 at protein level. In this study, OY-TES-1 polyclonal antibody was generated. The expression of OY-TES-1 mRNA and protein was detected by RT-PCR and immunohistochemistry in 60 CRC and paired adjacent non-tumor tissues, 24 colorectal adenoma and 3 normal colon tissues, respectively. Sera from 73 CRC patients were also tested for OY-TES-1 antibody by ELISA. Our results showed that the frequency of OY-TES-1 mRNA expression was statistically higher in CRC (73.3%, 44/60) than that in adjacent non-tumor tissue (55.0%, 33/60) and colorectal adenoma (45.8%, 11/24). For the first time, OY-TES-1 protein expression was found in (43.3%, 26/60) of CRC tissues, but absent in any of adjacent non-tumor and colorectal adenoma tissues. No OY-TES-1 expression was found in normal colon by either RT-PCR or immunohistochemistry. Furthermore, OY-TES-1 protein expression was correlated with tumor invasion stage (P=0.004) and histological grade (P=0.040). Anti-OY-TES-1 antibody was detected in (9.6%, 7/73) of CRC patients’ sera but not in 76 healthy donors. This finding demonstrates that OY-TES-1 is frequently expressed in CRC and is able to induce humoral immune response spontaneously in CRC patients, suggesting that it might be a promising immunotherapy target for CRC. PMID:24294369