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Sample records for p27kip1 reveals cancer

  1. [Prognostic value of the inmunohïstochemical expression of protein p27KIP1 in laryngeal cancer].

    PubMed

    García Lozano, M C; Orradre Romero, J L; Caro García, M; Martínez Alvarez, A; Lasso Luis, O; Piris Pinilla, M A

    2006-01-01

    In this paper we carried out an immunohistochemical study of protein p27KIP1 expression in a series of 195 patients with laryngeal carcinoma that were diagnosticated, treated and followed at the Department of Otolaryngology at "Virgen de la Salud" Hospital (Toledo, Spain). In the cases with lymph node metastasis we also studied p27KIP1 expression at this level. Furthermore we have analysed the value of protein p27KIP1 expression as a prognostic factor (tumor recurrence, deads due to cancer and survival) and we evaluate the relationship between p27KIP1 expression and other clinic and pathologic parameters. PMID:16566195

  2. CacyBP/SIP nuclear translocation regulates p27Kip1 stability in gastric cancer cells

    PubMed Central

    Niu, Ying-Lin; Li, Ya-Jun; Wang, Jing-Bo; Lu, Yuan-Yuan; Liu, Zhen-Xiong; Feng, Shan-Shan; Hu, Jian-Guo; Zhai, Hui-Hong

    2016-01-01

    AIM: To investigate the mechanism of calcyclin binding protein/Siah-1 interacting protein (CacyBP/SIP) nuclear translocation in promoting the proliferation of gastric cancer (GC) cells. METHODS: The effect of CacyBP/SIP nuclear translocation on cell cycle was investigated by cell cycle analysis. Western blot analysis was used to assess the change in expression of cell cycle regulatory proteins and proteasome-mediated degradation of p27Kip1. Co-immunoprecipitation (co-IP) analysis was performed to examine the binding of CacyBP/SIP with Skp1. A CacyBP/SIP truncation mutant which lacked the Skp1 binding site was constructed and fused to a fluorescent protein. Subsequently, the effect on Skp1 binding with the fusion protein was examined by co-IP, while localization of fluorescent fusion protein observed by confocal laser microscopy, and change in p27Kip1 protein expression assessed by Western blot analysis. RESULTS: CacyBP/SIP nuclear translocation induced by gastrin promoted progression of GC cells from G1 phase. However, while CacyBP/SIP nuclear translocation was inhibited using siRNA to suppress CacyBP/SIP expression, cell cycle was clearly inhibited. CacyBP/SIP nuclear translocation significantly decreased the level of cell cycle inhibitor p27Kip1, increased Cyclin E protein expression whereas the levels of Skp1, Skp2, and CDK2 were not affected. Upon inhibition of CacyBP/SIP nuclear translocation, there were no changes in protein levels of p27Kip1 and Cyclin E, while p27Kip1 decrease could be prevented by the proteasome inhibitor MG132. Moreover, CacyBP/SIP was found to bind to Skp1 by immunoprecipitation, an event that was abolished by mutant CacyBP/SIP, which also failed to stimulate p27Kip1 degradation, even though the mutant could still translocate into the nucleus. CONCLUSION: CacyBP/SIP nuclear translocation contributes to the proliferation of GC cells, and CacyBP/SIP exerts this effect, at least in part, by stimulating ubiquitin-mediated degradation of p27

  3. A prostatic intraepithelial neoplasia-dependent p27kip1 checkpoint induces senescence, inhibits cell proliferation and cancer progression

    PubMed Central

    Majumder, Pradip K.; Grisanzio, Chiara; O’Connell, Fionnuala; Barry, Marc; Brito, Joseph M.; Xu, Qing; Guney, Isil; Berger, Raanan; Herman, Paula; Bikoff, Rachel; Fedele, Giuseppe; Baek, Won-Ki; Wang, Shunyou; Ellwood-Yen, Katharine; Wu, Hong; Sawyers, Charles L.; Signoretti, Sabina; Hahn, William C.; Loda, Massimo; Sellers, William R.

    2008-01-01

    SUMMARY Transgenic expression of activated AKT1 in the murine prostate induces Prostatic Intraepithelial Neoplasia (PIN) that does not progress to invasive prostate cancer (CaP). In luminal epithelial cells of Akt-driven PIN we show the concomitant induction of p27kip1 and senescence. Genetic ablation of p27Kip1 led to down regulation of senescence markers and progression to cancer. In humans, p27Kip1 and senescence markers were elevated in PIN not associated with CaP, but were decreased and absent, respectively in cancer-associated PIN and in CaP. Importantly, p27Kip1 up-regulation in mouse and human in situ lesions did not depend upon mTOR or Akt activation but was instead specifically associated with alterations in cellular polarity, architecture and adhesion molecules. These data suggest that a p27Kip1-driven checkpoint limits progression of PIN to CaP. PMID:18691549

  4. Phosphorylation and subcellular localization of p27Kip1 regulated by hydrogen peroxide modulation in cancer cells.

    PubMed

    Ibañez, Irene L; Bracalente, Candelaria; Notcovich, Cintia; Tropper, Ivanna; Molinari, Beatriz L; Policastro, Lucía L; Durán, Hebe

    2012-01-01

    The Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a key protein in the decision between proliferation and cell cycle exit. Quiescent cells show nuclear p27Kip1, but this protein is exported to the cytoplasm in response to proliferating signals. We recently reported that catalase treatment increases the levels of p27Kip1 in vitro and in vivo in a murine model. In order to characterize and broaden these findings, we evaluated the regulation of p27Kip1 by hydrogen peroxide (H(2)O(2)) in human melanoma cells and melanocytes. We observed a high percentage of p27Kip1 positive nuclei in melanoma cells overexpressing or treated with exogenous catalase, while non-treated controls showed a cytoplasmic localization of p27Kip1. Then we studied the levels of p27Kip1 phosphorylated (p27p) at serine 10 (S10) and at threonine 198 (T198) because phosphorylation at these sites enables nuclear exportation of this protein, leading to accumulation and stabilization of p27pT198 in the cytoplasm. We demonstrated by western blot a decrease in p27pS10 and p27pT198 levels in response to H(2)O(2) removal in melanoma cells, associated with nuclear p27Kip1. Melanocytes also exhibited nuclear p27Kip1 and lower levels of p27pS10 and p27pT198 than melanoma cells, which showed cytoplasmic p27Kip1. We also showed that the addition of H(2)O(2) (0.1 µM) to melanoma cells arrested in G1 by serum starvation induces proliferation and increases the levels of p27pS10 and p27pT198 leading to cytoplasmic localization of p27Kip1. Nuclear localization and post-translational modifications of p27Kip1 were also demonstrated by catalase treatment of colorectal carcinoma and neuroblastoma cells, extending our findings to these other human cancer types. In conclusion, we showed in the present work that H(2)O(2) scavenging prevents nuclear exportation of p27Kip1, allowing cell cycle arrest, suggesting that cancer cells take advantage of their intrinsic pro-oxidant state to favor cytoplasmic localization of p27

  5. Phosphorylation and Subcellular Localization of p27Kip1 Regulated by Hydrogen Peroxide Modulation in Cancer Cells

    PubMed Central

    Ibañez, Irene L.; Bracalente, Candelaria; Notcovich, Cintia; Tropper, Ivanna; Molinari, Beatriz L.; Policastro, Lucía L.; Durán, Hebe

    2012-01-01

    The Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a key protein in the decision between proliferation and cell cycle exit. Quiescent cells show nuclear p27Kip1, but this protein is exported to the cytoplasm in response to proliferating signals. We recently reported that catalase treatment increases the levels of p27Kip1 in vitro and in vivo in a murine model. In order to characterize and broaden these findings, we evaluated the regulation of p27Kip1 by hydrogen peroxide (H2O2) in human melanoma cells and melanocytes. We observed a high percentage of p27Kip1 positive nuclei in melanoma cells overexpressing or treated with exogenous catalase, while non-treated controls showed a cytoplasmic localization of p27Kip1. Then we studied the levels of p27Kip1 phosphorylated (p27p) at serine 10 (S10) and at threonine 198 (T198) because phosphorylation at these sites enables nuclear exportation of this protein, leading to accumulation and stabilization of p27pT198 in the cytoplasm. We demonstrated by western blot a decrease in p27pS10 and p27pT198 levels in response to H2O2 removal in melanoma cells, associated with nuclear p27Kip1. Melanocytes also exhibited nuclear p27Kip1 and lower levels of p27pS10 and p27pT198 than melanoma cells, which showed cytoplasmic p27Kip1. We also showed that the addition of H2O2 (0.1 µM) to melanoma cells arrested in G1 by serum starvation induces proliferation and increases the levels of p27pS10 and p27pT198 leading to cytoplasmic localization of p27Kip1. Nuclear localization and post-translational modifications of p27Kip1 were also demonstrated by catalase treatment of colorectal carcinoma and neuroblastoma cells, extending our findings to these other human cancer types. In conclusion, we showed in the present work that H2O2 scavenging prevents nuclear exportation of p27Kip1, allowing cell cycle arrest, suggesting that cancer cells take advantage of their intrinsic pro-oxidant state to favor cytoplasmic localization of p27Kip1. PMID

  6. Rheb promotes cancer cell survival through p27Kip1-dependent activation of autophagy.

    PubMed

    Campos, Tania; Ziehe, Javiera; Palma, Mario; Escobar, David; Tapia, Julio C; Pincheira, Roxana; Castro, Ariel F

    2016-02-01

    We previously found that the small GTPase Rheb regulates the cell-cycle inhibitor p27KIP1 (p27) in colon cancer cells by a mTORC1-independent mechanism. However, the biological function of the Rheb/p27 axis in cancer cells remains unknown. Here, we show that siRNA-mediated depletion of Rheb decreases survival of human colon cancer cells under serum deprivation. As autophagy can support cell survival, we analyzed the effect of Rheb on this process by detecting the modification of the autophagy marker protein LC3 by western blot and imunofluorescence. We found that Rheb promotes autophagy in several human cancer cell lines under serum deprivation. Accordingly, blocking autophagy inhibited the pro-survival effect of Rheb in colon cancer cells. We then analyzed whether p27 was involved in the biological effect of Rheb. Depletion of p27 inhibited colon cancer cell survival, and Rheb induction of autophagy. These results suggest that p27 has an essential role in the effect of Rheb in response to serum deprivation. In addition, we demonstrated that the role of p27 in autophagy stands on the N-terminal portion of the protein, where the CDK-inhibitory domain is located. Our results indicate that a Rheb/p27 axis accounts for the activation of autophagy that supports cancer cell survival. Our work therefore highlights a biological function of Rheb and prompts the need for future studies to address whether the mTORC1-independent Rheb/p27 axis could contribute to tumorigenesis and/or resistance to mTOR inhibitors. PMID:25594310

  7. eIF3a improve cisplatin sensitivity in ovarian cancer by regulating XPC and p27Kip1 translation

    PubMed Central

    Zhang, Yu; Yu, Jing-Jing; Tian, Yan; Li, Zheng-Zheng; Zhang, Cai-Yi; Zhang, Shu-Fen; Cao, Lan-Qin; Zhang, Yi; Qian, Chen-Yue; Zhang, Wei; Zhou, Hong-Hao; Yin, Ji-Ye; Liu, Zhao-Qian

    2015-01-01

    The eukaryotic translation initiation factor 3a (eIF3a) is one of the core subunits of the translation initiation complex eIF3, responsible for ribosomal subunit joining and mRNA recruitment to the ribosome. Our previous study identified that it was correlated with platinum response in lung cancer. The current study aims to test the hypothesis that eIF3a may affect the drug response and prognosis of ovarian cancer patients receiving platinum-based chemotherapy by regulating xeroderma pigmentosum complementation group C (XPC) and p27Kip1. Immunohistochemistry and western blot was used to determine the expression of eIF3a in 126 human ovarian cancer tissues followed by association analysis of eIF3a expression with patient's response and survival. Ectopic over-expression and RNA interference knockdown of eIF3a were carried out in A2780/cisplatin (DDP) and its parental A2780 cells, respectively, to determine the effect of altered eIF3a expression on cellular response to cisplatin by employing MTT assay. Western Blot analyses were also carried out to determine the regulation of eIF3a on XPC and p27Kip1. eIF3a expression was associated with response of ovarian cancer patients to DDP-based chemotherapy and their survival. Overexpression and knockdown of eIF3a increased and decreased the cellular response to cisplatin in A2780/DDP and A2780 cells, respectively. In addition, XPC and p27Kip1 were down regulated by eIF3a. eIF3a improves ovarian cancer patients' response to DDP-based chemotherapy via down regulating XPC and p27Kip1. PMID:26213845

  8. Regulation of the p27Kip1 tumor suppressor by miR-221 and miR-222 promotes cancer cell proliferation

    PubMed Central

    le Sage, Carlos; Nagel, Remco; Egan, David A; Schrier, Mariette; Mesman, Elly; Mangiola, Annunziato; Anile, Corrado; Maira, Giulio; Mercatelli, Neri; Ciafrè, Silvia Anna; Farace, Maria Giulia; Agami, Reuven

    2007-01-01

    MicroRNAs (miRNAs) are potent post-transcriptional regulators of protein coding genes. Patterns of misexpression of miRNAs in cancer suggest key functions of miRNAs in tumorigenesis. However, current bioinformatics tools do not entirely support the identification and characterization of the mode of action of such miRNAs. Here, we used a novel functional genetic approach and identified miR-221 and miR-222 (miR-221&222) as potent regulators of p27Kip1, a cell cycle inhibitor and tumor suppressor. Using miRNA inhibitors, we demonstrate that certain cancer cell lines require high activity of miR-221&222 to maintain low p27Kip1 levels and continuous proliferation. Interestingly, high levels of miR-221&222 appear in glioblastomas and correlate with low levels of p27Kip1 protein. Thus, deregulated expression of miR-221&222 promotes cancerous growth by inhibiting the expression of p27Kip1. PMID:17627278

  9. Upregulation of p27Kip1 by demethylation sensitizes cisplatin-resistant human ovarian cancer SKOV3 cells.

    PubMed

    Zhao, Yan; Li, Qiaoyan; Wu, Xiaoying; Chen, Puxiang

    2016-08-01

    Ovarian cancer has a poor prognosis due to its chemoresistance, and p27Kip1 (p27) has been implicated in tumor prognosis and drug-resistance. However, the regulatory mechanisms of p27 in drug‑resistance in ovarian cancer remain unknown. The current study successfully established chemoresistant cell lines using paclitaxel (TAX), cisplatin (DDP) and carboplatin (CBP) in SKOV3 ovarian cancer cells. The results indicated that the expression levels of p27 were dramatically downregulated in chemoresistant cells. However, 5-aza-2'-deoxycytidine (5-aza) treatment restored p27 expression in DDP-resistant cells, and increased their sensitivity to DDP. In addition, it was observed that the methylation of DDP‑resistant cells, which was downregulated by 5‑aza treatment, was significantly higher compared with SKOV3 cells. Additionally, the overexpression of p27 arrested the cell cycle in S phase and promoted an apoptotic response to DDP. In conclusion, p27 was involved in chemoresistance of SKOV3 cells. Upregulated p27 expression induced by demethylation may enhance sensitivity to DDP through the regulation of the cell cycle. PMID:27314502

  10. A novel miRNA-mediated STOP sign in lung cancer: miR-340 inhibits the proliferation of lung cancer cells through p27KIP1

    PubMed Central

    Fernandez, Serena; Risolino, Maurizio; Verde, Pasquale

    2015-01-01

    Oncosuppressor miRNAs inhibit cancer cell proliferation by targeting key components of the cell cycle machinery. In our recent report we showed that miR-340 is a novel tumor suppressor in non-small cell lung cancer. miR-340 inhibits neoplastic cell proliferation and induces p27KIP1 by targeting multiple translational and post-translational regulators of this cyclin-dependent kinase inhibitor. PMID:27308439

  11. miR-222 confers the resistance of breast cancer cells to Adriamycin through suppression of p27(kip1) expression.

    PubMed

    Wang, Dan-Dan; Li, Jian; Sha, Huan-Huan; Chen, Xiu; Yang, Su-Jin; Shen, Hong-Yu; Zhong, Shan-Liang; Zhao, Jian-Hua; Tang, Jin-Hai

    2016-09-15

    Adriamycin (Adr) is a potent chemotherapeutic agent for chemotherapy of breast cancer patients. Despite impressive initial clinical responses, some developed drug resistance to Adr-based therapy and the mechanisms underlying breast cancer cells resistance to Adr are not well known. In our previous study, in vitro, we verified that miR-222 was upregulated in Adr-resistant breast cancer cells (MCF-7/Adr) compared with the sensitive parental cells (MCF-7/S). Here, miR-222 inhibitors or mimics were transfected into MCF-7 cell lines. RT-qPCR and western blot were used to detect the expression of p27(kip1). Immunofluorescence showed that miR-222 altered the subcellular location of p27(kip1) in nucleus. MTT was employed to verify the sensitivity of breast cancer cell lines to Adr. Flow cytometry showed the apoptosis and cell cycles of the cells after adding Adr. The results showed that downregulation of miR-222 in MCF-7/Adr increased sensitivity to Adr and Adr-induced apoptosis, and arrested the cells in G1 phase, accompanied by more expressions of p27(kip1), especially in nucleus. Furthermore, overexpressed miR-222 in MCF-7/S had the inverse results. Taken together, the results found that miR-222 induced Adr-resistance at least in part via suppressing p27(kip1) expression and altering its subcellular localization, and miR-222 inhibitors could reverse Adr-resistance of breast cancer cells. These results disclosed that the future holds much promise for the targeted therapeutic in the treatment of Adr-resistant breast cancer. PMID:27282281

  12. Cooperation of p27Kip1 and p18INK4c in Progestin-Mediated Cell Cycle Arrest in T-47D Breast Cancer Cells

    PubMed Central

    Swarbrick, Alexander; Lee, Christine S. L.; Sutherland, Robert L.; Musgrove, Elizabeth A.

    2000-01-01

    The steroid hormone progesterone regulates proliferation and differentiation in the mammary gland and uterus by cell cycle phase-specific actions. The long-term effect of progestins on T-47D breast cancer cells is inhibition of cellular proliferation. This is accompanied by decreased G1 cyclin-dependent kinase (CDK) activities, redistribution of the CDK inhibitor p27Kip1 among these CDK complexes, and alterations in the elution profile of cyclin E-Cdk2 upon gel filtration chromatography, such that high-molecular-weight complexes predominate. This study aimed to determine the relative contribution of CDK inhibitors to these events. Following progestin treatment, the majority of cyclin E- and D-CDK complexes were bound to p27Kip1 and few were bound to p21Cip1. In vitro, recombinant His6-p27 could quantitatively reproduce the effects on cyclin E-Cdk2 kinase activity and the shift in molecular weight observed following progestin treatment. In contrast, cyclin D-Cdk4 was not inhibited by His6-p27 in vitro or p27Kip1 in vivo. However, an increase in the expression of the Cdk4/6 inhibitor p18INK4c and its extensive association with Cdk4 and Cdk6 were apparent following progestin treatment. Recombinant p18INK4c led to the reassortment of cyclin-CDK-CDK inhibitor complexes in vitro, with consequent decrease in cyclin E-Cdk2 activity. These results suggest a concerted model of progestin action whereby p27Kip1 and p18INK4c cooperate to inhibit cyclin E-Cdk2 and Cdk4. Since similar models have been developed for growth inhibition by transforming growth factor β and during adipogenesis, interaction between the Cip/Kip and INK4 families of inhibitors may be a common theme in physiological growth arrest and differentiation. PMID:10713180

  13. Knockdown of CRM1 inhibits the nuclear export of p27(Kip1) phosphorylated at serine 10 and plays a role in the pathogenesis of epithelial ovarian cancer.

    PubMed

    Wang, You; Wang, Yingying; Xiang, Jingying; Ji, Fang; Deng, Yan; Tang, Chunhui; Yang, Shuyun; Xi, Qinghua; Liu, Rong; Di, Wen

    2014-02-01

    In a previous study, the nuclear export protein chromosomal region maintenance (CRM1) was correlated with p27(Kip1) in glioma. The aims of the present study were to investigate the expression of CRM1 and pSer10p27 and their functional roles in epithelial ovarian cancer (EOC) tissues. Using immunohistochemical analysis, CRM1 and pSer10p27 expression levels were shown to be associated with histologic stage and grade (P<0.05). High CRM1 and pSer10p27 expression levels were prognostic indicators of overall survival (P<0.05). Knockdown of CRM1 and pSer10p27 expression arrested cell cycle progression and inhibited the proliferation of SKOV3 cells both in vitro and in vivo. These data support the idea that pSer10p27 and CRM1 play cooperative roles in EOC. PMID:24018641

  14. KDM4B and KDM4A promote endometrial cancer progression by regulating androgen receptor, c-myc, and p27kip1

    PubMed Central

    Kwan, Suet-Ying; Chen, Limo; Chen, Jin-Hong; Ying, Zuo-Lin; Zhou, Ye; Gu, Wei; Wang, Li-Hua; Cheng, Wei-Wei; Zeng, Jianfang; Wan, Xiao-Ping; Mok, Samuel C.; Wong, Kwong-Kwok; Bao, Wei

    2015-01-01

    Epidemiological evidence suggests that elevated androgen levels and genetic variation related to the androgen receptor (AR) increase the risk of endometrial cancer (EC). However, the role of AR in EC is poorly understood. We report that two members of the histone demethylase KDM4 family act as major regulators of AR transcriptional activityin EC. In the MFE-296 cell line, KDM4B and AR upregulate c-myc expression, while in AN3CA cells KDM4A and AR downregulate p27kip1. Additionally, KDM4B expression is positively correlated with AR expression in EC cell lines with high baseline AR expression, while KDM4A and AR expression are positively correlated in low-AR cell lines. In clinical specimens, both KDM4B and KDM4A expression are significantly higher in EC tissues than that in normal endometrium. Finally, patients with alterations in AR, KDM4B, KDM4A, and c-myc have poor overall and disease-free survival rates. Together, these findings demonstrate that KDM4B and KDM4A promote EC progression by regulating AR activity. PMID:26397136

  15. KDM4B and KDM4A promote endometrial cancer progression by regulating androgen receptor, c-myc, and p27kip1.

    PubMed

    Qiu, Mei-Ting; Fan, Qiong; Zhu, Zhu; Kwan, Suet-Ying; Chen, Limo; Chen, Jin-Hong; Ying, Zuo-Lin; Zhou, Ye; Gu, Wei; Wang, Li-Hua; Cheng, Wei-Wei; Zeng, Jianfang; Wan, Xiao-Ping; Mok, Samuel C; Wong, Kwong-Kwok; Bao, Wei

    2015-10-13

    Epidemiological evidence suggests that elevated androgen levels and genetic variation related to the androgen receptor (AR) increase the risk of endometrial cancer (EC). However, the role of AR in EC is poorly understood. We report that two members of the histone demethylase KDM4 family act as major regulators of AR transcriptional activityin EC. In the MFE-296 cell line, KDM4B and AR upregulate c-myc expression, while in AN3CA cells KDM4A and AR downregulate p27kip1. Additionally, KDM4B expression is positively correlated with AR expression in EC cell lines with high baseline AR expression, while KDM4A and AR expression are positively correlated in low-AR cell lines. In clinical specimens, both KDM4B and KDM4A expression are significantly higher in EC tissues than that in normal endometrium. Finally, patients with alterations in AR, KDM4B, KDM4A, and c-myc have poor overall and disease-free survival rates. Together, these findings demonstrate that KDM4B and KDM4A promote EC progression by regulating AR activity. PMID:26397136

  16. Cyclin dependent kinase inhibitor p27Kip1 expression in normal and neoplastic cervical epithelium.

    PubMed Central

    Troncone, G; Vetrani, A; de Rosa, G; Gerbasio, D; Palombini, L

    1999-01-01

    AIM: To investigate whether there is loss of the p27Kip1 protein in developing cervical cancer and whether p27Kip1 immunoreactivity has any relation to the proliferative indicator Ki-67. METHODS: The expression of p27Kip1 and Ki-67 was assessed by immunohistochemistry in serial sections from normal epithelium (13), low grade (27) and high grade (19) squamous intraepithelial lesions (LSIL, HSIL), and invasive cervical cancer (23). In the SIL cases the presence of human papillomavirus (HPV) genomic sequences was assessed by in situ hybridisation. The results were evaluated by image analysis, and reported as mean score of the percentage of p27Kip1 and of Ki-67 positive cells in each histological group. RESULTS: In general, p27Kip1 immunostaining was related to squamous differentation, and was intense in normal epithelium (47%), while it was reduced in SIL lesions as an effect of the decreased number of differentiating cells. However, decrease in the p27Kip1 expression was more evident in LSIL (36%) than in HSIL (39%); in the latter, p27Kip1 had a different intraepithelial distribution in that the staining extended to the basal cells. The average levels of p27Kip1 were similar in SIL lesions associated to low, intermediate, and high risk HPV types. Compared with normal epithelium and dysplasia, invasive cancer showed significantly lower p27Kip1 levels (23%). There was no relation between p27Kip1 and Ki-67 labelling indices in any of the histological groups examined. CONCLUSIONS: A reduction in p27Kip1 protein occurs in cervical cancer independently of the proliferative status. The changes in p27Kip1 expression may be related to the unregulated kinetics of developing cervical cancer. Images PMID:10711250

  17. Down-regulation of both p21/Cip1 and p27/Kip1 produces a more aggressive prostate cancer phenotype

    PubMed Central

    Roy, Srirupa; Singh, Rana P.; Agarwal, Chapla; Siriwardana, Sunitha; Sclafani, Robert; Agarwal, Rajesh

    2009-01-01

    Roles of cyclin dependent kinase inhibitors, p21/Cip1 (p21) and p27/Kip1 (p27) in prostate cancer (PCa) progression is still not clear. Lower p27 protein expression in PCa tissues is often associated with poor prognosis, but prognostic significance of p21 is still controversial. Herein, we investigated the role of these molecules in determining PCa growth characteristics. We generated human PCa DU145 cell variants with knocked down levels of p21 (DU-p21) or p27 (DU-p27), or both (DU-p21+p27) via retroviral transduction of respective shRNAs and compared their various characteristics with empty vector-transduced DU145 (DU-EV) cells in vitro as well as in vivo. Knocking down either p21 or p27 did not show any significant change in doubling time, clonogenicity and cell cycle progression in DU145 cells, but simultaneous knock-down of both p21 and p27 significantly enhanced these parameters. In athymic mice, DU-p21+p27 tumors showed higher growth rate than the comparable growth of DU-EV, DU-p21 and DU-p27 tumors. Concurrently, DU-p21+p27 tumors had significantly higher proliferation rate, showing 54% and 48% increase in proliferating cell nuclear antigen (PCNA) and Ki-67-positive cells, respectively, compared to DU-EV tumors. DU-p21+p27 tumors also showed higher microvessel density and increased expression of vascular endothelial growth factor (VEGF). Proliferation and angiogenic status of DU-p21 and DU-p27 tumors was comparable to DU-EV tumors. Both in vitro and in vivo results implicate that p21 and p27 have compensatory roles in advanced prostate cancer cells, and ablation or down-modulation of both these molecules essentially enhances the aggressive prostate carcinoma phenotype. PMID:18583941

  18. Cell cycle arrest in Metformin treated breast cancer cells involves activation of AMPK, downregulation of cyclin D1, and requires p27Kip1 or p21Cip1

    PubMed Central

    Zhuang, Yongxian; Miskimins, W Keith

    2008-01-01

    Background The antihyperglycemic drug metformin may have beneficial effects on the prevention and treatment of cancer. Metformin is known to activate AMP-activated protein kinase (AMPK). It has also been shown to inhibit cyclin D1 expression and proliferation of some cultured cancer cells. However, the mechanisms of action by which metformin mediates cell cycle arrest are not completely understood. Results In this study, metformin was found to inhibit proliferation of most cultured breast cancer cell lines. This was independent of estrogen receptor, HER2, or p53 status. Inhibition of cell proliferation was associated with arrest within G0/G1 phase of the cell cycle. As in previous studies, metformin treatment led to activation of (AMPK) and downregulation of cyclin D1. However, these events were not sufficient for cell cycle arrest because they were also observed in the MDA-MB-231 cell line, which is not sensitive to growth arrest by metformin. In sensitive breast cancer lines, the reduction in cyclin D1 led to release of sequestered CDK inhibitors, p27Kip1 and p21Cip1, and association of these inhibitors with cyclin E/CDK2 complexes. The metformin-resistant cell line MDA-MB-231 expresses significantly lower levels of p27Kip1 and p21Cip1 than the metformin-sensitive cell line, MCF7. When p27Kip1 or p21Cip1 were overexpressed in MDA-MB-231, the cells became sensitive to cell cycle arrest in response to metformin. Conclusion Cell cycle arrest in response to metformin requires CDK inhibitors in addition to AMPK activation and cyclin D1 downregulation. This is of interest because many cancers are associated with loss or downregulation of CDK inhibitors and the results may be relevant to the development of anti-tumor reagents that target the AMPK pathway. PMID:19046439

  19. Caffeic acid phenethyl ester induced cell cycle arrest and growth inhibition in androgen-independent prostate cancer cells via regulation of Skp2, p53, p21Cip1 and p27Kip1

    PubMed Central

    Su, Liang-Cheng; Jiang, Shih Sheng; Chan, Tzu-Min; Chang, Chung-Ho; Chen, Li-Tzong; Kung, Hsing-Jien; Wang, Horng-Dar; Chuu, Chih-Pin

    2015-01-01

    Prostate cancer (PCa) patients receiving the androgen ablation therapy ultimately develop recurrent castration-resistant prostate cancer (CRPC) within 1–3 years. Treatment with caffeic acid phenethyl ester (CAPE) suppressed cell survival and proliferation via induction of G1 or G2/M cell cycle arrest in LNCaP 104-R1, DU-145, 22Rv1, and C4–2 CRPC cells. CAPE treatment also inhibited soft agar colony formation and retarded nude mice xenograft growth of LNCaP 104-R1 cells. We identified that CAPE treatment significantly reduced protein abundance of Skp2, Cdk2, Cdk4, Cdk7, Rb, phospho-Rb S807/811, cyclin A, cyclin D1, cyclin H, E2F1, c-Myc, SGK, phospho-p70S6kinase T421/S424, phospho-mTOR Ser2481, phospho-GSK3α Ser21, but induced p21Cip1, p27Kip1, ATF4, cyclin E, p53, TRIB3, phospho-p53 (Ser6, Ser33, Ser46, Ser392), phospho-p38 MAPK Thr180/Tyr182, Chk1, Chk2, phospho-ATM S1981, phospho-ATR S428, and phospho-p90RSK Ser380. CAPE treatment decreased Skp2 and Akt1 protein expression in LNCaP 104-R1 tumors as compared to control group. Overexpression of Skp2, or siRNA knockdown of p21Cip1, p27Kip1, or p53 blocked suppressive effect of CAPE treatment. Co-treatment of CAPE with PI3K inhibitor LY294002 or Bcl-2 inhibitor ABT737 showed synergistic suppressive effects. Our finding suggested that CAPE treatment induced cell cycle arrest and growth inhibition in CRPC cells via regulation of Skp2, p53, p21Cip1, and p27Kip1. PMID:25788262

  20. O-GlcNAc glycosylation of p27(kip1) promotes astrocyte migration and functional recovery after spinal cord contusion.

    PubMed

    Mao, Xingxing; Zhang, Dongmei; Tao, Tao; Liu, Xiaojuan; Sun, Xiaolei; Wang, Youhua; Shen, Aiguo

    2015-12-10

    Glial scar formation derived from astrocyte proliferation and migration influences the functional recovery after spinal cord injury. Cyclin-dependent kinase inhibitor p27(kip1), whose activity is closely related to its phosphorylation state, reportedly regulates astrocyte proliferation and migration. In this study, we reported that p27(Kip1) undergoes O-GlcNAc modification at Ser 2, Ser 110 and Thr 197. Inhibiting O-GlcNAcylation on Ser 2 by gene mutation (S2A) attenuated the phosphorylation of Ser 10, and vice versa. Interestingly, compared with wild type p27(Kip1), S2A p27(Kip1) displayed a decreased interaction with CRM1 and reduced nuclear export following serum starvation and release. In addition, the interaction between stathmin and S2A p27(Kip1) was also decreased. Cytoskeletal proteins microtubules appeared high density in astrocytes transfected with S2A p27(Kip1) especially at the leading edge of the scratch wound. Accordingly, scratch-wound assay revealed that the motility of astrocytes transfected with S2A p27(Kip1) was faster than that of control. Finally, we injected lentiviral vectors immediately after spinal cord contusion, and found the lesion volume of the rat injected with S2A p27(Kip1) was smaller than that of rat injected with wild type p27(Kip1). Besides, the BBB and CBS behavioral tests showed greater functional recovery in S2A p27(Kip1) treated rats. Taken together, our findings revealed a novel function of O-GlcNAc modification of p27(Kip1) in mediating astrocytes migration and functional recovery after spinal cord contusion. PMID:26562163

  1. Structural Basis of the Cks1-Dependent Recognition of P27Kip1 by the SCF skp2 Ubiquitin Ligase

    SciTech Connect

    Hao,B.; Zheng, N.; Schulman, B.; Wu, G.; Miller, J.; Pagano, M.; Pavletich, N.

    2005-01-01

    The ubiquitin-mediated proteolysis of the Cdk2 inhibitor p27(Kip1) plays a central role in cell cycle progression, and enhanced degradation of p27(Kip1) is associated with many common cancers. Proteolysis of p27(Kip1) is triggered by Thr187 phosphorylation, which leads to the binding of the SCF(Skp2) (Skp1-Cul1-Rbx1-Skp2) ubiquitin ligase complex. Unlike other known SCF substrates, p27(Kip1) ubiquitination also requires the accessory protein Cks1. The crystal structure of the Skp1-Skp2-Cks1 complex bound to a p27(Kip1) phosphopeptide shows that Cks1 binds to the leucine-rich repeat (LRR) domain and C-terminal tail of Skp2, whereas p27(Kip1) binds to both Cks1 and Skp2. The phosphorylated Thr187 side chain of p27(Kip1) is recognized by a Cks1 phosphate binding site, whereas the side chain of an invariant Glu185 inserts into the interface between Skp2 and Cks1, interacting with both. The structure and biochemical data support the proposed model that Cdk2-cyclin A contributes to the recruitment of p27(Kip1) to the SCF(Skp2)-Cks1 complex.

  2. The interplay between TEAD4 and KLF5 promotes breast cancer partially through inhibiting the transcription of p27Kip1

    PubMed Central

    Zhou, Zhongmei; Zhang, Hailin; Liu, Rong; Wu, Jing; Qin, Junying; Ma, Yun; Chen, Liang; Li, Shumo; Chen, Wenlin; Li, Fubing; Shi, Peiguo; Wu, Yingying; Shen, Jian; Chen, Ceshi

    2015-01-01

    Growing evidence suggests that YAP/TAZ are mediators of the Hippo pathway and promote breast cancer. However, the roles of YAP/TAZ transcription factor partners TEADs in breast cancer remain unclear. Here we found that TEAD4 was expressed in breast cancer cell lines, especially in triple negative breast cancers (TNBC) cell lines. TEAD4 binds to KLF5. Knockdown of either TEAD4 or KLF5 in HCC1937 and HCC1806 cells induced the expression of CDK inhibitor p27. Depletion of either TEAD4 or KLF5 activated the p27 gene promoter and increased the p27 mRNA levels. Depletion of p27 partially prevents growth inhibition caused by TEAD4 and KLF5 knockdown. TEAD4 overexpression stimulated proliferation in vitro and tumor growth in mice, while stable knockdown of TEAD4 inhibited proliferation in vitro and tumor growth in mice. Thus TEAD4 and KLF5, in collaboration, promoted TNBC cell proliferation and tumor growth in part by inhibiting p27 gene transcription. TEAD4 is a potential target and biomarker for the development of novel therapeutics for breast cancer. PMID:25970772

  3. Thrombin Induces Tumor Cell Cycle Activation and Spontaneous Growth by Down-regulation of p27Kip1, in Association with the Up-regulation of Skp2 and MiR-222

    PubMed Central

    Hu, Liang; Ibrahim, Sherif; Liu, Cynthia; Skaar, Jeffrey; Pagano, Michele; Karpatkin, Simon

    2009-01-01

    The effect of thrombin on tumor cell cycle activation and spontaneous growth was examined in synchronized serum-starved tumor cell lines and a model of spontaneous prostate cancer development in TRAMP mice. BrdUrd incorporation and propidium iodide staining of prostate LNCaP cells arrested in G0 and treated with thrombin or serum revealed a 48- and 29-fold increase in S phase cells, respectively, at 8 hours. Similar results were obtained with TRAMP cells and a glioblastoma cell line, T98G. Cell cycle kinases and inhibitors in synchronized tumor cells revealed high levels of p27Kip1 and low levels of Skp2 and cyclins D1 and A. Addition of thrombin, TFLLRN, or serum down-regulated p27Kip1 with concomitant induction of Skp2, Cyclin D1, and Cyclin A with similar kinetics. LNCaP p27Kip1-transfected cells or Skp2 knockdown cells were refractory to thrombin-induced cell cycle activation. MicroRNA 222, an inhibitor of p27Kip1, was robustly up-regulated by thrombin. The in vitro observations were tested in vivo with transgenic TRAMP mice. Repetitive thrombin injection enhanced prostate tumor volume 6- to 8-fold (P < 0.04). Repetitive hirudin, a specific potent antithrombin, decreased tumor volume 13- to 24-fold (P < 0.04). Thus, thrombin stimulates tumor cell growth in vivo by down-regulation of p27Kip1. PMID:19351827

  4. A new tumour suppression mechanism by p27Kip1: EGFR down-regulation mediated by JNK/c-Jun pathway inhibition

    PubMed Central

    Fang, Yong; Wang, Yihong; Wang, Yulei; Meng, Yan; Zhu, Junlan; Jin, Honglei; Li, Jingxia; Zhang, Dongyun; Yu, Yonghui; Wu, Xue-Ru; Huang, Chuanshu

    2014-01-01

    p27Kip1 is a potent inhibitor of cyclin-dependent kinases that drive G1-to-S cell-cycle transition. Reduced p27Kip1 expression is prevalent in a wide range of human tumours; however, the exact mechanism(s) of p27Kip1-mediated tumour suppression remains obscure. In the present study, we identified a close inverse relationship between p27Kip1 and EGFR (epidermal growth factor receptor) expression: the parental T24 human bladder cancer cells had high p27Kip1 expression but low EGFR expression and, in striking contrast, the metastatic derivative of T24 (T24T) had low p27Kip1 expression but high EGFR expression. This relationship was also found in various human cancer tissues, and was not only just correlative but also causal; depletion of p27Kip1 in MEF (mouse embryonic fibroblast) cells resulted in markedly elevated EGFR expression, a result reproducible with an Egfr promoter-luciferase reporter in both T24 and MEF cells, suggesting transcriptional repression of EGFR by p27Kip1. Indeed, p27Kip1 was found to regulate EGFR expression via the JNK (c-Jun N-terminal kinase)/c-Jun transcription factor: p27Kip1 deficiency activated JNK/c-Jun, whereas inhibition of JNK/c-Jun by dominant-negative mutants dramatically repressed Egfr transcription. Furthermore, the proximal promoter of the Egfr gene was crucial for its transcription, where the recruiting activity of c-Jun was much greater in p27Kip1−/− cells than in p27Kip1+/+ cells. Introduction of GFP–p27Kip1 into T24T cells suppressed JNK/c-Jun activation, EGFR expression and anchorage-independent growth. The results of the present study demonstrate that p27Kip1 suppresses JNK/c-Jun activation and EGFR expression in MEFs and human bladder cancer cells, and the results obtained are consistent with those from human cancer specimens. The present study provides new insights into p27Kip1 suppression of cancer cell growth, migration and metastasis. PMID:25121353

  5. p27Kip1 Is Required to Mediate a G1 Cell Cycle Arrest Downstream of ATM following Genotoxic Stress.

    PubMed

    Cassimere, Erica K; Mauvais, Claire; Denicourt, Catherine

    2016-01-01

    The DNA damage response (DDR) is a coordinated signaling network that ensures the maintenance of genome stability under DNA damaging stress. In response to DNA lesions, activation of the DDR leads to the establishment of cell cycle checkpoints that delay cell-cycle progression and allow repair of the defects. The tumor suppressor p27Kip1 is a cyclin-CDK inhibitor that plays an important role in regulating quiescence in a variety of tissues. Several studies have suggested that p27Kip1 also plays a role in the maintenance of genomic integrity. Here we demonstrate that p27Kip1 is essential for the establishment of a G1 checkpoint arrest after DNA damage. We also uncovered that ATM phosphorylates p27Kip1 on a previously uncharacterized residue (Ser-140), which leads to its stabilization after induction of DNA double-strand breaks. Inhibition of this stabilization by replacing endogenous p27Kip1 with a Ser-140 phospho-mutant (S140A) significantly sensitized cells to IR treatments. Our findings reveal a novel role for p27Kip1 in the DNA damage response pathway and suggest that part of its tumor suppressing functions relies in its ability to mediate a G1 arrest after the induction of DNA double strand breaks. PMID:27611996

  6. Huntingtin with an expanded polyglutamine repeat affects the Jab1-p27(Kip1) pathway.

    PubMed

    Cong, S Y; Pepers, B A; Zhou, T T; Kerkdijk, H; Roos, R A; van Ommen, G J; Dorsman, J C

    2012-06-01

    Expansion of polyglutamine repeats is the cause of at least nine inherited human neurodegenerative disorders, including Huntington's disease (HD). It is widely accepted that deregulation of the transcriptional coactivator CBP by expanded huntingtin (htt) plays an important role in HD molecular pathogenesis. In this study, we report on a novel target of expanded polyglutamine stretches, the transcriptional coactivator Jun activation domain-binding protein 1 (Jab1), which shares DNA-sequence-specific transcription factor targets with CBP. Jab1 also plays a major role in the degradation of the cyclin-dependent-kinase inhibitor and putative transcription cofactor p27(Kip1). We found that Jab1 accumulates in aggregates when co-expressed with either expanded polyglutamine stretches or N-terminal fragments of mutant htt. In addition, the coactivator function of Jab1 was suppressed both by aggregated expanded polyglutamine solely and by mutant htt. Inhibition by mutant htt even preceded the appearance of microscopic aggregation. In an exon 1 HD cell model, we found that endogenous Jab1 could be recruited into aggregates and that this was accompanied by the accumulation of p27(Kip1). Accumulation of p27(Kip1) was also found in brains derived from HD patients. The repression of Jab1 by various mechanisms coupled with an increase of p27(Kip1) at late stages may have important transcriptional effects. In addition, the interference with the Jab1-p27(Kip1) pathway may contribute to the observed lower incidence of cancer in HD patients and may also be relevant for the understanding of the molecular pathogenesis of polyglutamine disorders in general. PMID:22426400

  7. p27Kip1 expression as a prognostic marker for squamous cell carcinoma of the head and neck

    PubMed Central

    DE ALMEIDA, MIGUEL REIS; PÉREZ-SAYÁNS, MARIO; SUÁREZ-PEÑARANDA, JOSÉ MANUEL; SOMOZA-MARTÍN, JOSÉ MANUEL; GARCÍA-GARCÍA, ABEL

    2015-01-01

    Regulation of the cell cycle is essential for carcinogenesis. The cell cycle is controlled by cyclin-dependent kinases (CDKs), which are upregulated by cyclins and downregulated by CDK inhibitors (CDKIs). Decreased p27Kip1 expression has been associated with survival rate, tumor size, histological differentiation and the presence of lymph node metastasis in patients with various types of cancer. The aim of the current study is to provide a literature review on the association between p27Kip1 expression and the clinical and pathological aspects of head and neck squamous cell carcinoma (HNSCC), and the expression of other CDKIs of the Cip/Kip family and cyclins. Throughout the literature, different methodologies were used to determine the immunohistochemical expression of p27Kip1; thus, results concerning p27Kip1 expression in HNSCC vary widely. However, it has now been confirmed that p27Kip1 is underexpressed in SCC cells. p27 may be a promising marker for determining the prognosis of HNSCC, despite the marked variability of the results obtained. An association between p27 expression and survival rate, time to recurrence and tumor stage has been observed. Based on the information currently available, it is premature to recommend the analysis of p27Kip1 expression in guiding HNSCC treatment planning. However, although relatively unstudied, the correlation between p27Kip1 expression and other tumor suppressor genes may turn out to be important in determining the prognosis of HNSCC. Further prospective studies utilizing standardized laboratory methodologies and statistics that facilitate meta-analyses are required to confirm this proposal. PMID:26722226

  8. Histone Deacetylase Inhibitors Increase p27Kip1 by Affecting Its Ubiquitin-Dependent Degradation through Skp2 Downregulation

    PubMed Central

    Borriello, Adriana; Naviglio, Silvio; Bencivenga, Debora; Caldarelli, Ilaria; Tramontano, Annunziata; Speranza, Maria Carmela; Stampone, Emanuela; Sapio, Luigi; Negri, Aide; Oliva, Adriana; Sinisi, Antonio Agostino; Spina, Annamaria; Della Ragione, Fulvio

    2016-01-01

    Histone deacetylase inhibitors (HDACIs) represent an intriguing class of pharmacologically active compounds. Currently, some HDACIs are FDA approved for cancer therapy and many others are in clinical trials, showing important clinical activities at well tolerated doses. HDACIs also interfere with the aging process and are involved in the control of inflammation and oxidative stress. In vitro, HDACIs induce different cellular responses including growth arrest, differentiation, and apoptosis. Here, we evaluated the effects of HDACIs on p27Kip1, a key cyclin-dependent kinase inhibitor (CKI). We observed that HDACI-dependent antiproliferative activity is associated with p27Kip1 accumulation due to a reduced protein degradation. p27Kip1 removal requires a preliminary ubiquitination step due to the Skp2-SCF E3 ligase complex. We demonstrated that HDACIs increase p27Kip1 stability through downregulation of Skp2 protein levels. Skp2 decline is only partially due to a reduced Skp2 gene expression. Conversely, the protein decrease is more profound and enduring compared to the changes of Skp2 transcript. This argues for HDACIs effects on Skp2 protein posttranslational modifications and/or on its removal. In summary, we demonstrate that HDACIs increase p27Kip1 by hampering its nuclear ubiquitination/degradation. The findings might be of relevance in the phenotypic effects of these compounds, including their anticancer and aging-modulating activities. PMID:26682002

  9. Reduced levels of the cell-cycle inhibitor p27Kip1 in epithelial dysplasia and carcinoma of the oral cavity.

    PubMed Central

    Jordan, R. C.; Bradley, G.; Slingerland, J.

    1998-01-01

    Recent studies have shown that the cyclin-dependent kinase (cdk) inhibitors play important roles in cell cycle progression in normal cells. Alterations in the cdk inhibitors also appear to be important in cancer development in a number of human tumors. p27Kip1 is a member of the CIP/KIP family of cdk inhibitors that negatively regulates cyclin-cdk complexes. Reduced levels of p27Kip1 protein have been identified in a number of human cancers, and in some cases reduced p27Kip1 is associated with an increase in proliferative fraction. In the present study, we examined p27Kip1 protein by immunohistochemistry in 10 normal and 36 dysplastic epithelia and in 8 squamous cell carcinomas from one anatomical site within the oral cavity, the floor of the mouth. Proliferative activity was assessed in serial sections by determining the expression of the cell cycle proteins Ki-67 and cyclin A. p27kip1 protein was significantly reduced in oral dysplasias and carcinomas compared with that in normal epithelial controls. In addition, there was a significant reduction in p27Kip1 protein between low- and high-grade dysplasias, suggesting that changes in p27Kip1 expression may be an early event in oral carcinogenesis. There was increasing expression of Ki-67 and cyclin A proteins with increasingly severe grades of dysplasia compared with normal controls. Although there was a strong correlation between Ki-67 and cyclin A scores (r2= 0.61) for all categories of disease, there was a weak negative correlation between Ki-67 and p27Kip1 levels (r2 = 0.29) and between cyclin A and p27Kip1 levels (r2 = 0.25). In conclusion, this study has found that a reduction in the proportion of cells expressing p27Kip1 protein is frequently associated with oral dysplasia and carcinoma from the floor of the mouth. Furthermore, reductions in p27Kip1 levels are associated with increased cell proliferation, although other changes likely contribute to altered cell kinetics during carcinogenesis at this site

  10. Homeodomain-interacting protein kinase-2 stabilizes p27(kip1) by its phosphorylation at serine 10 and contributes to cell motility.

    PubMed

    Pierantoni, Giovanna Maria; Esposito, Francesco; Tornincasa, Mara; Rinaldo, Cinzia; Viglietto, Giuseppe; Soddu, Silvia; Fusco, Alfredo

    2011-08-19

    HIPK2 is a serine/threonine kinase that acts as a coregulator of an increasing number of factors involved in cell survival and proliferation during development and in response to different types of stress. Here we report on a novel target of HIPK2, the cyclin-dependent kinase inhibitor p27(kip1). HIPK2 phosphorylates p27(kip1) in vitro and in vivo at serine 10, an event that accounts for 80% of the total p27(kip1) phosphorylation and plays a crucial role in the stability of the protein. Indeed, HIPK2 depletion by transient or stable RNA interference in tumor cells of different origin was consistently associated with strong reduction of p27(kip1) phosphorylation at serine 10 and of p27(kip1) stability. An initial evaluation of the functional relevance of this HIPK2-mediated regulation of p27(kip1) revealed a contribution to cell motility, rather than to cell proliferation, but only in cells that do not express wild-type p53. PMID:21715331

  11. Nanog induces suppression of senescence through downregulation of p27KIP1 expression

    PubMed Central

    Münst, Bernhard; Thier, Marc Christian; Winnemöller, Dirk; Helfen, Martina; Thummer, Rajkumar P.; Edenhofer, Frank

    2016-01-01

    ABSTRACT A comprehensive analysis of the molecular network of cellular factors establishing and maintaining pluripotency as well as self renewal of pluripotent stem cells is key for further progress in understanding basic stem cell biology. Nanog is necessary for the natural induction of pluripotency in early mammalian development but dispensable for both its maintenance and its artificial induction. To gain further insight into the molecular activity of Nanog, we analyzed the outcomes of Nanog gain-of-function in various cell models employing a recently developed biologically active recombinant cell-permeant protein, Nanog-TAT. We found that Nanog enhances the proliferation of both NIH 3T3 and primary fibroblast cells. Nanog transduction into primary fibroblasts results in suppression of senescence-associated β-galactosidase activity. Investigation of cell cycle factors revealed that transient activation of Nanog correlates with consistent downregulation of the cell cycle inhibitor p27KIP1 (also known as CDKN1B). By performing chromatin immunoprecipitation analysis, we confirmed bona fide Nanog-binding sites upstream of the p27KIP1 gene, establishing a direct link between physical occupancy and functional regulation. Our data demonstrates that Nanog enhances proliferation of fibroblasts through transcriptional regulation of cell cycle inhibitor p27 gene. PMID:26795560

  12. MicroRNAs 221 and 222 target p27Kip1 in Marek's disease virus-transformed tumour cell line MSB-1.

    PubMed

    Lambeth, Luke S; Yao, Yongxiu; Smith, Lorraine P; Zhao, Yuguang; Nair, Venugopal

    2009-05-01

    MicroRNAs (miRNAs) are a class of short RNAs that function as post-transcriptional suppressors of protein expression and are involved in a variety of biological processes, including oncogenesis. Several recent studies have implicated the involvement of miR-221 and miR-222 in tumorigenesis as these miRNAs are upregulated in a number of cancers and affect the expression of cell cycle regulatory proteins such as the cyclin-dependent kinase (cdk) inhibitor p27(Kip1). Marek's disease virus (MDV) is a highly oncogenic herpesvirus that affects poultry, causing acute neoplastic disease with lymphomatous lesions in several organs. MDV-encoded oncogenes such as Meq are directly implicated in the neoplastic transformation of T cells and have been well studied. More recently, however, the involvement of both host and virus-encoded miRNAs in the induction of MD lymphomas is being increasingly recognized. We analysed the miRNA expression profiles in the MDV-transformed lymphoblastoid cell line MSB-1 and found that endogenous miRNAs miR-221 and miR-222 were significantly upregulated. Demonstration of the conserved binding sites for these miRNAs in the chicken p27(Kip1) 3'-untranslated region sequence and the repression of luciferase activity of reporter constructs indicated that miR-221 and miR-222 target p27(Kip1) in these cells. We also found that overexpression of miR-221 and miR-222 decreased p27(Kip1) levels and that treatment with retrovirally expressed antagomiRs partially alleviated this suppression. These data show that an oncogenic herpesvirus, as in the case of many cancers, can exploit the miRNA machinery for suppressing cell cycle regulatory molecules such as p27(Kip1) in the induction and progression of T-cell lymphomas. PMID:19264608

  13. p27(Kip1) participates in the regulation of endoreplication in differentiating chick retinal ganglion cells.

    PubMed

    Ovejero-Benito, María C; Frade, José M

    2015-01-01

    Nuclear DNA duplication in the absence of cell division (i.e. endoreplication) leads to somatic polyploidy in eukaryotic cells. In contrast to some invertebrate neurons, whose nuclei may contain up to 200,000-fold the normal haploid DNA amount (C), polyploid neurons in higher vertebrates show only 4C DNA content. To explore the mechanism that prevents extra rounds of DNA synthesis in these latter cells we focused on the chick retina, where a population of tetraploid retinal ganglion cells (RGCs) has been described. We show that differentiating chick RGCs that express the neurotrophic receptors p75 and TrkB while lacking retinoblastoma protein, a feature of tetraploid RGCs, also express p27(Kip1). Two different short hairpin RNAs (shRNA) that significantly downregulate p27(Kip1) expression facilitated DNA synthesis and increased ploidy in isolated chick RGCs. Moreover, this forced DNA synthesis could not be prevented by Cdk4/6 inhibition, thus suggesting that it is triggered by a mechanism similar to endoreplication. In contrast, p27(Kip1) deficiency in mouse RGCs does not lead to increased ploidy despite previous observations have shown ectopic DNA synthesis in RGCs from p27(Kip1-/-) mice. This suggests that a differential mechanism is used for the regulation of neuronal endoreplication in mammalian versus avian RGCs. PMID:25946375

  14. PRMT5 Is Upregulated in Malignant and Metastatic Melanoma and Regulates Expression of MITF and p27Kip1

    PubMed Central

    Nicholas, Courtney; Yang, Jennifer; Peters, Sara B.; Bill, Matthew A.; Baiocchi, Robert A.; Yan, Fengting; Sïf, Saïd; Tae, Sookil; Gaudio, Eugenio; Wu, Xin; Grever, Michael R.; Young, Gregory S.; Lesinski, Gregory B.

    2013-01-01

    Protein arginine methyltransferase-5 (PRMT5) is a Type II arginine methyltransferase that regulates various cellular functions. We hypothesized that PRMT5 plays a role in regulating the growth of human melanoma cells. Immunohistochemical analysis indicated significant upregulation of PRMT5 in human melanocytic nevi, malignant melanomas and metastatic melanomas as compared to normal epidermis. Furthermore, nuclear PRMT5 was significantly decreased in metastatic melanomas as compared to primary cutaneous melanomas. In human metastatic melanoma cell lines, PRMT5 was predominantly cytoplasmic, and associated with its enzymatic cofactor Mep50, but not STAT3 or cyclin D1. However, histologic examination of tumor xenografts from athymic mice revealed heterogeneous nuclear and cytoplasmic PRMT5 expression. Depletion of PRMT5 via siRNA inhibited proliferation in a subset of melanoma cell lines, while it accelerated growth of others. Loss of PRMT5 also led to reduced expression of MITF (microphthalmia-associated transcription factor), a melanocyte-lineage specific oncogene, and increased expression of the cell cycle regulator p27Kip1. These results are the first to report elevated PRMT5 expression in human melanoma specimens and indicate this protein may regulate MITF and p27Kip1 expression in human melanoma cells. PMID:24098663

  15. Cell-permeable Carboxyl-terminal p27Kip1 Peptide Exhibits Anti-tumor Activity by Inhibiting Pim-1 Kinase*

    PubMed Central

    Morishita, Daisuke; Takami, Miho; Yoshikawa, Seiko; Katayama, Ryohei; Sato, Shigeo; Kukimoto-Niino, Mutsuko; Umehara, Takashi; Shirouzu, Mikako; Sekimizu, Kazuhisa; Yokoyama, Shigeyuki; Fujita, Naoya

    2011-01-01

    The incidence and death rate of prostate cancer is increasing rapidly. In addition, the low sensitivity of prostate cancer to chemotherapy makes it difficult to treat this condition. The serine/threonine kinase Pim-1 plays an important role in cell cycle progression and apoptosis inhibition, resulting in prostate tumorigenesis. Therefore, Pim-1 inhibition has been expected to be an attractive target for developing new anti-cancer drugs. However, no small compounds targeting Pim-1 have progressed to clinical use because of their lack of specificity. Here, we have reported a new cell-permeable Pim-1 inhibitory p27Kip1 peptide that could interfere with the binding of Pim-1 to its substrates and act as an anti-cancer drug. The peptide could bind to Pim-1 and inhibit phosphorylation of endogenous p27Kip1 and Bad by Pim-1. Treatment of prostate cancer with the peptide induces G1 arrest and subsequently apoptosis in vitro. However, the peptide showed almost no growth inhibitory or apoptosis-inducing effects in normal cells. The peptide could inhibit tumor growth in in vivo prostate cancer xenograft models. Moreover, the peptide treatment could overcome resistance to taxol, one of the first line chemotherapeutic agents for prostate cancer, and a combination of the peptide with taxol synergistically inhibited prostate cancer growth in vivo. These results indicate that a Pim-1 inhibitory p27Kip1 peptide could be developed as an anti-cancer drug against prostate cancer. PMID:21062737

  16. Augmentation of antitumor activity of 5-fluorouracil by interferon alpha is associated with up-regulation of p27Kip1 in human hepatocellular carcinoma cells.

    PubMed

    Eguchi, H; Nagano, H; Yamamoto, H; Miyamoto, A; Kondo, M; Dono, K; Nakamori, S; Umeshita, K; Sakon, M; Monden, M

    2000-07-01

    Several clinical trials have demonstrated the effectiveness of combination therapy with 5-fluorouracil (5-FU) and IFN-alpha in colon cancer, hepatocellular carcinoma (HCC), and other malignancies. In our preliminary clinical studies, we have observed outstanding effects with this combination therapy in patients with advanced HCC. However, the underlying mechanism by which IFN-alpha modulates the effects of 5-FU is unknown. We, therefore, conducted a mechanistic study using two HCC cell lines, PLC/PRF/5 and HuH7. IFN-alpha significantly enhanced the growth inhibitory effect of 5-FU in PLC/PRF/5 cells but not in HuH7 cells, and the isobolographic analysis indicated that this effect was synergistic. Flow cytometric analysis showed a delay in the progression of G0-G1 to S phase in PLC/PRF/5, and a sustained, induction of the cyclin-dependent kinase inhibitor p27-Kip1 and down-regulation of cyclin D1 was observed. Moreover, increased expression of p27Kip1 was associated with reduced CDK-2-associated kinase activity. Another difference in the two cell types was that PLC/PRF/5 expressed abundant IFN receptors, but HuH7 did not. Apoptosis assays were not helpful in explaining the mechanism. Our results suggest that the synergistic effects of 5-FU and IFN-alpha may in part be attributable to alterations in cell cycle progression via up-regulation of p27Kip1. PMID:10914738

  17. Insect peptide CopA3-induced protein degradation of p27Kip1 stimulates proliferation and protects neuronal cells from apoptosis

    SciTech Connect

    Nam, Seung Taek; Kim, Dae Hong; Lee, Min Bum; Nam, Hyo Jung; Kang, Jin Ku; Park, Mi Jung; Lee, Ik Hwan; Seok, Heon; Lee, Dong Gun; Hwang, Jae Sam; Kim, Ho

    2013-07-19

    Highlights: •CopA3 peptide isolated from the Korean dung beetle has antimicrobial activity. •Our study reported that CopA3 has anticancer and immunosuppressive effects. •We here demonstrated that CopA3 has neurotropic and neuroprotective effects. •CopA3 degrades p27Kip1 protein and this mediates effects of CopA3 on neuronal cells. -- Abstract: We recently demonstrated that the antibacterial peptide, CopA3 (a D-type disulfide dimer peptide, LLCIALRKK), inhibits LPS-induced macrophage activation and also has anticancer activity in leukemia cells. Here, we examined whether CopA3 could affect neuronal cell proliferation. We found that CopA3 time-dependently increased cell proliferation by up to 31 ± 2% in human neuroblastoma SH-SY5Y cells, and up to 29 ± 2% in neural stem cells isolated from neonatal mouse brains. In both cell types, CopA3 also significantly inhibited the apoptosis and viability losses caused by 6-hydroxy dopamine (a Parkinson disease-mimicking agent) and okadaic acid (an Alzheimer’s disease-mimicking agent). Immunoblotting revealed that the p27Kip1 protein (a negative regulator of cell cycle progression) was markedly degraded in CopA3-treated SH-SY5Y cells. Conversely, an adenovirus expressing p27Kip1 significantly inhibited the antiapoptotic effects of CopA3 against 6-hydroxy dopamine- and okadaic acid-induced apoptosis, and decreased the neurotropic effects of CopA3. These results collectively suggest that CopA3-mediated protein degradation of p27Kip1 may be the main mechanism through which CopA3 exerts neuroprotective and neurotropic effects.

  18. Differential modulation of paclitaxel-mediated apoptosis by p21Waf1 and p27Kip1.

    PubMed

    Schmidt, M; Lu, Y; Liu, B; Fang, M; Mendelsohn, J; Fan, Z

    2000-05-11

    The impact of the cyclin dependent kinase (CDK) inhibitors p21Waf1 and p27Kip1 on paclitaxel-mediated cytotoxicity was investigated in RKO human colon adenocarcinoma cells with the ecdysone-inducible expression of p21Waf1 or p27Kip1. Ectopic expression of p27Kip1 arrested cells at G1 phase, whereas p21Waf1 expression arrested cells at G1 and G2. Expression of p21Waf1 after paclitaxel treatment produced much greater resistance to paclitaxel than did expression of p27Kip1. We attributed this difference to the additional block at G2 induced by p21Waf1, which prevented cells from entering M phase and becoming paclitaxel susceptible. Expression of p21Waf1 inhibited p34cdc2 activity and markedly reduced paclitaxel-mediated mitotic arrest, from 87.5 to 23%. In contrast, p27Kip1 expression also inhibited p34cdc2 but reduced mitotic arrest only slightly, from 87. 4 to 74.5%. We concluded that the G2 block produced by p21Waf1, but not by p27Kip1, contributed to their unequal modulation of sensitivity to paclitaxel-mediated apoptosis in RKO cells, and there is no causal relationship between paclitaxel-mediated cytotoxicity and elevation of p34cdc2 activity. PMID:10828884

  19. The Relevance of Women's Diseases, Jun Activation-domain Binding Protein 1 (JAB1) and p27kip1

    PubMed Central

    Kim, Mijin; Lee, Hae-Hyeog

    2016-01-01

    The Jun activation-domain binding protein 1 (Jab1) recognize a potential coactivator of activator protein 1 (AP-1) such as c-fos, c-jun transcription factor and the fifth subunit of the COP9 signalosome complex. Also, Jab1 activate the c-jun gene resulted cell proliferation. Not only a powerful tumor suppressor but also regulator of apoptosis negative cdk inhibitor p27kip1 are involved in the cell cycle. This is Jab1 and p27kip1 interact with each other, Jab1 accelerate p27kip1 from nuclear to cytoplasm through ubiquitin/proteasome pathway. However, information about the relationship between Jab1 and p27kip1 is not known much. Taken together, the results of this study identify function and structure of Jab1 and p27kip1 were described in a recent article on the basis of relevant. Besides Jab1 and p27kip1 will organize the relationship between the disease and women. PMID:27152307

  20. Cell cycle regulatory protein p27KIP1 is a substrate and interacts with the protein kinase CK2.

    PubMed

    Tapia, Julio C; Bolanos-Garcia, Victor M; Sayed, Muhammed; Allende, Catherine C; Allende, Jorge E

    2004-04-01

    The protein kinase CK2 is constituted by two catalytic (alpha and/or alpha') and two regulatory (beta) subunits. CK2 phosphorylates more than 300 proteins with important functions in the cell cycle. This study has looked at the relation between CK2 and p27(KIP1), which is a regulator of the cell cycle and a known inhibitor of cyclin-dependent kinases (Cdk). We demonstrated that in vitro recombinant Xenopus laevis CK2 can phosphorylate recombinant human p27(KIP1), but this phosphorylation occurs only in the presence of the regulatory beta subunit. The principal site of phosphorylation is serine-83. Analysis using pull down and surface plasmon resonance (SPR) techniques showed that p27(KIP1) interacts with the beta subunit through two domains present in the amino and carboxyl ends, while CD spectra showed that p27(KIP1) phosphorylation by CK2 affects its secondary structure. Altogether, these results suggest that p27(KIP1) phosphorylation by CK2 probably involves a docking event mediated by the CK2beta subunit. The phosphorylation of p27(KIP1) by CK2 may affect its biological activity. PMID:15034923

  1. CRM-1 knockdown inhibits extrahepatic cholangiocarcinoma tumor growth by blocking the nuclear export of p27Kip1.

    PubMed

    Luo, Jian; Chen, Yongjun; Li, Qiang; Wang, Bing; Zhou, Yanqiong; Lan, Hongzhen

    2016-08-01

    Cholangiocarcinoma is a deadly disease which responds poorly to surgery and conventional chemotherapy or radiotherapy. Early diagnosis is difficult due to the anatomical and biological characteristics of cholangiocarcinoma. Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a cyclin‑dependent kinase inhibitor and in the present study, we found that p27Kip1 expression was suppressed in the nucleus and increased in the cytoplasm in 53 samples of cholangiocarcinoma from patients with highly malignant tumors (poorly-differentiated and tumor-node-metastsis (TNM) stage III-IV) compared with that in samples from 10 patients with chronic cholangitis. The expression of phosphorylated (p-)p27Kip1 (Ser10), one of the phosphorylated forms of p27Kip1, was increased in the patient samples with increasing malignancy and clinical stage. Coincidentally, chromosome region maintenance 1 (CRM-1; also referred to as exportin 1 or Xpo1), a critical protein responsible for protein translocation from the nucleus to the cytoplasm, was also overexpressed in the tumor samples which were poorly differentiated and of a higher clinical stage. Through specific short hairpin RNA (shRNA)-mediated knockdown of CRM-1 in the cholangiocarcinoma cell line QBC939, we identified an elevation of cytoplasmic p27Kip1 and a decrease of nuclear p27Kip1. Furthermore, the viability and colony formation ability of QBC939 cells was largely reduced with G1 arrest. Consistent with the findings of the in vitro experiments, in a xenograft mouse model, the tumors formed in the CRM-1 knockdown group were markedly smaller and weighed less than those in the control group in vivo. Taken together, these findings demonstrated that the interplay between CRM-1 and p27Kip1 may provide potentially potent biomarkers and functional targets for the development of future cholangiocarcinoma treatments. PMID:27279267

  2. CRM-1 knockdown inhibits extrahepatic cholangiocarcinoma tumor growth by blocking the nuclear export of p27Kip1

    PubMed Central

    Luo, Jian; Chen, Yongjun; Li, Qiang; Wang, Bing; Zhou, Yanqiong; Lan, Hongzhen

    2016-01-01

    Cholangiocarcinoma is a deadly disease which responds poorly to surgery and conventional chemotherapy or radiotherapy. Early diagnosis is difficult due to the anatomical and biological characteristics of cholangiocarcinoma. Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a cyclin-dependent kinase inhibitor and in the present study, we found that p27Kip1 expression was suppressed in the nucleus and increased in the cytoplasm in 53 samples of cholangiocarcinoma from patients with highly malignant tumors (poorly-differentiated and tumor-node-metastsis (TNM) stage III–IV) compared with that in samples from 10 patients with chronic cholangitis. The expression of phosphorylated (p-)p27Kip1 (Ser10), one of the phosphorylated forms of p27Kip1, was increased in the patient samples with increasing malignancy and clinical stage. Coincidentally, chromosome region maintenance 1 (CRM-1; also referred to as exportin 1 or Xpo1), a critical protein responsible for protein translocation from the nucleus to the cytoplasm, was also overexpressed in the tumor samples which were poorly differentiated and of a higher clinical stage. Through specific short hairpin RNA (shRNA)-mediated knockdown of CRM-1 in the cholangiocarcinoma cell line QBC939, we identified an elevation of cytoplasmic p27Kip1 and a decrease of nuclear p27Kip1. Furthermore, the viability and colony formation ability of QBC939 cells was largely reduced with G1 arrest. Consistent with the findings of the in vitro experiments, in a xenograft mouse model, the tumors formed in the CRM-1 knockdown group were markedly smaller and weighed less than those in the control group in vivo. Taken together, these findings demonstrated that the interplay between CRM-1 and p27Kip1 may provide potentially potent biomarkers and functional targets for the development of future cholangiocarcinoma treatments. PMID:27279267

  3. Epidermal growth factor upregulates Skp2/Cks1 and p27kip1 in human extrahepatic cholangiocarcinoma cells

    PubMed Central

    Kim, Ja-yeon; Kim, Hong Joo; Park, Jung Ho; Park, Dong Il; Cho, Yong Kyun; Sohn, Chong Il; Jeon, Woo Kyu; Kim, Byung Ik; Kim, Dong Hoon; Chae, Seoung Wan; Sohn, Jin Hee

    2014-01-01

    AIM: To evaluate the expression status of S-phase kinase-associated protein 2 (Skp2)/cyclin-dependent kinases regulatory subunit 1 (Cks1) and p27kip1, and assess the prognostic significance of Skp2/Cks1 expression with p27kip1 in patients with extrahepatic cholangiocarcinoma. METHODS: Seventy-six patients who underwent curative resection for histologically confirmed extrahepatic cholangiocarcinoma at our institution from December 1994 to March 2008 were enrolled. Immunohistochemical staining for Skp2, Cks1, p27kip1, and Ki67, along with other relevant molecular biologic experiments, were performed. RESULTS: By Cox regression analyses, advanced age (> 65 years), advanced AJCC tumor stage, poorly differentiated histology, and higher immunostaining intensity of Skp2 were identified as independent prognostic factors in patients with extrahepatic cholangiocarcinoma. Exogenous epidermal growth factor (EGF, especially 0.1-10 ng/mL) significantly increased the proliferation indices by MTT assay and the mRNA levels of Skp2/Cks1 and p27kip1 in SNU-1196, SNU-1079, and SNU-245 cells. The protein levels of Skp2/Cks1 (from nuclear lysates) and p27kip1 (from cytosolic lysate) were also significantly increased in these cells. There were significant reductions in the protein levels of Skp2/Cks1 and p27kip1 (from nuclear lysate) after the treatment of LY294002. By chromatin immunoprecipitation assay, we found that E2F1 transcription factor directly binds to the promoter site of Skp2. CONCLUSION: Higher immunostaining intensity of Skp2/Cks1 was an independent prognostic factor for patients with extrahepatic cholangiocarcinoma. EGF upregulates the mRNA and protein levels of Skp2/Cks1 and p27kip1 via the PI3K/Akt pathway and direct binding of E2F1 transcription factor with the Skp2 promoter. PMID:24574749

  4. MARCKS Signaling Differentially Regulates Vascular Smooth Muscle and Endothelial Cell Proliferation through a KIS-, p27kip1- Dependent Mechanism

    PubMed Central

    Yu, Dan; Makkar, George; Dong, Tuo; Strickland, Dudley K.; Sarkar, Rajabrata; Monahan, Thomas Stacey

    2015-01-01

    Background Overexpression of the myristolated alanine-rich C kinase substrate (MARCKS) occurs in vascular proliferative diseases such as restenosis after bypass surgery. MARCKS knockdown results in arrest of vascular smooth muscle cell (VSMC) proliferation with little effect on endothelial cell (EC) proliferation. We sought to identify the mechanism of differential regulation by MARCKS of VSMC and EC proliferation in vitro and in vivo. Methods and Results siRNA-mediated MARCKS knockdown in VSMCs inhibited proliferation and prevented progression from phase G0/G1 to S. Protein expression of the cyclin-dependent kinase inhibitor p27kip1, but not p21cip1 was increased by MARCKS knockdown. MARCKS knockdown did not affect proliferation in VSMCs derived from p27kip1-/- mice indicating that the effect of MARCKS is p27kip1-dependent. MARCKS knockdown resulted in decreased phosphorylation of p27kip1 at threonine 187 and serine 10 as well as, kinase interacting with stathmin (KIS), cyclin D1, and Skp2 expression. Phosphorylation of p27kip1 at serine 10 by KIS is required for nuclear export and degradation of p27kip1. MARCKS knockdown caused nuclear trapping of p27kip1. Both p27kip1 nuclear trapping and cell cycle arrest were released by overexpression of KIS, but not catalytically inactive KIS. In ECs, MARCKS knockdown paradoxically increased KIS expression and cell proliferation. MARCKS knockdown in a murine aortic injury model resulted in decreased VSMC proliferation determined by bromodeoxyuridine (BrdU) integration assay, and inhibition of vascular wall thickening. MARCKS knockdown increased the rate of re-endothelialization. Conclusions MARCKS knockdown arrested VSMC cell cycle by decreasing KIS expression. Decreased KIS expression resulted in nuclear trapping of p27kip1 in VSMCs. MARCKS knockdown paradoxically increased KIS expression in ECs resulting in increased EC proliferation. MARCKS knockdown significantly attenuated the VSMC proliferative response to vascular

  5. Depletion of p18/LAMTOR1 promotes cell survival via activation of p27(kip1) -dependent autophagy under starvation.

    PubMed

    Zada, Sahib; Noh, Hae Sook; Baek, Seon Mi; Ha, Ji Hye; Hahm, Jong Ryeal; Kim, Deok Ryong

    2015-11-01

    The MAPK and mTOR signal pathways in endosomes or lysosomes play a crucial role in cell survival and death. They are also closely associated with autophagy, a catabolic process highly regulated under various cellular stress or nutrient deprivation. Recently we have isolated a protein, named p18/LAMTOR1, that specifically regulates the ERK or mTOR pathway in lysosomes. p18/LAMTOR1 also interacts with p27(kip1) . Here we examined how p18/LAMTOR1 plays a role in autophagy under nutrient deprivation. The p18(+/+) MEF cells were more susceptible to cell death under starvation or in the presence of AICAR in comparison with p18(-/-) MEF cells. Cleavage of caspase-3 was increased in p18(+/+) MEF cells under starvation, and phosphorylation at the threonine 198 of p27(kip1) was highly elevated in starved p18(-/-) MEF cells. Furthermore, LC3-II formation and other autophagy-associated proteins were largely increased in p18-deficient cells, and suppression of p27(kip1) expression in p18(-/-) MEF cells mitigated starvation-induced cell death. These data suggest that ablation of p18/LAMTOR1 suppresses starvation-induced cell death by stimulating autophagy through modulation of p27(kip1) activity. PMID:26032166

  6. CPEB1 restrains proliferation of Glioblastoma cells through the regulation of p27Kip1 mRNA translation

    PubMed Central

    Galardi, Silvia; Petretich, Massimo; Pinna, Guillaume; D’Amico, Silvia; Loreni, Fabrizio; Michienzi, Alessandro; Groisman, Irina; Ciafrè, Silvia Anna

    2016-01-01

    The cytoplasmic element binding protein 1 (CPEB1) regulates many important biological processes ranging from cell cycle control to learning and memory formation, by controlling mRNA translation efficiency via 3′ untranslated regions (3′UTR). In the present study, we show that CPEB1 is significantly downregulated in human Glioblastoma Multiforme (GBM) tissues and that the restoration of its expression impairs glioma cell lines growth. We demonstrate that CPEB1 promotes the expression of the cell cycle inhibitor p27Kip1 by specifically targeting its 3′UTR, and competes with miR-221/222 binding at an overlapping site in the 3′UTR, thus impairing miR-221/222 inhibitory activity. Upon binding to p27Kip1 3′UTR, CPEB1 promotes elongation of poly-A tail and the subsequent translation of p27Kip1 mRNA. This leads to higher levels of p27Kip1 in the cell, in turn significantly inhibiting cell proliferation, and confers to CPEB1 a potential value as a tumor suppressor in Glioblastoma. PMID:27142352

  7. CPEB1 restrains proliferation of Glioblastoma cells through the regulation of p27(Kip1) mRNA translation.

    PubMed

    Galardi, Silvia; Petretich, Massimo; Pinna, Guillaume; D'Amico, Silvia; Loreni, Fabrizio; Michienzi, Alessandro; Groisman, Irina; Ciafrè, Silvia Anna

    2016-01-01

    The cytoplasmic element binding protein 1 (CPEB1) regulates many important biological processes ranging from cell cycle control to learning and memory formation, by controlling mRNA translation efficiency via 3' untranslated regions (3'UTR). In the present study, we show that CPEB1 is significantly downregulated in human Glioblastoma Multiforme (GBM) tissues and that the restoration of its expression impairs glioma cell lines growth. We demonstrate that CPEB1 promotes the expression of the cell cycle inhibitor p27(Kip1) by specifically targeting its 3'UTR, and competes with miR-221/222 binding at an overlapping site in the 3'UTR, thus impairing miR-221/222 inhibitory activity. Upon binding to p27(Kip1) 3'UTR, CPEB1 promotes elongation of poly-A tail and the subsequent translation of p27(Kip1) mRNA. This leads to higher levels of p27(Kip1) in the cell, in turn significantly inhibiting cell proliferation, and confers to CPEB1 a potential value as a tumor suppressor in Glioblastoma. PMID:27142352

  8. Discovery of Small Molecules that Inhibit the Disordered Protein, p27Kip1

    DOE PAGESBeta

    Iconaru, Luigi I.; Ban, David; Bharatham, Kavitha; Ramanathan, Arvind; Zhang, Weixing; Shelat, Anang A.; Zuo, Jian; Kriwacki, Richard W.

    2015-10-28

    In disordered proteins we see that they are highly prevalent in biological systems. They control myriad signaling and regulatory processes, and their levels and/or cellular localization are often altered in human disease. In contrast to folded proteins, disordered proteins, due to conformational heterogeneity and dynamics, are not considered viable drug targets. We challenged this paradigm by identifying through NMR-based screening small molecules that bound specifically, albeit weakly, to the disordered cell cycle regulator, p27Kip1 (p27). Moreover, two groups of molecules bound to sites created by transient clusters of aromatic residues within p27. Conserved chemical features within these two groups ofmore » small molecules exhibited complementarity to their binding sites within p27, establishing structure-activity relationships for small molecule: disordered protein interactions. Finally, one compound counteracted the Cdk2/cyclin A inhibitory function of p27 in vitro, providing proof-of- principle that small molecules can inhibit the function of a disordered protein (p27) through sequestration in a conformation incapable of folding and binding to a natural regulatory target (Cdk2/cyclin A).« less

  9. Discovery of Small Molecules that Inhibit the Disordered Protein, p27Kip1

    PubMed Central

    Iconaru, Luigi I.; Ban, David; Bharatham, Kavitha; Ramanathan, Arvind; Zhang, Weixing; Shelat, Anang A.; Zuo, Jian; Kriwacki, Richard W.

    2015-01-01

    Disordered proteins are highly prevalent in biological systems, they control myriad signaling and regulatory processes, and their levels and/or cellular localization are often altered in human disease. In contrast to folded proteins, disordered proteins, due to conformational heterogeneity and dynamics, are not considered viable drug targets. We challenged this paradigm by identifying through NMR-based screening small molecules that bound specifically, albeit weakly, to the disordered cell cycle regulator, p27Kip1 (p27). Two groups of molecules bound to sites created by transient clusters of aromatic residues within p27. Conserved chemical features within these two groups of small molecules exhibited complementarity to their binding sites within p27, establishing structure-activity relationships for small molecule:disordered protein interactions. Finally, one compound counteracted the Cdk2/cyclin A inhibitory function of p27 in vitro, providing proof-of-principle that small molecules can inhibit the function of a disordered protein (p27) through sequestration in a conformation incapable of folding and binding to a natural regulatory target (Cdk2/cyclin A). PMID:26507530

  10. MYC antagonizes the differentiation induced by imatinib in chronic myeloid leukemia cells through downregulation of p27(KIP1.).

    PubMed

    Gómez-Casares, M T; García-Alegria, E; López-Jorge, C E; Ferrándiz, N; Blanco, R; Alvarez, S; Vaqué, J P; Bretones, G; Caraballo, J M; Sánchez-Bailón, P; Delgado, M D; Martín-Perez, J; Cigudosa, J C; León, J

    2013-04-25

    Chronic myeloid leukemia (CML) progresses from a chronic to a blastic phase where the leukemic cells are proliferative and undifferentiated. The CML is nowadays successfully treated with BCR-ABL kinase inhibitors as imatinib and dasatinib. In the CML-derived K562 cell line, low concentrations of imatinib induce proliferative arrest and erythroid differentiation. We found that imatinib upregulated the cell cycle inhibitor p27(KIP1) (p27) in a time- and -concentration dependent manner, and that the extent of imatinib-mediated differentiation was severely decreased in cells with depleted p27. MYC (c-Myc) is a transcription factor frequently deregulated in human cancer. MYC is overexpressed in untreated CML and is associated to poor response to imatinib. Using K562 sublines with conditional MYC expression (induced by Zn(2+) or activated by 4-hydroxy-tamoxifen) we show that MYC prevented the erythroid differentiation induced by imatinib and dasatinib. The differentiation inhibition is not due to increased proliferation of MYC-expressing clones or enhanced apoptosis of differentiated cells. As p27 overexpression is reported to induce erythroid differentiation in K562, we explored the effect of MYC on imatinib-dependent induction of p27. We show that MYC abrogated the imatinib-induced upregulation of p27 concomitantly with the differentiation inhibition, suggesting that MYC inhibits differentiation by antagonizing the imatinib-mediated upregulation of p27. This effect occurs mainly by p27 protein destabilization. This was in part due to MYC-dependent induction of SKP2, a component of the ubiquitin ligase complex that targets p27 for degradation. The results suggest that, although MYC deregulation does not directly confer resistance to imatinib, it might be a factor that contributes to progression of CML through the inhibition of differentiation. PMID:22710719

  11. Cryptic sequence features within the disordered protein p27Kip1 regulate cell cycle signaling

    PubMed Central

    Das, Rahul K.; Huang, Yongqi; Phillips, Aaron H.; Kriwacki, Richard W.; Pappu, Rohit V.

    2016-01-01

    Peptide motifs embedded within intrinsically disordered regions (IDRs) of proteins are often the sites of posttranslational modifications that control cell-signaling pathways. How do IDR sequences modulate the functionalities of motifs? We answer this question using the polyampholytic C-terminal IDR of the cell cycle inhibitory protein p27Kip1 (p27). Phosphorylation of Thr-187 (T187) within the p27 IDR controls entry into S phase of the cell division cycle. Additionally, the conformational properties of polyampholytic sequences are predicted to be influenced by the linear patterning of oppositely charged residues. Therefore, we designed sequence variants of the p27 IDR to alter charge patterning outside the primary substrate motif containing T187. Computer simulations and biophysical measurements confirm predictions regarding the impact of charge patterning on the global dimensions of IDRs. Through functional studies, we uncover cryptic sequence features within the p27 IDR that influence the efficiency of T187 phosphorylation. Specifically, we find a positive correlation between T187 phosphorylation efficiency and the weighted net charge per residue of an auxiliary motif. We also find that accumulation of positive charges within the auxiliary motif can diminish the efficiency of T187 phosphorylation because this increases the likelihood of long-range intra-IDR interactions that involve both the primary and auxiliary motifs and inhibit their contributions to function. Importantly, our findings suggest that the cryptic sequence features of the WT p27 IDR negatively regulate T187 phosphorylation signaling. Our approaches provide a generalizable strategy for uncovering the influence of sequence contexts on the functionalities of primary motifs in other IDRs. PMID:27140628

  12. Cryptic sequence features within the disordered protein p27Kip1 regulate cell cycle signaling.

    PubMed

    Das, Rahul K; Huang, Yongqi; Phillips, Aaron H; Kriwacki, Richard W; Pappu, Rohit V

    2016-05-17

    Peptide motifs embedded within intrinsically disordered regions (IDRs) of proteins are often the sites of posttranslational modifications that control cell-signaling pathways. How do IDR sequences modulate the functionalities of motifs? We answer this question using the polyampholytic C-terminal IDR of the cell cycle inhibitory protein p27(Kip1) (p27). Phosphorylation of Thr-187 (T187) within the p27 IDR controls entry into S phase of the cell division cycle. Additionally, the conformational properties of polyampholytic sequences are predicted to be influenced by the linear patterning of oppositely charged residues. Therefore, we designed sequence variants of the p27 IDR to alter charge patterning outside the primary substrate motif containing T187. Computer simulations and biophysical measurements confirm predictions regarding the impact of charge patterning on the global dimensions of IDRs. Through functional studies, we uncover cryptic sequence features within the p27 IDR that influence the efficiency of T187 phosphorylation. Specifically, we find a positive correlation between T187 phosphorylation efficiency and the weighted net charge per residue of an auxiliary motif. We also find that accumulation of positive charges within the auxiliary motif can diminish the efficiency of T187 phosphorylation because this increases the likelihood of long-range intra-IDR interactions that involve both the primary and auxiliary motifs and inhibit their contributions to function. Importantly, our findings suggest that the cryptic sequence features of the WT p27 IDR negatively regulate T187 phosphorylation signaling. Our approaches provide a generalizable strategy for uncovering the influence of sequence contexts on the functionalities of primary motifs in other IDRs. PMID:27140628

  13. Notch Signaling Activates Stem Cell Properties of Müller Glia through Transcriptional Regulation and Skp2-mediated Degradation of p27Kip1

    PubMed Central

    Parameswaran, Sowmya; Mathews, Saumi; Xia, Xiaohuan; Zheng, Li; Neville, Andrew J.; Ahmad, Iqbal

    2016-01-01

    Müller glia (MG), the sole glial cells generated by retinal progenitors, have emerged as a viable cellular target for therapeutic regeneration in degenerative blinding diseases, as they possess dormant stem cell properties. However, the mammalian MG does not display the neurogenic potential of their lower vertebrate counterparts, precluding their practical clinical use. The answer to this barrier may be found in two interlinked processes underlying the neurogenic potential, i.e., the activation of the dormant stem cell properties of MG and their differentiation along the neuronal lineage. Here, we have focused on the former and examined Notch signaling-mediated activation of MG. We demonstrate that one of the targets of Notch signaling is the cyclin-dependent kinase inhibitor (CKI), p27Kip1, which is highly expressed in quiescent MG. Notch signaling facilitates the activation of MG by inhibiting p27Kip1 expression. This is likely achieved through the Notch- p27Kip1 and Notch-Skp2-p27Kip1 axes, the former inhibiting the expression of p27Kip1 transcripts and the latter levels of p27Kip1 proteins by Skp2-mediated proteasomal degradation. Thus, Notch signaling may facilitate re-entry of MG into the cell cycle by inhibiting p27Kip1 expression both transcriptionally and post-translationally. PMID:27011052

  14. Notch Signaling Activates Stem Cell Properties of Müller Glia through Transcriptional Regulation and Skp2-mediated Degradation of p27Kip1.

    PubMed

    Del Debbio, Carolina Beltrame; Mir, Qulsum; Parameswaran, Sowmya; Mathews, Saumi; Xia, Xiaohuan; Zheng, Li; Neville, Andrew J; Ahmad, Iqbal

    2016-01-01

    Müller glia (MG), the sole glial cells generated by retinal progenitors, have emerged as a viable cellular target for therapeutic regeneration in degenerative blinding diseases, as they possess dormant stem cell properties. However, the mammalian MG does not display the neurogenic potential of their lower vertebrate counterparts, precluding their practical clinical use. The answer to this barrier may be found in two interlinked processes underlying the neurogenic potential, i.e., the activation of the dormant stem cell properties of MG and their differentiation along the neuronal lineage. Here, we have focused on the former and examined Notch signaling-mediated activation of MG. We demonstrate that one of the targets of Notch signaling is the cyclin-dependent kinase inhibitor (CKI), p27Kip1, which is highly expressed in quiescent MG. Notch signaling facilitates the activation of MG by inhibiting p27Kip1 expression. This is likely achieved through the Notch- p27Kip1 and Notch-Skp2-p27Kip1 axes, the former inhibiting the expression of p27Kip1 transcripts and the latter levels of p27Kip1 proteins by Skp2-mediated proteasomal degradation. Thus, Notch signaling may facilitate re-entry of MG into the cell cycle by inhibiting p27Kip1 expression both transcriptionally and post-translationally. PMID:27011052

  15. A single-nucleotide polymorphism in the human p27kip1 gene (-838C>A) affects basal promoter activity and the risk of myocardial infarction

    PubMed Central

    González, Pelayo; Díez-Juan, Antonio; Coto, Eliecer; Álvarez, Victoria; Reguero, Julian R; Batalla, Alberto; Andrés, Vicente

    2004-01-01

    Background Excessive proliferation of vascular smooth muscle cells and leukocytes within the artery wall is a major event in the development of atherosclerosis. The growth suppressor p27kip1 associates with several cyclin-dependent kinase/cyclin complexes, thereby abrogating their capacity to induce progression through the cell cycle. Recent studies have implicated p27kip1 in the control of neointimal hyperplasia. For instance, p27kip1 ablation in apolipoprotein-E-null mice enhanced arterial cell proliferation and accelerated atherogenesis induced by dietary cholesterol. Therefore, p27kip1 is a candidate gene to modify the risk of developing atherosclerosis and associated ischaemic events (i.e., myocardial infarction and stroke). Results In this study we found three common single-nucleotide polymorphisms in the human p27kip1 gene (+326T>G [V109G], -79C>T, and -838C>A). The frequency of -838A carriers was significantly increased in myocardial infarction patients compared to healthy controls (odds ratio [OR] = 1.73, 95% confidence interval [95%CI] = 1.12–2.70). In addition, luciferase reporter constructs driven by the human p27kip1 gene promoter containing A at position -838 had decreased basal transcriptional activity when transiently transfected in Jurkat cells, compared with constructs bearing C in -838 (P = 0.04). Conclusions These data suggest that -838A is associated with reduced p27kip1 promoter activity and increased risk of myocardial infarction. PMID:15061869

  16. Differential roles of p21(Waf1) and p27(Kip1) in modulating chemosensitivity and their possible application in drug discovery studies.

    PubMed

    Schmidt, M; Lu, Y; Parant, J M; Lozano, G; Bacher, G; Beckers, T; Fan, Z

    2001-11-01

    In this study, the differential role of the cyclin-dependent kinase (CDK) inhibitors p21(Waf1) and p27(Kip1) in cell cycle regulation was proposed for use in screening natural or synthetic compounds for cell cycle-dependent (particularly M phase-dependent) antineoplastic activity. p21(Waf1) or p27(Kip1) was ectopically expressed with an ecdysone-inducible mammalian expression system in a human colon adenocarcinoma cell line. Induction of p21(Waf1) or p27(Kip1) expression inhibited the activities of CDK2 and completely arrested cells at G(1) phase of the cell cycle by p27(Kip1) and at G(1) and G(2) phases by p21(Waf1). We examined the sensitivity of these cells to several antineoplastic agents known to be cell cycle-dependent or -independent. Substantially increased resistance to cell cycle-dependent antineoplastic agents was found in the cells when the expression of p21(Waf1) or p27(Kip1) was induced. In contrast, only a desensitization to cell cycle-independent antineoplastic agents was found in the cells arrested by p21(Waf1) or p27(Kip1). Because p21(Waf1) induces an additional block at G(2) phase that inhibits cell entry into M phase, we further examined the difference between p21(Waf1)- and p27(Kip1)-induced cells in their sensitivity to D-24851, a novel M phase-dependent compound. We found that induction of p21(Waf1) after exposure of the cells to D-24851 conferred stronger resistance than did induction of p27(Kip1). Taken together, our results suggest that the differential effect of p21(Waf1) and p27(Kip1) on cell cycle regulation may be advantageous for screening chemical libraries for novel antineoplastic candidates that are cell cycle-dependent, and M phase-dependent in particular. PMID:11641417

  17. Rat Protein Tyrosine Phosphatase η Suppresses the Neoplastic Phenotype of Retrovirally Transformed Thyroid Cells through the Stabilization of p27Kip1

    PubMed Central

    Trapasso, Francesco; Iuliano, Rodolfo; Boccia, Angelo; Stella, Antonella; Visconti, Roberta; Bruni, Paola; Baldassarre, Gustavo; Santoro, Massimo; Viglietto, Giuseppe; Fusco, Alfredo

    2000-01-01

    The r-PTPη gene encodes a rat receptor-type protein tyrosine phosphatase whose expression is negatively regulated by neoplastic cell transformation. Here we first demonstrate a dramatic reduction in DEP-1/HPTPη (the human homolog of r-PTPη) expression in a panel of human thyroid carcinomas. Subsequently, we show that the reexpression of the r-PTPη gene in highly malignant rat thyroid cells transformed by retroviruses carrying the v-mos and v-ras-Ki oncogenes suppresses their malignant phenotype. Cell cycle analysis demonstrated that r-PTPη caused G1 growth arrest and increased the cyclin-dependent kinase inhibitor p27Kip1 protein level by reducing the proteasome-dependent degradation rate. We propose that the r-PTPη tumor suppressor activity is mediated by p27Kip1 protein stabilization, because suppression of p27Kip1 protein synthesis using p27-specific antisense oligonucleotides blocked the growth-inhibitory effect induced by r-PTPη. Furthermore, we provide evidence that in v-mos- or v-ras-Ki-transformed thyroid cells, the p27Kip1 protein level was regulated by the mitogen-activated protein (MAP) kinase pathway and that r-PTPη regulated p27Kip1 stability by preventing v-mos- or v-ras-Ki-induced MAP kinase activation. PMID:11094075

  18. Brk/Protein tyrosine kinase 6 phosphorylates p27KIP1, regulating the activity of cyclin D-cyclin-dependent kinase 4.

    PubMed

    Patel, Priyank; Asbach, Benedikt; Shteyn, Elina; Gomez, Cindy; Coltoff, Alexander; Bhuyan, Sadia; Tyner, Angela L; Wagner, Ralf; Blain, Stacy W

    2015-05-01

    Cyclin D and cyclin-dependent kinase 4 (cdk4) are overexpressed in a variety of tumors, but their levels are not accurate indicators of oncogenic activity because an accessory factor such as p27(Kip1) is required to assemble this unstable dimer. Additionally, tyrosine (Y) phosphorylation of p27 (pY88) is required to activate cdk4, acting as an "on/off switch." We identified two SH3 recruitment domains within p27 that modulate pY88, thereby modulating cdk4 activity. Via an SH3-PXXP interaction screen, we identified Brk (breast tumor-related kinase) as a high-affinity p27 kinase. Modulation of Brk in breast cancer cells modulates pY88 and increases resistance to the cdk4 inhibitor PD 0332991. An alternatively spliced form of Brk (Alt Brk) which contains its SH3 domain blocks pY88 and acts as an endogenous cdk4 inhibitor, identifying a potentially targetable regulatory region within p27. Brk is overexpressed in 60% of breast carcinomas, suggesting that this facilitates cell cycle progression by modulating cdk4 through p27 Y phosphorylation. p27 has been considered a tumor suppressor, but our data strengthen the idea that it should also be considered an oncoprotein, responsible for cyclin D-cdk4 activity. PMID:25733683

  19. Knockdown of AMPKα2 Promotes Pulmonary Arterial Smooth Muscle Cells Proliferation via mTOR/Skp2/p27Kip1 Signaling Pathway

    PubMed Central

    Ke, Rui; Liu, Lu; Zhu, Yanting; Li, Shaojun; Xie, Xinming; Li, Fangwei; Song, Yang; Yang, Lan; Gao, Li; Li, Manxiang

    2016-01-01

    It has been shown that activation of adenosine monophosphate-activated protein kinase (AMPK) suppresses proliferation of a variety of tumor cells as well as nonmalignant cells. In this study, we used post-transcriptional gene silencing with small interfering RNA (siRNA) to specifically examine the effect of AMPK on pulmonary arterial smooth muscle cells (PASMCs) proliferation and to further elucidate its underlying molecular mechanisms. Our results showed that knockdown of AMPKα2 promoted primary cultured PASMCs proliferation; this was accompanied with the elevation of phosphorylation of mammalian target of rapamycin (mTOR) and S-phase kinase-associated protein 2 (Skp2) protein level and reduction of p27Kip1. Importantly, prior silencing of mTOR with siRNA abolished AMPKα2 knockdown-induced Skp2 upregulation, p27Kip1 reduction as well as PASMCs proliferation. Furthermore, pre-depletion of Skp2 by siRNA also eliminated p27Kip1 downregulation and PASMCs proliferation caused by AMPKα2 knockdown. Taken together, our study indicates that AMPKα2 isoform plays an important role in regulation of PASMCs proliferation by modulating mTOR/Skp2/p27Kip1 axis, and suggests that activation of AMPKα2 might have potential value in the prevention and treatment of pulmonary arterial hypertension. PMID:27258250

  20. Butein inhibits cell proliferation and induces cell cycle arrest in acute lymphoblastic leukemia via FOXO3a/p27kip1 pathway

    PubMed Central

    Wang, Li-Na; Tian, Yun; Shi, Dingbo; Wang, Jingshu; Qin, Ge; Li, Anchuan; Liang, Yan-Ni; Zhou, Huan-Juan; Ke, Zhi-Yong; Huang, Wenlin; Deng, Wuguo; Luo, Xue-Qun

    2016-01-01

    Acute lymphoblastic leukemia (ALL) is a common hematological malignancy characterized by the uncontrolled proliferation of leukemia cells in children. Discovering and developing effective chemotherapeutic drugs are needed for ALL. In this study, we investigated the anti-leukemic activity of butein and its action mechanisms in ALL. Butein was found to significantly suppress the cellular proliferation of ALL cell lines and primary ALL blasts in a dose-dependent manner. It also induced cell cycle arrest by decreasing the expression of cyclin E and CDK2. We also found that butein promoted nuclear Forkhead Class box O3a (FOXO3a) localization, enhanced the binding of FOXO3a on the p27kip1 gene promoter and then increased the expression of p27kip1. Moreover, we showed that FOXO3a knockdown significantly decreased the proliferation inhibition by butein, whereas overexpression of FOXO3a enhanced the butein-mediated proliferation inhibition. However, overexpression of FOXO3a mutation (C-terminally truncated FOXO3a DNA-binding domain) decreased the proliferation inhibition by butein through decreasing the expression of p27kip1. Our results therefore demonstrate the therapeutic potential of butein for ALL via FOXO3a/p27kip1 pathway. PMID:26919107

  1. HBx-upregulated lncRNA UCA1 promotes cell growth and tumorigenesis by recruiting EZH2 and repressing p27Kip1/CDK2 signaling

    PubMed Central

    Hu, Jiao-Jiao; Song, Wei; Zhang, Shao-Dan; Shen, Xiao-Hui; Qiu, Xue-Mei; Wu, Hua-Zhang; Gong, Pi-Hai; Lu, Sen; Zhao, Zhu-Jiang; He, Ming-Liang; Fan, Hong

    2016-01-01

    It is well accepted that HBx plays the major role in hepatocarcinogenesis associated with hepatitis B virus (HBV) infections. However, little was known about its role in regulating long noncoding RNAs (lncRNAs), a large group of transcripts regulating a variety of biological processes including carcinogenesis in mammalian cells. Here we report that HBx upregulates UCA1 genes and downregulates p27 genes in hepatic LO2 cells. Further studies show that the upregulated UCA1 promotes cell growth by facilitating G1/S transition through CDK2 in both hepatic and hepatoma cells. Knock down of UCA1 in HBx-expressing hepatic and hepatoma cells resulted in markedly increased apoptotic cells by elevating the cleaved caspase-3 and caspase-8. More importantly, UCA1 is found to be physically associated with enhancer of zeste homolog 2 (EZH2), which suppresses p27Kip1 through histone methylation (H3K27me3) on p27Kip1 promoter. We also show that knockdown of UCA1 in hepatoma cells inhibits tumorigenesis in nude mice. In a clinic study, UCA1 is found to be frequently up-regulated in HBx positive group tissues in comparison with the HBx negative group, and exhibits an inverse correlation between UCA1 and p27Kip1 levels. Our findings demonstrate an important mechanism of hepatocarcinogenesis through the signaling of HBx-UCA1/EZH2-p27Kip1 axis, and a potential target of HCC. PMID:27009634

  2. Involvement of p27KIP1 in G1 arrest mediated by an anti-epidermal growth factor receptor monoclonal antibody.

    PubMed

    Wu, X; Rubin, M; Fan, Z; DeBlasio, T; Soos, T; Koff, A; Mendelsohn, J

    1996-04-01

    Activation of the cyclin dependent kinases (CDK4/CDK6 and CDK2) is required for G1 phase progression and entry into S-phase. The activation of these kinases is regulated by checkpoints that monitor environmental and intracellular conditions. Progression into S-phase is controlled, in part, by the availability of growth factors, and we have investigated the relationship between growth factor availability and the activation of the CDK kinases. Blocking activation of epidermal growth factor (EGF) receptor tyrosine kinase with anti-EGF receptor monoclonal antibody (mAb) 225 induces G1 phase cell cycle arrest in DiFi human colon adenocarcinoma cells. When DiFi cells are treated with mAb 225 for 24 h, we observe marked decreases in the activities of CDK2 kinase and cyclin E-associated CDK kinase which are not accompanied by reduced levels of cyclin E and CDK2 proteins. However, the amount of cyclin/CDK kinase inhibitor p27KIP1 increases in the mAb-treated cells and p27KIP1 is bound to CDK2 in increasing amounts. Immunodepletion of p27KIP1 removes an inhibitory activity from lysates of mAb-treated cells: the immunodepleted and heated lysates lose the capacity to inhibit cyclin E/CDK2 activity in an in vitro assay. The results suggest that G1 arrest in the cell cycle induced by EGF receptor blockade involves p27KIP1. PMID:8622855

  3. HBx-upregulated lncRNA UCA1 promotes cell growth and tumorigenesis by recruiting EZH2 and repressing p27Kip1/CDK2 signaling.

    PubMed

    Hu, Jiao-Jiao; Song, Wei; Zhang, Shao-Dan; Shen, Xiao-Hui; Qiu, Xue-Mei; Wu, Hua-Zhang; Gong, Pi-Hai; Lu, Sen; Zhao, Zhu-Jiang; He, Ming-Liang; Fan, Hong

    2016-01-01

    It is well accepted that HBx plays the major role in hepatocarcinogenesis associated with hepatitis B virus (HBV) infections. However, little was known about its role in regulating long noncoding RNAs (lncRNAs), a large group of transcripts regulating a variety of biological processes including carcinogenesis in mammalian cells. Here we report that HBx upregulates UCA1 genes and downregulates p27 genes in hepatic LO2 cells. Further studies show that the upregulated UCA1 promotes cell growth by facilitating G1/S transition through CDK2 in both hepatic and hepatoma cells. Knock down of UCA1 in HBx-expressing hepatic and hepatoma cells resulted in markedly increased apoptotic cells by elevating the cleaved caspase-3 and caspase-8. More importantly, UCA1 is found to be physically associated with enhancer of zeste homolog 2 (EZH2), which suppresses p27Kip1 through histone methylation (H3K27me3) on p27Kip1 promoter. We also show that knockdown of UCA1 in hepatoma cells inhibits tumorigenesis in nude mice. In a clinic study, UCA1 is found to be frequently up-regulated in HBx positive group tissues in comparison with the HBx negative group, and exhibits an inverse correlation between UCA1 and p27Kip1 levels. Our findings demonstrate an important mechanism of hepatocarcinogenesis through the signaling of HBx-UCA1/EZH2-p27Kip1 axis, and a potential target of HCC. PMID:27009634

  4. Prognostic implications of expression of the cell cycle inhibitor protein p27Kip1.

    PubMed

    Cariou, S; Catzavelos, C; Slingerland, J M

    1998-01-01

    Mitogenic and growth inhibitory signals influence the activity of a family of cyclin dependent kinases (cdks). p27 is an important cdk inhibitor, acting in G1 to inhibit cyclin-cdks. As negative growth regulators, the cdk inhibitors may function as tumor suppressors. While the p16 gene plays a tumor suppressor role in cancers, p27 gene mutations have been identified only rarely. While high levels of p27 protein are expressed in normal human mammary epithelium, loss of p27 is frequent and is of independent prognostic significance in breast cancers. Low p27 is also a poor prognostic factor in colon, gastric, esophageal, lung, and prostate carcinomas, and enhanced proteasomal degradation may underlie loss of p27 in tumor cells. Loss of p27 has not been significantly correlated with tumor proliferation in a number of studies and may reflect alterations in differentiation and adhesion-dependent growth regulation germane to oncogenesis and tumor progression. Efforts to confirm the prognostic value of p27 are under way in a number of large breast cancer studies. These studies may also indicate whether loss of p27 in association with other traditional or novel markers has greater prognostic potential than each factor alone. p27 immunostaining is inexpensive and reliable and may become part of the routine histopathologic processing of tumors in the near future. Widespread application of p27 in prognostic testing will require greater uniformity in scoring techniques and determination of the cut off levels which distinguish individuals at high and low risk of cancer recurrence and death. Finally, the greatest utility of p27 may lie in the information it sheds on the biology of aberrant growth regulation in breast cancer and the potential to use this in the generation of novel therapeutic strategies. PMID:10066070

  5. Embryonic Lethal Abnormal Vision-like HuR-dependent mRNA Stability Regulates Post-transcriptional Expression of Cyclin-dependent Kinase Inhibitor p27Kip1

    PubMed Central

    Ziegeler, Gudrun; Ming, Jie; Koseki, Jana C.; Sevinc, Sema; Chen, Ting; Ergun, Suleyman; Qin, Xuebin; Aktas, Bertal H.

    2010-01-01

    The cyclin-dependent kinase inhibitor p27Kip1 plays a critical role in regulating entry into and exit from the cell cycle. Post-transcriptional regulation of p27Kip1 expression is of significant interest. The embryonic lethal abnormal vision (ELAV)-like RNA-binding protein HuR is thought be important for the translation of p27Kip1, however, different reports attributed diametrically opposite roles to HuR. We report here an alternative mechanism wherein HuR regulates stability of the p27Kip1 mRNA. Specifically, human and mouse p27Kip1 mRNAs interact with HuR protein through multiple U-rich elements in both 5′ and 3′ untranslated regions (UTR). These interactions, which occur in vitro and in vivo, stabilize p27Kip1 mRNA and play a critical role in its accumulation. Deleting HuR binding sites or knocking down HuR expression destabilizes p27Kip1 mRNA and reduces its accumulation. We also identified a CT repeat in the 5′ UTR of full-length p27Kip1 mRNA isoforms that interact with a ∼41-kDa protein and represses p27Kip1 expression. This CT-rich element and diffuse elements in the 3′ UTR regulate post-transcriptional expression of p27Kip1 at the level of translation. This is the first demonstration that HuR-dependent mRNA stability and HuR-independent mRNA translation plays a critical role in the regulation of post-transcriptional p27Kip1 expression. PMID:20332085

  6. NSun2 delays replicative senescence by repressing p27 (KIP1) translation and elevating CDK1 translation

    PubMed Central

    Xing, Junyue; Liu, Zhenyun; Jiang, Bin; Dou, Yali; Gorospe, Myriam; Wang, Wengong

    2015-01-01

    A rise in the levels of the cyclin-dependent kinase (CDK) inhibitor p27KIP1 is important for the growth arrest of senescent cells, but the mechanisms responsible for this increase are poorly understood. Here, we show that the tRNA methyltransferase NSun2 represses the expression of p27 in replicative senescence. NSun2 methylated the 5′-untranslated region (UTR) of p27 mRNA at cytosine C64 in vitro and in cells, thereby repressing the translation of p27. During replicative senescence, increased p27 protein levels were accompanied by decreased NSun2 protein levels. Knockdown of NSun2 in human diploid fibroblasts (HDFs) elevated p27 levels and reduced the expression of CDK1 (encoded by CDK1 mRNA, a previously reported target of NSun2), which in turn further repressed cell proliferation and accelerated replicative senescence, while overexpression of NSun2 exerted the opposite effect. Ectopic overexpression of the p27 5′UTR fragment rescued the effect of NSun2 overexpression in lowering p27, increasing CDK1, promoting cell proliferation, and delaying replicative senescence. Our findings indicate that NSun2-mediated mRNA methylation regulates p27 and CDK1 levels during replicative senescence. PMID:26687548

  7. TGF-β activates APC through Cdh1 binding for Cks1 and Skp2 proteasomal destruction stabilizing p27kip1 for normal endometrial growth.

    PubMed

    Pavlides, Savvas C; Lecanda, Jon; Daubriac, Julien; Pandya, Unnati M; Gama, Patricia; Blank, Stephanie; Mittal, Khushbakhat; Shukla, Pratibha; Gold, Leslie I

    2016-04-01

    We previously reported that aberrant TGF-β/Smad2/3 signaling in endometrial cancer (ECA) leads to continuous ubiquitylation of p27(kip1)(p27) by the E3 ligase SCF-Skp2/Cks1 causing its degradation, as a putative mechanism involved in the pathogenesis of this cancer. In contrast, normal intact TGF-β signaling prevents degradation of nuclear p27 by SCF-Skp2/Cks1 thereby accumulating p27 to block Cdk2 for growth arrest. Here we show that in ECA cell lines and normal primary endometrial epithelial cells, TGF-β increases Cdh1 and its binding to APC/C to form the E3 ligase complex that ubiquitylates Cks1 and Skp2 prompting their proteasomal degradation and thus, leaving p27 intact. Knocking-down Cdh1 in ECA cell lines increased Skp2/Cks1 E3 ligase activity, completely diminished nuclear and cytoplasmic p27, and obviated TGF-β-mediated inhibition of proliferation. Protein synthesis was not required for TGF-β-induced increase in nuclear p27 and decrease in Cks1 and Skp2. Moreover, half-lives of Cks1 and Skp2 were extended in the Cdh1-depleted cells. These results suggest that the levels of p27, Skp2 and Cks1 are strongly or solely regulated by proteasomal degradation. Finally, an inverse relationship of low p27 and high Cks1 in the nucleus was shown in patients in normal proliferative endometrium and grade I-III ECAs whereas differentiated secretory endometrium showed the reverse. These studies implicate Cdh1 as the master regulator of TGF-β-induced preservation of p27 tumor suppressor activity. Thus, Cdh1 is a potential therapeutic target for ECA and other human cancers showing an inverse relationship between Cks1/Skp2 and p27 and/or dysregulated TGF-β signaling. PMID:26963853

  8. SKP2 Oncogene Is a Direct MYC Target Gene and MYC Down-regulates p27KIP1 through SKP2 in Human Leukemia Cells*

    PubMed Central

    Bretones, Gabriel; Acosta, Juan C.; Caraballo, Juan M.; Ferrándiz, Nuria; Gómez-Casares, M. Teresa; Albajar, Marta; Blanco, Rosa; Ruiz, Paula; Hung, Wen-Chun; Albero, M. Pilar; Perez-Roger, Ignacio; León, Javier

    2011-01-01

    SKP2 is the ubiquitin ligase subunit that targets p27KIP1 (p27) for degradation. SKP2 is induced in the G1-S transit of the cell cycle, is frequently overexpressed in human cancer, and displays transformation activity in experimental models. Here we show that MYC induces SKP2 expression at the mRNA and protein levels in human myeloid leukemia K562 cells with conditional MYC expression. Importantly, in these systems, induction of MYC did not activate cell proliferation, ruling out SKP2 up-regulation as a consequence of cell cycle entry. MYC-dependent SKP2 expression was also detected in other cell types such as lymphoid, fibroblastic, and epithelial cell lines. MYC induced SKP2 mRNA expression in the absence of protein synthesis and activated the SKP2 promoter in luciferase reporter assays. With chromatin immunoprecipitation assays, MYC was detected bound to a region of human SKP2 gene promoter that includes E-boxes. The K562 cell line derives from human chronic myeloid leukemia. In a cohort of chronic myeloid leukemia bone marrow samples, we found a correlation between MYC and SKP2 mRNA levels. Analysis of cancer expression databases also indicated a correlation between MYC and SKP2 expression in lymphoma. Finally, MYC-induced SKP2 expression resulted in a decrease in p27 protein in K562 cells. Moreover, silencing of SKP2 abrogated the MYC-mediated down-regulation of p27. Our data show that SKP2 is a direct MYC target gene and that MYC-mediated SKP2 induction leads to reduced p27 levels. The results suggest the induction of SKP2 oncogene as a new mechanism for MYC-dependent transformation. PMID:21245140

  9. SKP2 oncogene is a direct MYC target gene and MYC down-regulates p27(KIP1) through SKP2 in human leukemia cells.

    PubMed

    Bretones, Gabriel; Acosta, Juan C; Caraballo, Juan M; Ferrándiz, Nuria; Gómez-Casares, M Teresa; Albajar, Marta; Blanco, Rosa; Ruiz, Paula; Hung, Wen-Chun; Albero, M Pilar; Perez-Roger, Ignacio; León, Javier

    2011-03-18

    SKP2 is the ubiquitin ligase subunit that targets p27(KIP1) (p27) for degradation. SKP2 is induced in the G(1)-S transit of the cell cycle, is frequently overexpressed in human cancer, and displays transformation activity in experimental models. Here we show that MYC induces SKP2 expression at the mRNA and protein levels in human myeloid leukemia K562 cells with conditional MYC expression. Importantly, in these systems, induction of MYC did not activate cell proliferation, ruling out SKP2 up-regulation as a consequence of cell cycle entry. MYC-dependent SKP2 expression was also detected in other cell types such as lymphoid, fibroblastic, and epithelial cell lines. MYC induced SKP2 mRNA expression in the absence of protein synthesis and activated the SKP2 promoter in luciferase reporter assays. With chromatin immunoprecipitation assays, MYC was detected bound to a region of human SKP2 gene promoter that includes E-boxes. The K562 cell line derives from human chronic myeloid leukemia. In a cohort of chronic myeloid leukemia bone marrow samples, we found a correlation between MYC and SKP2 mRNA levels. Analysis of cancer expression databases also indicated a correlation between MYC and SKP2 expression in lymphoma. Finally, MYC-induced SKP2 expression resulted in a decrease in p27 protein in K562 cells. Moreover, silencing of SKP2 abrogated the MYC-mediated down-regulation of p27. Our data show that SKP2 is a direct MYC target gene and that MYC-mediated SKP2 induction leads to reduced p27 levels. The results suggest the induction of SKP2 oncogene as a new mechanism for MYC-dependent transformation. PMID:21245140

  10. p27kip1 Maintains a Subset of Leukemia Stem Cells in the Quiescent State in Murine MLL-Leukemia

    PubMed Central

    Zhang, Jun; Seet, Christopher; Sun, Clare; Li, Jing; You, Dewen; Volk, Andrew; Breslin, Peter; Li, Xingyu; Wei, Wei; Qian, Zhijian; Zeleznik-Le, Nancy J.; Zhang, Zhou; Zhang, Jiwang

    2013-01-01

    MLL (mixed-lineage leukemia)-fusion genes induce the development of leukemia through deregulation of normal MLL target genes, such as HOXA9 and MEIS1. Both HOXA9 and MEIS1 are required for MLL-fusion gene-induced leukemogenesis. Co-expression of HOXA9 and MEIS1 induces acute myeloid leukemia (AML) similar to that seen in mice in which MLL-fusion genes are over-expressed. p27kip1 (p27 hereafter), a negative regulator of the cell cycle, has also been defined as an MLL target, the expression of which is up-regulated in MLL leukemic cells (LCs). To investigate whether p27 plays a role in the pathogenesis of MLL-leukemia, we examined the effects of p27 deletion (p27-/-) on MLL-AF9 (MA9)-induced murine AML development. HOXA9/MEIS1 (H/M)-induced, p27 wild-type (p27+/+) and p27-/- AML were studied in parallel as controls. We found that LCs from both MA9-AML and H/M-AML can be separated into three fractions, a CD117-CD11bhi differentiated fraction as well as CD117+CD11bhi and CD117+CD11blo, two less differentiated fractions. The CD117+CD11blo fraction, comprising only 1-3% of total LCs, expresses higher levels of early hematopoietic progenitor markers but lower levels of mature myeloid cell markers compared to other populations of LCs. p27 is expressed and is required for maintaining the quiescent and drug-resistant states of the CD117+CD11blo fraction of MA9-LCs but not of H/M-LCs. p27 deletion significantly compromises the leukemogenic capacity of CD117+CD11blo MA9-LCs by reducing the frequency of leukemic stem cells (LSCs) but does not do so in H/M-LCs. In addition, we found that p27 is highly expressed and required for cell cycle arrest in the CD117-CD11bhi fraction in both types of LCs. Furthermore, we found that c-Myc expression is required for maintaining LCs in an undifferentiated state independently of proliferation. We concluded that p27 represses the proliferation of LCs, which is specifically required for maintaining the quiescent and drug-resistant states of a

  11. A vitamin D3 analog induces a G1-phase arrest in CaCo-2 cells by inhibiting cdk2 and cdk6: roles of cyclin E, p21Waf1, and p27Kip1.

    PubMed

    Scaglione-Sewell, B A; Bissonnette, M; Skarosi, S; Abraham, C; Brasitus, T A

    2000-11-01

    Previous studies by our laboratory have shown that a noncalcemic fluorinated analog of 1alpha,25-dihydroxyvitamin D3, 1alpha,25-dihydroxy-16-ene-23-yne-26,27-hexafluorocholcal ciferol (F6-D3), significantly reduced the frequency of colonic adenomas and completely abolished the development of colonic adenocarcinomas in rats treated with azoxymethane. The mechanisms involved in this analog's chemopreventive actions, however, remain unclear. In the present study, we now show that although both 1alpha,25-dihydroxyvitamin D3 and F6-D3 inhibited the proliferation of CaCo-2 cells, a human colonic adenocarcinoma cell line, by increasing their doubling times, only F6-D3 caused an arrest of these cells in the G1 phase of their cell cycle. This arrest was accompanied by an increase in the expression of the cyclin-dependent kinase (cdk) inhibitor proteins, p2Waf1 and p27Kip1, which served to decrease the activity of cyclin-dependent kinase 2 and cyclin-dependent kinase 6, whereas the expression and phosphorylation of pRB were unchanged. In contrast to the increased expression of these cdk inhibitors, the expression of cyclin E was decreased, which further inhibited the activity of cyclin-dependent kinase 2. Collectively, the inhibition of these cyclin-dependent kinases served to arrest the CaCo-2 cells, independent of changes in pRB. Furthermore, antibody neutralization studies suggest that transforming growth factor-beta may mediate the coassociations between cdk2 and p27Kip1 and cyclin E induced by F6-D3. These data indicate that cell cycle arrest may, at least in part, underlie the chemopreventive actions of F6-D3 observed in the azoxymethane model of colon cancer. Furthermore, if the antiproliferative action observed in CaCo-2 cells also occurs in human colonic epithelium, F6-D3 may have chemopreventive potential against human colon cancer, as well. PMID:11089522

  12. Ganglioside GM3 modulates tumor suppressor PTEN-mediated cell cycle progression--transcriptional induction of p21(WAF1) and p27(kip1) by inhibition of PI-3K/AKT pathway.

    PubMed

    Choi, Hee-Jung; Chung, Tae-Wook; Kang, Sung-Koo; Lee, Young-Choon; Ko, Jeong-Heon; Kim, Jong-Guk; Kim, Cheorl-Ho

    2006-07-01

    The simple ganglioside GM3 has been shown to have anti-proliferative effects in several in vitro and in vivo cancer models. Although the exogenous ganglioside GM3 has an inhibitory effect on cancer cell proliferation, the exact mechanism by which it prevents cell proliferation remains unclear. Previous studies showed that MDM2 is an oncoprotein that controls tumorigenesis through both p53-dependent and p53-independent mechanisms, and tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN), a dual-specificity phosphatase that antagonizes phosphatidylinositol 3-kinase (PI-3K)/AKT signaling, is capable of blocking MDM2 nuclear translocation and destabilizing the MDM2 protein. Results from our current study show that GM3 treatment dramatically increases cyclin-dependent kinase (CDK) inhibitor (CKI) p21(WAF1) expression through the accumulation of p53 protein by the PTEN-mediated inhibition of the PI-3K/AKT/MDM2 survival signaling in HCT116 colon cancer cells. Moreover, the data herein clearly show that ganglioside GM3 induces p53-dependent transcriptional activity of p21(WAF1), as evidenced by the p21(WAF1) promoter-driven luciferase reporter plasmid (full-length p21(WAF1) promoter and a construct lacking the p53-binding sites). Additionally, ganglioside GM3 enhances expression of CKI p27(kip1) through the PTEN-mediated inhibition of the PI-3K/AKT signaling. Furthermore, the down-regulation of the cyclin E and CDK2 was clearly observed in GM3-treated HCT116 cells, but the down-regulation of cyclin D1 and CDK4 was not. On the contrary, suppression of PTEN levels by RNA interference restores the enhanced expression of p53-dependent p21(WAF1) and p53-independent p27(kip1) through inactivating the effect of PTEN on PI-3K/AKT signaling modulated by ganglioside GM3. These results suggest that ganglioside GM3-stimulated PTEN expression modulates cell cycle regulatory proteins, thus inhibiting cell growth. We conclude that ganglioside GM3 represents a

  13. The Down syndrome-related protein kinase DYRK1A phosphorylates p27Kip1 and Cyclin D1 and induces cell cycle exit and neuronal differentiation

    PubMed Central

    Soppa, Ulf; Schumacher, Julian; Florencio Ortiz, Victoria; Pasqualon, Tobias; Tejedor, Francisco J; Becker, Walter

    2014-01-01

    A fundamental question in neurobiology is how the balance between proliferation and differentiation of neuronal precursors is maintained to ensure that the proper number of brain neurons is generated. Substantial evidence implicates DYRK1A (dual specificity tyrosine-phosphorylation-regulated kinase 1A) as a candidate gene responsible for altered neuronal development and brain abnormalities in Down syndrome. Recent findings support the hypothesis that DYRK1A is involved in cell cycle control. Nonetheless, how DYRK1A contributes to neuronal cell cycle regulation and thereby affects neurogenesis remains poorly understood. In the present study we have investigated the mechanisms by which DYRK1A affects cell cycle regulation and neuronal differentiation in a human cell model, mouse neurons, and mouse brain. Dependent on its kinase activity and correlated with the dosage of overexpression, DYRK1A blocked proliferation of SH-SY5Y neuroblastoma cells within 24 h and arrested the cells in G1 phase. Sustained overexpression of DYRK1A induced G0 cell cycle exit and neuronal differentiation. Furthermore, we provide evidence that DYRK1A modulated protein stability of cell cycle-regulatory proteins. DYRK1A reduced cellular Cyclin D1 levels by phosphorylation on Thr286, which is known to induce proteasomal degradation. In addition, DYRK1A phosphorylated p27Kip1 on Ser10, resulting in protein stabilization. Inhibition of DYRK1A kinase activity reduced p27Kip1 Ser10 phosphorylation in cultured hippocampal neurons and in embryonic mouse brain. In aggregate, these results suggest a novel mechanism by which overexpression of DYRK1A may promote premature neuronal differentiation and contribute to altered brain development in Down syndrome. PMID:24806449

  14. The inhibition of activated hepatic stellate cells proliferation by arctigenin through G0/G1 phase cell cycle arrest: persistent p27(Kip1) induction by interfering with PI3K/Akt/FOXO3a signaling pathway.

    PubMed

    Li, Ao; Wang, Jun; Wu, Mingjun; Zhang, Xiaoxun; Zhang, Hongzhi

    2015-01-15

    Proliferation of hepatic stellate cells (HSCs) is vital for the development of fibrosis during liver injury. In this study, we describe that arctigenin (ATG), a major bioactive component of Fructus Arctii, exhibited selective cytotoxic activity via inhibiting platelet-derived growth factor-BB (PDGF-BB)-activated HSCs proliferation and arrested cell cycle at G0/G1 phase, which could not be observed in normal human hepatocytes in vitro. The cyclin-dependent kinase (CDK) 4/6 activities could be strongly inhibited by ATG through down-regulation of cyclin D1 and CDK4/6 expression in early G1 phase arrest. In the ATG-treated HSCs, the expression level of p27(Kip1) and the formation of CDK2-p27(Kip1) complex were also increased. p27(Kip1) silencing significantly attenuated the effect of ATG, including cell cycle arrest and suppression of proliferation in activated HSCs. We also found that ATG suppressed PDGF-BB-induced phosphorylation of Akt and its downstream transcription factor Forkhead box O 3a (FOXO3a), decreased binding of FOXO3a to 14-3-3 protein, and stimulated nuclear translocation of FOXO3a in activated HSCs. Furthermore, knockdown of FOXO3a expression by FOXO3a siRNA attenuated ATG-induced up-regulation of p27(Kip1) in activated HSCs. All the above findings suggested that ATG could increase the levels of p27(Kip1) protein through inhibition of Akt and improvement of FOXO3a activity, in turn inhibited the CDK2 kinase activity, and eventually caused an overall inhibition of HSCs proliferation. PMID:25498792

  15. FGFR1 signaling stimulates proliferation of human mesenchymal stem cells by inhibiting the cyclin-dependent kinase inhibitors p21(Waf1) and p27(Kip1).

    PubMed

    Dombrowski, Christian; Helledie, Torben; Ling, Ling; Grünert, Martin; Canning, Claire A; Jones, C Michael; Hui, James H; Nurcombe, Victor; van Wijnen, Andre J; Cool, Simon M

    2013-12-01

    Signaling through fibroblast growth factor receptor one (FGFR1) is a known inducer of proliferation in both embryonic and human adult mesenchymal stem cells (hMSCs) and positively regulates maintenance of stem cell viability. Leveraging the mitogenic potential of FGF2/FGFR1 signaling in stem cells for therapeutic applications necessitates a mechanistic understanding of how this receptor stimulates cell cycle progression. Using small interfering RNA (siRNA) depletion, antibody-inhibition, and small molecule inhibition, we establish that FGFR1 activity is rate limiting for self-renewal of hMSCs. We show that FGFR1 promotes stem cell proliferation through multiple mechanisms that unite to antagonize cyclin-dependent kinase (CDK) inhibitors. FGFR1 not only stimulates c-Myc to suppress transcription of the CDK inhibitors p21(Waf1) and p27(Kip1), thus promoting cell cycle progression but also increases the activity of protein kinase B (AKT) and the level of S-phase kinase-associated protein 2 (Skp2), resulting in the nuclear exclusion and reduction of p21(Waf1). The in vivo importance of FGFR1 signaling for the control of proliferation in mesenchymal progenitor populations is underscored by defects in ventral mesoderm formation during development upon inhibition of its signaling. Collectively, these studies demonstrate that FGFR1 signaling mediates the continuation of MSC growth and establishes a receptor target for enhancing the expansion of mesenchymal progenitors while maintaining their multilineage potential. PMID:23939995

  16. Cdc6 Protein Activates p27KIP1-bound Cdk2 Protein Only after the Bound p27 Protein Undergoes C-terminal Phosphorylation*

    PubMed Central

    Uranbileg, Baasanjav; Yamamoto, Hanako; Park, Jung-ha; Mohanty, Atish R.; Arakawa-Takeuchi, Shiho; Jinno, Shigeki; Okayama, Hiroto

    2012-01-01

    In mammalian cells Cdk2 activity during the G1-S transition is mainly controlled by p27KIP1. Although the amount and subcellular localization of p27 influence Cdk2 activity, how Cdk2 activity is regulated during this phase transition still remains virtually unknown. Here we report an entirely new mechanism for this regulation. Cdc6 the AAA+ ATPase, known to assemble prereplicative complexes on chromosomal replication origins and activate p21CIP1-bound Cdk2, also activated p27-bound Cdk2 in its ATPase and cyclin binding motif-dependent manner but only after the p27 bound to the Cdk2 was phosphorylated at the C terminus. ROCK, which mediates a signal for cell anchorage to the extracellular matrix and activates the mTORC1 cascade as well as controls cytoskeleton assembly, was partly responsible for C-terminal phosphorylation of the p27. In vitro reconstitution demonstrated ROCK (Rho-associated kinase)-mediated phosphorylation of Cdk2-bound p27 at the C terminus and subsequent activation of the Cdk2 by Cdc6. PMID:22223646

  17. Expression of p27Kip1, a cell cycle repressor protein, is inversely associated with potential carcinogenic risk in the genetic rodent models of obesity and long-lived Ames dwarf mice

    PubMed Central

    Eto, Isao

    2014-01-01

    Introduction The association of genetic rodent models of obesity and cancer still remains a controversial issue. Although this controversy has largely been resolved in recent years for homozygous leptin receptor-deficient obese Zucker rats and homozygous long-lived Ames dwarf mice, it is still unresolved for homozygous leptin-deficient obese ob/ob mice. Objective The objective of the present study described below was to investigate whether the expression of the cell cycle repressor protein p27(Kip1) is (a) down-regulated in the tumor-free homozygous leptin receptor-deficient obese Zucker rats as well as tumor-free homozygous leptin-deficient obese ob/ob mice and (b) up-regulated in the tumor-free homozygous long-lived Ames dwarf mice. Methods To achieve this objective, we first performed western immunoblot analysis of the hepatic expression of p27. We then performed western immunoblot analysis and proteomic analysis of the hepatic expression of the proteins involved in the upstream molecular signaling pathways for the expression of p27. Lastly, we analyzed the serum levels of glucose, insulin, and branched-chain amino acids, all of which have been shown to regulate, causally and inversely, the expression of p27. Results/Conclusions The results indicated that the hepatic expression of p27 was down-regulated in the homozygous leptin receptor-deficient obese Zucker rats and up-regulated in the homozygous long-lived Ames dwarf mice as expected. We also found that the hepatic expression of p27 was down-regulated in the homozygous leptin-deficient obese ob/ob mice. This last observation was not completely consistent with all of the results of the published studies where homozygous leptin-deficient obese ob/ob mice were used. PMID:23357529

  18. The MLL fusion gene, MLL-AF4, regulates cyclin-dependent kinase inhibitor CDKN1B (p27kip1) expression

    PubMed Central

    Xia, Zhen-Biao; Popovic, Relja; Chen, Jing; Theisler, Catherine; Stuart, Tara; Santillan, Donna A.; Erfurth, Frank; Diaz, Manuel O.; Zeleznik-Le, Nancy J.

    2005-01-01

    MLL, involved in many chromosomal translocations associated with acute myeloid and lymphoid leukemia, has >50 known partner genes with which it is able to form in-frame fusions. Characterizing important downstream target genes of MLL and of MLL fusion proteins may provide rational therapeutic strategies for the treatment of MLL-associated leukemia. We explored downstream target genes of the most prevalent MLL fusion protein, MLL-AF4. To this end, we developed inducible MLL-AF4 fusion cell lines in different backgrounds. Overexpression of MLL-AF4 does not lead to increased proliferation in either cell line, but rather, cell growth was slowed compared with similar cell lines inducibly expressing truncated MLL. We found that in the MLL-AF4-induced cell lines, the expression of the cyclin-dependent kinase inhibitor gene CDKN1B was dramatically changed at both the RNA and protein (p27kip1) levels. In contrast, the expression levels of CDKN1A (p21) and CDKN2A (p16) were unchanged. To explore whether CDKN1B might be a direct target of MLL and of MLL-AF4, we used chromatin immunoprecipitation (ChIP) assays and luciferase reporter gene assays. MLL-AF4 binds to the CDKN1B promoter in vivo and regulates CDKN1B promoter activity. Further, we confirmed CDKN1B promoter binding by ChIP in MLL-AF4 as well as in MLL-AF9 leukemia cell lines. Our results suggest that CDKN1B is a downstream target of MLL and of MLL-AF4, and that, depending on the background cell type, MLL-AF4 inhibits or activates CDKN1B expression. This finding may have implications in terms of leukemia stem cell resistance to chemotherapy in MLL-AF4 leukemias. PMID:16169901

  19. Ras activity late in G1 phase required for p27kip1 downregulation, passage through the restriction point, and entry into S phase in growth factor-stimulated NIH 3T3 fibroblasts.

    PubMed Central

    Takuwa, N; Takuwa, Y

    1997-01-01

    It is well documented that Ras functions as a molecular switch for reentry into the cell cycle at the border between G0 and G1 by transducing extracellular growth stimuli into early G1 mitogenic signals. In the present study, we investigated the role of Ras during the late stage of the G1 phase by using NIH 3T3 (M17) fibroblasts in which the expression of a dominant negative Ras mutant, p21(Ha-Ras[Asn17]), is induced in response to dexamethasone treatment. We found that delaying the expression of Ras(Asn17) until late in the G1 phase by introducing dexamethasone 3 h after the addition of epidermal growth factor (EGF) abolished the downregulation of the p27kip1 cyclin-dependent kinase (CDK) inhibitor which normally occurred during this period, with resultant suppression of cyclin Ds/CDK4 and cyclin E/CDK2 and G1 arrest. The immunodepletion of p27kip1 completely eliminated the CDK inhibitor activity from EGF-stimulated, dexamethasone-treated cell lysate. The failure of p27kip1 downregulation and G1 arrest was also observed in cells in which Ras(Asn17) was induced after growth stimulation with a phorbol ester or alpha-thrombin and was mimicked by the addition late in the G1 phase of inhibitors for phosphatidylinositol-3-kinase. Ras-mediated downregulation of p27kip1 involved both the suppression of synthesis and the stimulation of the degradation of the protein. Unlike the earlier expression of Ras(Asn17) at the border between G0 and G1, its delayed expression did not compromise the EGF-stimulated transient activation of extracellular signal-regulated kinases or inhibit the stimulated expression of a principal D-type cyclin, cyclin D1, until close to the border between G1 and S. We conclude that Ras plays temporally distinct, phase-specific roles throughout the G1 phase and that Ras function late in G1 is required for p27kip1 downregulation and passage through the restriction point, a prerequisite for entry into the S phase. PMID:9271412

  20. Fad24, a Positive Regulator of Adipogenesis, Is Required for S Phase Re-entry of C2C12 Myoblasts Arrested in G0 Phase and Involved in p27(Kip1) Expression at the Protein Level.

    PubMed

    Ochiai, Natsuki; Nishizuka, Makoto; Osada, Shigehiro; Imagawa, Masayoshi

    2016-05-01

    Factor for adipocyte differentiation 24 (fad24) is a positive regulator of adipogenesis. We previously found that human fad24 is abundantly expressed in skeletal muscle. However, the function of fad24 in skeletal muscle remains largely unknown. Because skeletal muscle is a highly regenerative tissue, we focused on the function of fad24 in skeletal muscle regeneration. In this paper, we investigated the role of fad24 in the cell cycle re-entry of quiescent C2C12 myoblasts-mimicked satellite cells. The expression levels of fad24 and histone acetyltransferase binding to ORC1 (hbo1), a FAD24-interacting factor, were elevated at the early phase of the regeneration process in response to cardiotoxin-induced muscle injury. The knockdown of fad24 inhibited the proliferation of quiescent myoblasts, whereas fad24 knockdown did not affect differentiation. S phase entry following serum activation is abrogated by fad24 knockdown in quiescent cells. Furthermore, fad24 knockdown cells show a marked accumulation of p27(Kip1) protein. These results suggest that fad24 may have an important role in the S phase re-entry of quiescent C2C12 cells through the regulation of p27(Kip1) at the protein level. PMID:26902224

  1. Molecular cloning and functional characterization of a cell-permeable superoxide dismutase targeted to lung adenocarcinoma cells. Inhibition cell proliferation through the Akt/p27kip1 pathway.

    PubMed

    Lu, Min; Gong, Xingguo; Lu, Yuwen; Guo, Jianjun; Wang, Chenhui; Pan, Yuanjiang

    2006-05-12

    In clinical oncology, many trials with superoxide dismutase (SOD) have failed to demonstrate antitumor ability and in many cases even caused deleterious effects because of low tumor-targeting ability. In the current research, the Nostoc commune Fe-SOD coding sequence was amplified from genomic DNA. In addition, the single chain variable fragment (ScFv) was constructed from the cDNA of an LC-1 hybridoma cell line secreting anti-lung adenocarcinoma monoclonal antibody. After modification, the SOD and ScFv were fused and co-expressed, and the resulting fusion protein produced SOD and LC-1 antibody activity. Tracing SOD-ScFv by fluorescein isothiocyanate and superoxide anions (O2*-) in SPC-A-1 cells showed that the fusion protein could recognize and enter SPC-A-1 cells to eliminate O2*-. The lower oxidative stress resulting from the decrease in cellular O2*- delayed the cell cycle at G1 and significantly slowed SPC-A-1 cell growth in association with the dephosphorylation of the serine-threonine protein kinase Akt and expression of p27kip1. The tumor-targeting fusion protein resulting from this research overcomes two disadvantages of SODs previously used in the clinical setting, the inability to target tumor cells or permeate the cell membrane. These findings lay the groundwork for development of an efficient antitumor drug targeted by the ScFv. PMID:16551617

  2. Structural basis of the Cks1-dependent recognition of p27(Kip1) by the SCF(Skp2) ubiquitin ligase

    SciTech Connect

    Hao, B.; Zheng, N.; Schulman, B.A.; Wu, G.; Miller, J.J.; Pagano, M.; Pavletich, N.P.

    2010-07-19

    The ubiquitin-mediated proteolysis of the Cdk2 inhibitor p27{sup Kip1} plays a central role in cell cycle progression, and enhanced degradation of p27{sup Kip1} is associated with many common cancers. Proteolysis of p27{sup Kip1} is triggered by Thr187 phosphorylation, which leads to the binding of the SCF{sup Skp2} (Skp1-Cul1-Rbx1-Skp2) ubiquitin ligase complex. Unlike other known SCF substrates, p27{sup Kip1} ubiquitination also requires the accessory protein Cks1. The crystal structure of the Skp1-Skp2-Cks1 complex bound to a p27{sup Kip1} phosphopeptide shows that Cks1 binds to the leucine-rich repeat (LRR) domain and C-terminal tail of Skp2, whereas p27{sup Kip1} binds to both Cks1 and Skp2. The phosphorylated Thr187 side chain of p27{sup Kip1} is recognized by a Cks1 phosphate binding site, whereas the side chain of an invariant Glu185 inserts into the interface between Skp2 and Cks1, interacting with both. The structure and biochemical data support the proposed model that Cdk2-cyclin A contributes to the recruitment of p27{sup Kip1} to the SCF{sup Skp2}-Cks1 complex.

  3. Targeting the cyclin-binding groove site to inhibit the catalytic activity of CDK2/cyclin A complex using p27(KIP1)-derived peptidomimetic inhibitors.

    PubMed

    Karthiga, Arumugasamy; Tripathi, Sunil Kumar; Shanmugam, Ramasamy; Suryanarayanan, Venkatesan; Singh, Sanjeev Kumar

    2015-01-01

    Functionally activated cyclin-dependent kinase 2 (CDK2)/cyclin A complex has been validated as an interesting therapeutic target to develop the efficient antineoplastic drug based on the cell cycle arrest. Cyclin A binds to CDK2 and activates the kinases as well as recruits the substrate and inhibitors using a hydrophobic cyclin-binding groove (CBG). Blocking the cyclin substrate recruitment on CBG is an alternative approach to override the specificity hurdle of the currently available ATP site targeting CDK2 inhibitors. Greater understanding of the interaction of CDK2/cyclin A complex with p27 (negative regulator) reveals that the Leu-Phe-Gly (LFG) motif region of p27 binds with the CBG site of cyclin A to arrest the malignant cell proliferation that induces apoptosis. In the present study, Replacement with Partial Ligand Alternatives through Computational Enrichment (REPLACE) drug design strategies have been applied to acquire LFG peptide-derived peptidomimetics library. The peptidomimetics function is equivalent with respect to substrate p27 protein fashion but does not act as an ATP antagonist. The combined approach of molecular docking, molecular dynamics (MD), and molecular electrostatic potential and ADME/T prediction were carried out to evaluate the peptidomimetics. Resultant interaction and electrostatic potential maps suggested that smaller substituent is desirable at the position of phenyl ring to interact with Trp217, Arg250, and Gln254 residues in the active site. The best docked poses were refined by the MD simulations which resulted in conformational changes. After equilibration, the structure of the peptidomimetic and receptor complex was stable. The results revealed that the various substrate protein-derived peptidomimetics could serve as perfect leads against CDK2 protein. PMID:25584078

  4. KLRG1 impairs CD4+ T cell responses via p16ink4a and p27kip1 pathways: role in hepatitis B vaccine failure in individuals with hepatitis C virus infection.

    PubMed

    Shi, Lei; Wang, Jia M; Ren, Jun P; Cheng, Yong Q; Ying, Ruo S; Wu, Xiao Y; Lin, Shu M; Griffin, Jeddidiah W D; Li, Guang Y; Moorman, Jonathan P; Yao, Zhi Q

    2014-01-15

    Coinfection of hepatitis B virus (HBV) with hepatitis C virus (HCV) is quite common, leading to an increase in morbidity and mortality. As such, HBV vaccination is recommended in HCV-infected individuals. However, HBV vaccine responses in HCV-infected individuals are often blunted compared with uninfected populations. The mechanism for this failure of vaccine response in HCV-infected subjects remains unclear. In this study, we investigated the expression and function of an inhibitory receptor, killer cell lectin-like receptor subfamily G member 1 (KLRG1), in the regulation of CD4(+) T cells and HBV vaccine responses during HCV infection. We demonstrated that KLRG1 was overexpressed on CD4(+) T cells from HCV-infected, HBV vaccine nonresponders compared with HBV vaccine responders. The capacity of CD4(+) T cells to proliferate and secrete IL-2 cytokine was inversely associated with the level of KLRG1 expression. Importantly, blocking KLRG1 signaling resulted in a significant improvement in CD4(+) T cell proliferation and IL-2 production in HCV-infected, HBV vaccine nonresponders in response to TCR stimulation. Moreover, blockade of KLRG1 increased the phosphorylation of Akt (Ser(473)) and decreased the expression of cell cycle inhibitors p16(ink4a) and p27(kip1), which subsequently enhanced the expression of cyclin-dependent kinase 2 and cyclin E. These results suggest that the KLRG1 pathway impairs CD4(+) T cell responses to neoantigen and induces a state of immune senescence in individuals with HCV infection, raising the possibility that blocking this negative-signaling pathway might improve HBV vaccine responses in the setting of chronic viral infection. PMID:24337749

  5. KLRG1 impairs CD4+ T cell responses via p16ink4a and p27kip1 pathways - Role in hepatitis B vaccine failure in individuals with hepatitis C virus infection

    PubMed Central

    Shi, Lei; Wang, Jia M.; Ren, Jun P.; Cheng, Yong Q.; Ying, Ruo S.; Wu, Xiao Y.; Lin, Shu M.; Griffin, Jeddidiah WD; Li, Guang Y.; Moorman, Jonathan P.; Yao, Zhi Q.

    2014-01-01

    Co-infection of hepatitis B virus (HBV) with hepatitis C virus (HCV) is quite common, leading to an increase in morbidity and mortality. As such, HBV vaccination is recommended in HCV-infected individuals. HBV vaccine responses in HCV-infected individuals, however, are often blunted when compared to uninfected populations. The mechanism for this failure of vaccine response in HCV-infected subjects remains unclear. In this study, we investigated the expression and function of an inhibitory receptor, killer cell lectin-like receptor subfamily G member 1 (KLRG1), in regulation of CD4+ T cells and HBV vaccine responses during HCV infection. We demonstrated that KLRG1 was over-expressed on CD4+ T cells from HCV-infected, HBV vaccine non-responders (HBV-NR) compared to those responders (HBV-R). The capacity of CD4+ T cell to proliferate and secrete IL-2 cytokine was inversely associated with the level of KLRG1 expression. Importantly, blocking KLRG1 signaling resulted in a significant improvement of CD4+ T cell proliferation and IL-2 production in HCV-infected, HBV-NR in response to T cell receptor (TCR) stimulation. Moreover, blockade of KLRG1 increased the phosphorylation of Akt (Ser473) and decreased the expression of cell cycle inhibitors p16ink4a and p27kip1, which subsequently enhanced CDK 2 and cyclin E expressions. These results suggest that the KLRG1 pathway impairs CD4+ T cell responses to neo-antigen and induces a state of immune senescence in individuals with HCV infection, raising the possibility that blocking this negative signaling pathway might improve HBV vaccine responses in the setting of chronic viral infection. PMID:24337749

  6. The expression of the ubiquitin ligase subunit Cks1 in human breast cancer

    PubMed Central

    Slotky, Merav; Shapira, Ma'anit; Ben-Izhak, Ofer; Linn, Shai; Futerman, Boris; Tsalic, Medy; Hershko, Dan D

    2005-01-01

    Introduction Loss of the cell-cycle inhibitory protein p27Kip1 is associated with a poor prognosis in breast cancer. The decrease in the levels of this protein is the result of increased proteasome-dependent degradation, mediated and rate-limited by its specific ubiquitin ligase subunits S-phase kinase protein 2 (Skp2) and cyclin-dependent kinase subunit 1 (Cks1). Skp2 was recently found to be overexpressed in breast cancers, but the role of Cks1 in these cancers is unknown. The present study was undertaken to examine the role of Cks1 expression in breast cancer and its relation to p27Kip1 and Skp2 expression and to tumor aggressiveness. Methods The expressions of Cks1, Skp2, and p27Kip1 were examined immunohistochemically on formalin-fixed, paraffin-wax-embedded tissue sections from 50 patients with breast cancer and by immunoblot analysis on breast cancer cell lines. The relation between Cks1 levels and patients' clinical and histological parameters were examined by Cox regression and the Kaplan–Meier method. Results The expression of Cks1 was strongly associated with Skp2 expression (r = 0.477; P = 0.001) and inversely with p27Kip1 (r = -0.726; P < 0.0001). Overexpression of Cks1 was associated with loss of tumor differentiation, young age, lack of expression of estrogen receptors and of progesterone receptors, and decreased disease-free (P = 0.0007) and overall (P = 0.041) survival. In addition, Cks1 and Skp2 expression were increased by estradiol in estrogen-dependent cell lines but were down-regulated by tamoxifen. Conclusion These results suggest that Cks1 is involved in p27Kip1 down-regulation and may have an important role in the development of aggressive tumor behavior in breast cancer. PMID:16168119

  7. The Effectiveness of Cucurbitacin B in BRCA1 Defective Breast Cancer Cells

    PubMed Central

    Chakrabarty, Subhas; Bögler, Oliver; Patmasiriwat, Pimpicha

    2013-01-01

    Cucurbitacin B (CuB) is one of the potential agents for long term anticancer chemoprevention. Cumulative evidences has shown that cucurbitacin B provides potent cellular biological activities such as hepatoprotective, anti-inflammatory and antimicrobial effects, but the precise mechanism of this agent is not clearly understood. We examine the biological effects on cancer cells of cucurbitacin B extracted from a Thai herb, Trichosanthes cucumerina L. The wild type (wt) BRCA1, mutant BRCA1, BRCA1 knocked-down and BRCA1 overexpressed breast cancer cells were treated with the cucurbitacin B and determined for the inhibitory effects on the cell proliferation, migration, invasion, anchorage-independent growth. The gene expressions in the treated cells were analyzed for p21/Waf1, p27Kip1 and survivin. Our previous study revealed that loss of BRCA1 expression leads to an increase in survivin expression, which is responsible for a reduction in sensitivity to paclitaxel. In this work, we showed that cucurbitacin B obviously inhibited knocked-down and mutant BRCA1 breast cancer cells rather than the wild type BRCA1 breast cancer cells in regards to the cellular proliferation, migration, invasion and anchorage-independent growth. Furthermore, forcing the cells to overexpress wild type BRCA1 significantly reduced effectiveness of cucurbitacin B on growth inhibition of the endogenous mutant BRCA1 cells. Interestingly, cucurbitacin B promotes the expression of p21/Waf1 and p27Kip1 but inhibit the expression of survivin. We suggest that survivin could be an important target of cucurbitacin B in BRCA1 defective breast cancer cells. PMID:23393598

  8. Chromosomal deletions and tumor suppressor genes in prostate cancer.

    PubMed

    Dong, J T

    2001-01-01

    Chromosomal deletion appears to be the earliest as well as the most frequent somatic genetic alteration during carcinogenesis. It inactivates a tumor suppressor gene in three ways, that is, revealing a gene mutation through loss of heterozygosity as proposed in the two-hit theory, inducing haploinsufficiency through quantitative hemizygous deletion and associated loss of expression, and truncating a genome by homozygous deletion. Whereas the two-hit theory has guided the isolation of many tumor suppressor genes, the haploinsufficiency hypothesis seems to be also useful in identifying target genes of chromosomal deletions, especially for the deletions detected by comparative genomic hybridization (CGH). At present, a number of chromosomal regions have been identified for their frequent deletions in prostate cancer, including 2q13-q33, 5q14-q23, 6q16-q22, 7q22-q32, 8p21-p22, 9p21-p22, 10q23-q24, 12p12-13, 13q14-q21, 16q22-24, and 18q21-q24. Strong candidate genes have been identified for some of these regions, including NKX3.1 from 8p21, PTEN from 10q23, p27/Kip1 from 12p13, and KLF5 from 13q21. In addition to their location in a region with frequent deletion, there are functional and/or genetic evidence supporting the candidacy of these genes. Thus far PTEN is the most frequently mutated gene in prostate cancer, and KLF5 showed the most frequent hemizygous deletion and loss of expression. A tumor suppressor role has been demonstrated for NKX3.1, PTEN, and p27/Kip1 in knockout mice models. Such genes are important targets of investigation for the development of biomarkers and therapeutic regimens. PMID:12085961

  9. Src kinases catalytic activity regulates proliferation, migration and invasiveness of MDA-MB-231 breast cancer cells.

    PubMed

    Sánchez-Bailón, María Pilar; Calcabrini, Annarica; Gómez-Domínguez, Daniel; Morte, Beatriz; Martín-Forero, Esther; Gómez-López, Gonzalo; Molinari, Agnese; Wagner, Kay-Uwe; Martín-Pérez, Jorge

    2012-06-01

    SFKs are frequently deregulated in cancer where they control cellular proliferation, migration, survival and metastasis. Here we study the role of SFKs catalytic activity in triple-negative/basal-like and metastatic human breast cancer MDA-MB-231 cells employing three well-established inhibitors: Dasatinib, PP2 and SU6656. These compounds inhibited migration and invasion. Concomitantly, they reduced Fak, paxillin, p130CAS, caveolin-1 phosphorylation and altered cytoskeletal structures. They also inhibited cell proliferation, but in different manners. Dasatinib and PP2 increased p27(Kip1) expression and reduced c-Myc levels, restraining G1–S transition. In contrast, SU6656 did not modify p27(Kip1) expression, slightly altered c-Myc levels and generated polyploid multinucleated cells, indicating inhibition of cytokinesis. These later effects were also observed in SYF fibroblasts, suggesting a SFKs-independent action. ZM447439, an Aurora B kinase inhibitor, produced similar cell cycle and morphological alterations in MDA-MB-231 cells, indicating that SU6656 blocked Aurora B kinase. This was confirmed by inhibition of histone H3 phosphorylation, the canonical Aurora B kinase substrate. Furthermore, hierarchical clustering analysis of gene expression profiles showed that SU6656 defined a set of genes that differed from Dasatinib and PP2. Additionally, Gene Set Enrichment Analyses revealed that SU6656 significantly reduces the Src pathway. Together, these results show the importance of SFKs catalytic activity for MDA-MB-231 proliferation, migration and invasiveness. They also illustrate that SU6656 acts as dual SFKs and Aurora B kinase inhibitor, suggesting its possible use as a therapeutic agent in breast cancer. PMID:22570868

  10. Decreased Skp2 Expression Is Necessary but Not Sufficient for Therapy-Induced Senescence in Prostate Cancer

    PubMed Central

    Ewald, Jonathan A; Jarrard, David F

    2012-01-01

    Therapy-induced senescence (TIS), a cytostatic stress response in cancer cells, is induced inefficiently by current anticancer agents and radiation. The mechanisms that mediate TIS in cancer cells are not well defined. Herein, we characterize a robust senescence response both in vitro and in vivo to the quinone diaziquone (AZQ), previously identified in a high-throughput senescence-induction small-molecule screen. Using AZQ and several other agents that induce senescence, we screened a series of cyclin-dependent kinase inhibitors and found that p27Kip1 was induced in all investigated prostate cancer cell lines. The ubiquitin-ligase Skp2 negatively regulates p27Kip1 and, during TIS, is translocated to the cytoplasm before its expression is decreased in senescent cells. Overexpression of Skp2 blocks the effects of AZQ on senescence and p27Kip1 induction. We also find that stable long-term short hairpin RNA knockdown of Skp2 decreases proliferation but does not generate the complete senescence phenotype. We conclude that Skp2 participates in regulating TIS but, alone, is insufficient to induce senescence in cancer cells. PMID:22937180

  11. Targeting prostate cancer based on signal transduction and cell cycle pathways

    PubMed Central

    Lee, John T.; Lehmann, Brian D.; Terrian, David M.; Chappell, William H.; Stivala, Franca; Libra, Massimo; Martelli, Alberto M.; Steelman, Linda S.

    2008-01-01

    Prostate cancer remains a leading cause of death in men despite increased capacity to diagnose at earlier stages. After prostate cancer has become hormone independent, which often occurs after hormonal ablation therapies, it is difficult to effectively treat. Prostate cancer may arise from mutations and dysregulation of various genes involved in regulation signal transduction (e.g., PTEN, Akt, etc.,) and the cell cycle (e.g., p53, p21Cip1, p27Kip1, Rb, etc.,). This review focuses on the aberrant interactions of signal transduction and cell cycle genes products and how they can contribute to prostate cancer and alter therapeutic effectiveness. PMID:18594202

  12. Protein Kinase C alpha (PKCα) dependent signaling mediates endometrial cancer cell growth and tumorigenesis

    PubMed Central

    Haughian, James M.; Reno, Elaine M.; Thorne, Alicia M.; Bradford, Andrew P.

    2009-01-01

    Endometrial cancer is the most common invasive gynecologic malignancy, yet molecular mechanisms and signaling pathways underlying its etiology and pathophysiology remain poorly characterized. We sought to define a functional role for the protein kinase C (PKC) isoform, PKCα, in an established cell model of endometrial adenocarcinoma. Ishikawa cells depleted of PKCα protein grew slower, formed fewer colonies in anchorage-independent growth assays and exhibited impaired xenograft tumor formation in nude mice. Consistent with impaired growth, PKCα knockdown increased levels of the cyclin dependent kinase (CDK) inhibitors p21Cip1/WAF1 (p21) and p27Kip1 (p27). Despite the absence of functional phosphatase and tensin homologue (PTEN) protein in Ishikawa cells, PKCα knockdown reduced Akt phosphorylation at serine 473 and concomitantly inhibited phosphorylation of the Akt target, glycogen synthase kinase-3β (GSK-3β). PKCα knockdown also resulted in decreased basal ERK phosphorylation and attenuated ERK activation following EGF stimulation. p21 and p27 expression was not increased by treatment of Ishikawa cells with ERK and Akt inhibitors, suggesting PKCα regulates CDK expression independently of Akt and ERK. Immunohistochemical analysis of grade 1 endometrioid adenocarcinoma revealed aberrant PKCα expression, with foci of elevated PKCα staining, not observed in normal endometrium. These studies demonstrate a critical role for PKCα signaling in endometrial tumorigenesis by regulating expression of CDK inhibitors p21 and p27 and activation of Akt and ERK dependent proliferative pathways. Thus, targeting PKCα may provide novel therapeutic options in endometrial tumors. PMID:19672862

  13. The soybean-derived peptide lunasin inhibits non-small cell lung cancer cell proliferation by suppressing phosphorylation of the retinoblastoma protein

    PubMed Central

    McConnell, Elizabeth J.; Devapatla, Bharat; Yaddanapudi, Kavitha; Davis, Keith R.

    2015-01-01

    Lunasin, a soybean bioactive peptide, has both chemopreventive and chemotherapeutic activities. The aim of this study was to determine the chemotherapeutic potential of lunasin against human lung cancer. Treatment of non-small cell lung cancer (NSCLC) cells with highly purified soybean-derived lunasin caused limited, cell-line specific anti-proliferative effects on anchorage-dependent growth whereas two normal bronchial epithelial cell lines were unaffected. Lunasin's antiproliferative effects were potentiated upon utilization of anchorage-independent conditions. Furthermore, NSCLC cell lines that were unaffected by lunasin in anchorage-dependent assays exhibited a dose-dependent inhibition in colony formation or colony size. Mouse xenograft studies revealed that 30 mg lunasin/kg body weight per day decreased NSCLC H1299 tumor volume by 63.0% at day 32. Mechanistic studies using cultured NSCLC H661 cells showed that lunasin inhibited cell cycle progression at the G1/S phase interface without inducing apoptosis. Immunoblot analyses of key cell-cycle proteins demonstrated that lunasin altered the expression of the G1 specific cyclin-dependent kinase complex components, increased levels of p27Kip1, reduced levels of phosphorylated Akt, and ultimately inhibited the sequential phosphorylation of the retinoblastoma protein (RB). These results establish for the first time that lunasin can inhibit NSCLC proliferation by suppressing cell-cycle dependent phosphorylation of RB. PMID:25609198

  14. Flavokawain A induces deNEDDylation and Skp2 degradation leading to inhibition of tumorigenesis and cancer progression in the TRAMP transgenic mouse model

    PubMed Central

    Li, Xuesen; Yokoyama, Noriko N.; Zhang, Saiyang; Ding, Lina; Liu, Hong-min; Lilly, Michael B.; Mercola, Dan; Zi, Xiaolin

    2015-01-01

    S phase kinase-associated protein 2 (Skp2) has been shown to be required for spontaneous tumor development that occurs in the retinoblastoma protein (pRb) deficient mice. Here we have demonstrated that flavokawain A (FKA), a novel chalcone from the kava plant, selectively inhibited the growth of pRb deficient cell lines and resulted in a proteasome-dependent and ubiquitination-mediated Skp2 degradation. Degradation of Skp2 by FKA was found to be involved in a functional Cullin1, but independent of Cdh1 expression. Further studies have demonstrated that FKA docked into the ATP binding pocket of the precursor cell-expressed developmentally down-regulated 8 (NEDD8)-activating enzyme (NAE) complex, inhibited NEDD8 conjugations to both Cullin1 and Ubc12 in PC3 cells and Ubc12 NEDDylation in an in vitro assay. Finally, dietary feeding of the autochthonous transgenic adenocarcinoma of the mouse prostate (TRAMP) mice with FKA inhibited the formation of high-grade prostatic intra-epithelial neoplasia lesions (HG-PIN) and prostate adenocarcinomas, reduced the tumor burden and completely abolished distant organ metastasis. Immunohistochemistry studies revealed that dietary FKA feeding resulted in marked anti-proliferative and apoptotic effects via down-regulation of Skp2 and NEDD8 and up-regulation of p27/Kip1 in the prostate of TRAMP mice. Our findings therefore provide evidence that FKA is a promising NEDDylation inhibitor for targeting Skp2 degradation in prostate cancer prevention and treatment. PMID:26497688

  15. Flavokawain A induces deNEDDylation and Skp2 degradation leading to inhibition of tumorigenesis and cancer progression in the TRAMP transgenic mouse model.

    PubMed

    Li, Xuesen; Yokoyama, Noriko N; Zhang, Saiyang; Ding, Lina; Liu, Hong-min; Lilly, Michael B; Mercola, Dan; Zi, Xiaolin

    2015-12-01

    S phase kinase-associated protein 2 (Skp2) has been shown to be required for spontaneous tumor development that occurs in the retinoblastoma protein (pRb) deficient mice. Here we have demonstrated that flavokawain A (FKA), a novel chalcone from the kava plant, selectively inhibited the growth of pRb deficient cell lines and resulted in a proteasome-dependent and ubiquitination-mediated Skp2 degradation. Degradation of Skp2 by FKA was found to be involved in a functional Cullin1, but independent of Cdh1 expression. Further studies have demonstrated that FKA docked into the ATP binding pocket of the precursor cell-expressed developmentally down-regulated 8 (NEDD8)-activating enzyme (NAE) complex, inhibited NEDD8 conjugations to both Cullin1 and Ubc12 in PC3 cells and Ubc12 NEDDylation in an in vitro assay. Finally, dietary feeding of the autochthonous transgenic adenocarcinoma of the mouse prostate (TRAMP) mice with FKA inhibited the formation of high-grade prostatic intra-epithelial neoplasia lesions (HG-PIN) and prostate adenocarcinomas, reduced the tumor burden and completely abolished distant organ metastasis. Immunohistochemistry studies revealed that dietary FKA feeding resulted in marked anti-proliferative and apoptotic effects via down-regulation of Skp2 and NEDD8 and up-regulation of p27/Kip1 in the prostate of TRAMP mice. Our findings therefore provide evidence that FKA is a promising NEDDylation inhibitor for targeting Skp2 degradation in prostate cancer prevention and treatment. PMID:26497688

  16. From microRNA functions to microRNA therapeutics: Novel targets and novel drugs in breast cancer research and treatment

    PubMed Central

    PIVA, ROBERTA; SPANDIDOS, DEMETRIOS A.; GAMBARI, ROBERTO

    MicroRNAs (miRNAs or miRs) are a family of small non-coding RNAs that regulate gene expression by the sequence-selective targeting of mRNAs, leading to translational repression or mRNA degradation, depending on the degree of complementarity with target mRNA sequences. miRNAs play a crucial role in cancer. In the case of breast tumors, several studies have demonstrated a correlation between: i) the expression profile of oncogenic miRNAs (oncomiRs) or tumor suppressor miRNAs and ii) the tumorigenic potential of triple-negative [estrogen receptor (ER), progesterone receptor (PR) and Her2/neu] primary breast cancers. Among the miRNAs involved in breast cancer, miR-221 plays a crucial role for the following reasons: i) miR-221 is significantly overexpressed in triple-negative primary breast cancers; ii) the oncosuppressor p27 Kip1 , a validated miR-221 target, is downregulated in aggressive cancer cell lines; and iii) the upregulation of a key transcription factor, Slug, appears to be crucial, since it binds to the miR-221/miR-222 promoter and is responsible for the high expression of the miR-221/miR-222 cluster in breast cancer cells. A Slug/miR-221 network has been suggested, linking miR-221 activity with the downregulation of a Slug repressor, leading to Slug/miR-221 upregulation and p27 Kip1 downregulation. Interference with this process can be achieved using antisense miRNA (antagomiR) molecules targeting miR-221, inducing the down-regulation of Slug and the upregulation of p27 Kip1 . PMID:23939688

  17. Hypersensitivity to contact inhibition provides a clue to cancer resistance of naked mole-rat.

    PubMed

    Seluanov, Andrei; Hine, Christopher; Azpurua, Jorge; Feigenson, Marina; Bozzella, Michael; Mao, Zhiyong; Catania, Kenneth C; Gorbunova, Vera

    2009-11-17

    The naked mole-rat is the longest living rodent with a maximum lifespan exceeding 28 years. In addition to its longevity, naked mole-rats have an extraordinary resistance to cancer as tumors have never been observed in these rodents. Furthermore, we show that a combination of activated Ras and SV40 LT fails to induce robust anchorage-independent growth in naked mole-rat cells, while it readily transforms mouse fibroblasts. The mechanisms responsible for the cancer resistance of naked mole-rats were unknown. Here we show that naked mole-rat fibroblasts display hypersensitivity to contact inhibition, a phenomenon we termed "early contact inhibition." Contact inhibition is a key anticancer mechanism that arrests cell division when cells reach a high density. In cell culture, naked mole-rat fibroblasts arrest at a much lower density than those from a mouse. We demonstrate that early contact inhibition requires the activity of p53 and pRb tumor suppressor pathways. Inactivation of both p53 and pRb attenuates early contact inhibition. Contact inhibition in human and mouse is triggered by the induction of p27(Kip1). In contrast, early contact inhibition in naked mole-rat is associated with the induction of p16(Ink4a). Furthermore, we show that the roles of p16(Ink4a) and p27(Kip1) in the control of contact inhibition became temporally separated in this species: the early contact inhibition is controlled by p16(Ink4a), and regular contact inhibition is controlled by p27(Kip1). We propose that the additional layer of protection conferred by two-tiered contact inhibition contributes to the remarkable tumor resistance of the naked mole-rat. PMID:19858485

  18. Proliferation and Cdk4 expression in microsatellite unstable colon cancers with TGFBR2 mutations.

    PubMed

    Grady, William M; Willis, Joseph E; Trobridge, Patty; Romero-Gallo, Judith; Munoz, Nina; Olechnowicz, Joseph; Ferguson, Kelly; Gautam, Shiva; Markowitz, Sanford D

    2006-02-01

    Approximately 15% of human colon cancers have microsatellite instability (MSI) and carry frameshift mutations in a polyadenine tract (BAT-RII) in the type II transforming growth factor beta (TGF-beta) receptor (TGFBR2), a required component of the TGF-beta receptor. The BAT-RII mutations in MSI colon cancers make the tumors resistant to the effects of TGF-beta. In cultured epithelial cells, TGF-beta can inhibit cell proliferation and induce apoptosis, and in vitro it can regulate the expression of a variety of cyclins, cyclin-dependent kinases (cdks) and cdk inhibitors. These effects are context- and tissue type-dependent, raising questions about which of these in vitro effects of TGF-beta signaling inactivation contribute to the formation of primary colon cancer. Thus, this study sought to determine the pathogenetically relevant effects of TGFBR2 inactivation in primary MSI colon cancers with mutant BAT-RII. Colon cancers with mutant BAT-RII were found to have increased proliferation compared to cancers with wild-type BAT-RII. Assessment of cdk4, cyclin D1 and p27(kip1) expression revealed that only cdk4 expression was increased in the cancers with mutant BAT-RII. In order to determine if TGFBR2 inactivation was the cause of these changes, TGFBR2 was reconstituted in an MSI colon cancer cell line, resulting in decreased proliferation and decreased cdk4 expression and kinase activity. These results suggest that TGFBR2 mutations in primary colon cancers may be responsible for the increased proliferation and cdk4 expression in these tumors and provide evidence that deregulation of cdk4 is a pathogenic in vivo consequence of TGFBR2 inactivation in primary colon cancer. PMID:16108056

  19. Organ differences in the impact of p27(kip1) deficiency on carcinogenesis induced by N-methyl-N-nitrosourea.

    PubMed

    Ogawa, Kumiko; Murasaki, Toshiya; Sugiura, Satoshi; Nakanishi, Makoto; Shirai, Tomoyuki

    2013-06-01

    To evaluate the impact of p27 on carcinogenesis in various organs, N-methyl-N-nitrosourea (MNU), a direct-acting alkylating agent, was given to p27 knock-out mice. Groups of 20-40 male and female mice with null, hetero- or wild-type p27 alleles were given drinking water containing 240 ppm MNU or distilled water every other week for five cycles. The incidence and multiplicity of the induced proliferative lesions were then histologically evaluated at weeks 14 and 20. MNU treatment induced various lesions including squamous hyperplasia and squamous cell carcinoma in the forestomach, atypical hyperplasia and adenocarcinomas in the fundic and pyloric glands, adenomas and adenocarcinomas in the duodenum, malignant lymphomas in the thymus, liver, kidney and spleen and alveolar hyperplasia, adenomas, adenocarcinomas and malignant lymphomas in the lung. Although the incidences of the lesions in the forestomach, fundic and pyloric glands did not differ among the p27 genotypes, those of alveolar hyperplasia of the lung and malignant lymphoma of the thymus were significantly increased in p27-null males as compared with both wild- and hetero-type animals. Moreover, in both p27(+/+) and p27(+/-) cases, the rates for p27-positive cells were obviously increased in proliferative lesions of the pyloric gland and the lung. However, an increased rate of p27-positive cells was not observed in malignant lymphoma of the thymus. These findings suggest that p27 does not control the cell cycle equally in all organs affected by MNU-induced carcinogenesis. PMID:22183835

  20. A pathway in quiescent cells that controls p27Kip1 stability, subcellular localization, and tumor suppression

    PubMed Central

    Besson, Arnaud; Gurian-West, Mark; Chen, Xueyan; Kelly-Spratt, Karen S.; Kemp, Christopher J.; Roberts, James M.

    2006-01-01

    We have created two knock-in mouse models to study the mechanisms that regulate p27 in normal cells and cause misregulation of p27 in tumors: p27S10A, in which Ser10 is mutated to Ala; and p27CK–, in which point mutations abrogate the ability of p27 to bind cyclins and CDKs. These two mutant alleles identify steps in a pathway that controls the proteasomal degradation of p27 uniquely in quiescent cells: Dephosphorylation of p27 on Ser10 inhibits p27 nuclear export and promotes its assembly into cyclin–CDK complexes, which is, in turn, necessary for p27 turnover. We further show that Ras-dependent lung tumorigenesis is associated with increased phosphorylation on Ser10 and cytoplasmic mislocalization of p27. Indeed, we find that p27S10A is refractory to Ras-induced cytoplasmic translocation and that p27S10A mice are tumor resistant. Thus, phosphorylation of p27 on Ser10 is an important event in the regulation of the tumor suppressor function of p27. PMID:16391232

  1. CDK2 and mTOR are direct molecular targets of isoangustone A in the suppression of human prostate cancer cell growth.

    PubMed

    Lee, Eunjung; Son, Joe Eun; Byun, Sanguine; Lee, Seung Joon; Kim, Yeong A; Liu, Kangdong; Kim, Jiyoung; Lim, Soon Sung; Park, Jung Han Yoon; Dong, Zigang; Lee, Ki Won; Lee, Hyong Joo

    2013-10-01

    Licorice extract which is used as a natural sweetener has been shown to possess inhibitory effects against prostate cancer, but the mechanisms responsible are poorly understood. Here, we report a compound, isoangustone A (IAA) in licorice that potently suppresses the growth of aggressive prostate cancer and sought to clarify its mechanism of action. We analyzed its inhibitory effects on the growth of PTEN-deleted human prostate cancer cells, in vitro and in vivo. Administration of IAA significantly attenuated the growth of prostate cancer cell cultures and xenograft tumors. These effects were found to be attributable to inhibition of the G1/S phase cell cycle transition and the accumulation of p27(kip1). The elevated p27(kip1) expression levels were concurrent with the decrease of its phosphorylation at threonine 187 through suppression of CDK2 kinase activity and the reduced phosphorylation of Akt at Serine 473 by diminishing the kinase activity of the mammalian target of rapamycin (mTOR). Further analysis using recombinant proteins and immunoprecipitated cell lysates determined that IAA exerts suppressive effects against CDK2 and mTOR kinase activity by direct binding with both proteins. These findings suggested that the licorice compound IAA is a potent molecular inhibitor of CDK2 and mTOR, with strong implications for the treatment of prostate cancer. Thus, licorice-derived extracts with high IAA content warrant further clinical investigation for nutritional sources for prostate cancer patients. PMID:23707764

  2. D-alpha-tocopheryl polyethylene glycol succinate (TPGS) induces cell cycle arrest and apoptosis selectively in Survivin-overexpressing breast cancer cells.

    PubMed

    Neophytou, Christiana M; Constantinou, Constantina; Papageorgis, Panagiotis; Constantinou, Andreas I

    2014-05-01

    D-alpha-tocopheryl polyethylene glycol succinate (TPGS) is a vitamin E derivative that has been intensively applied as a vehicle for drug delivery systems to enhance drug solubility and increase the oral bioavailability of anti-cancer drugs. Recently, it has been reported that TPGS acts as an anti-cancer agent alone or synergistically with chemotherapeutic drugs and increases the efficacy of nanoparticle formulations. In this study, we investigated the antitumor efficacy and the molecular mechanism of action of TPGS in breast cancer cell lines. Our results show that TPGS can induce G1/S cell cycle arrest and apoptosis in breast cancer cell lines (MCF-7 and MDA-MB-231) but not in "normal" (non-tumorigenic) immortalized cells (MCF-10A and MCF-12F). An investigation of the molecular mechanism of action of TPGS reveals that induction of G1/S phase cell cycle arrest is associated with upregulation of P21 and P27Kip1 proteins. Induction of apoptosis by TPGS involves the inhibition of phospho-AKT and the downregulation of the anti-apoptotic proteins Survivin and Bcl-2. Interestingly, our results also suggest that TPGS induces both caspase -dependent and -independent apoptotic signaling pathways and that this vitamin E derivative is selectively cytotoxic in breast cancer cell lines. When compared to the Survivin inhibitor YM155, TPGS was shown to be more selective for cancer cell growth inhibition. Overall our results suggest that TPGS may not only be useful as a carrier molecule for drug delivery, but may also exert intrinsic therapeutic effects suggesting that it may promote a synergistic interaction with formulated chemotherapeutic drugs. PMID:24560876

  3. Astaxanthin Inhibits Proliferation of Human Gastric Cancer Cell Lines by Interrupting Cell Cycle Progression

    PubMed Central

    Kim, Jung Ha; Park, Jong-Jae; Lee, Beom Jae; Joo, Moon Kyung; Chun, Hoon Jai; Lee, Sang Woo; Bak, Young-Tae

    2016-01-01

    Background/Aims Astaxanthin is a carotenoid pigment that has antioxidant, antitumoral, and anti-inflammatory properties. In this in vitro study, we investigated the mechanism of anticancer effects of astaxanthin in gastric carcinoma cell lines. Methods The human gastric adenocarcinoma cell lines AGS, KATO-III, MKN-45, and SNU-1 were treated with various concentrations of astaxanthin. A cell viability test, cell cycle analysis, and immunoblotting were performed. Results The viability of each cancer cell line was suppressed by astaxanthin in a dose-dependent manner with significantly decreased proliferation in KATO-III and SNU-1 cells. Astaxanthin increased the number of cells in the G0/G1 phase but reduced the proportion of S phase KATO-III and SNU-1 cells. Phosphorylated extracellular signal-regulated kinase (ERK) was decreased in an inverse dose-dependent correlation with astaxanthin concentration, and the expression of p27kip-1 increased the KATO-III and SNU-1 cell lines in an astaxanthin dose-dependent manner. Conclusions Astaxanthin inhibits proliferation by interrupting cell cycle progression in KATO-III and SNU-1 gastric cancer cells. This may be caused by the inhibition of the phosphorylation of ERK and the enhanced expression of p27kip-1. PMID:26470770

  4. Latex of Euphorbia antiquorum-induced S-phase arrest via active ATM kinase and MAPK pathways in human cervical cancer HeLa cells.

    PubMed

    Hsieh, Wen-Tsong; Lin, Hui-Yi; Chen, Jou-Hsuan; Lin, Wen-Chung; Kuo, Yueh-Hsiung; Wood, W Gibson; Lu, Hsu-Feng; Chung, Jing-Gung

    2015-09-01

    Latex of Euphorbia antiquorum (EA) has demonstrated great chemotherapeutic potential for cancer. However, the mechanisms of anti-proliferation of EA on cancer cell remain to be further investigated. The purpose of this study was to explore the influence of EA in human cervical cancer cells. Here, the cell cycle distribution by flow cytometry was examined and the protein expression by the western blotting methods was analyzed. From the cytometric results it was shown that EA-induced S-phase arrest in a concentration manner both in human cervical cancer HeLa and CaSki cells. According the western blot results it was illustrated that EA could downregulate early cyclin E1-Cdk2; and cyclin A-Cdc2 provides a significant additional quantity of S-phase promotion, that in turn promoted the expression of p21(waf1/cip1) and p27(kip1) which were the inhibitors in the complex of cyclin A and Cdc2 that led to cell cycle arrest. Moreover, EA promoted the activation of ataxia telangiectasia mutated (ATM) and check-point kinase-2 (Chk2); however, it negatively regulated the expression of Topoisomerases I and II, Cdc25A, and Cdc25C signaling. Caffeine, an ATM/ATR inhibitor significantly reversed EA downregulation in the levels of Cdc25A. Furthermore, JNK inhibitor SP600125 and p38 MAPK inhibitor SB203580 both could reverse the EA upregulation of the protein of Chk2 level, significantly. This study, therefore, revealed that EA could downregulate topoisomerase, and activate ATM kinase, which then induce parallel Chk 1/2 and MAPK signaling pathways to promote the degradation of Cdc25A to induced S-phase arrest in human cervical cancer HeLa cells. PMID:24706497

  5. Combination treatment with proteasome inhibitors and antiestrogens has a synergistic effect mediated by p21WAF1 in estrogen receptor-positive breast cancer.

    PubMed

    Maynadier, Marie; Basile, Ilaria; Gallud, Audrey; Gary-Bobo, Magali; Garcia, Marcel

    2016-08-01

    Although antiestrogens significantly improve the survival of patients with ER-positive breast cancer, therapeutic resistance remains a major limitation. The combinatorial use of antiestrogen with other therapies was proposed to increase their efficiency and more importantly, to prevent or delay the resistance phenomenon. In the present study, we addressed their combined effects with proteasome inhibitors (PIs). The effects of antiestrogens (hydroxyl-tamoxifen, raloxifen and fulvestrant) currently used in endocrine therapy were tested in combination with PIs, bortezomib or MG132, on the growth of three ER-positive breast cancer cell lines and in two cellular models of acquired antiestrogen resistance. When compared to single treatments, these combined treatments were significantly more effective in preventing the growth of the cell lines. The regulation of key cell cycle proteins, the cyclin-dependent kinase inhibitors, p21WAF1 and p27KIP1, were also studied. Bortezomib and MG132 drastically increased p21WAF1 expression through elevation of its mRNA concentration. Notably, p27KIP1 regulation was quite different from that of p21WAF1. Furthermore, the effect of bortezomib in combination with antiestrogen was evaluated on antiestrogen-resistant cell lines. The growth of two antiestrogen-resistant cell lines appeared responsive to proteasome inhibition and was strongly decreased by a combined therapy with an antiestrogen. Collectively, these findings provide new perspectives for the use of PIs in combination with endocrine therapies for breast cancer and possibly to overcome acquired hormonal resistance. PMID:27373750

  6. Ovarian cancer has frequent loss of heterozygosity at chromosome 12p12.3-13.1 (region of TEL and Kip1 loci) and chromosome 12q23-ter: evidence for two new tumour-suppressor genes.

    PubMed Central

    Hatta, Y.; Takeuchi, S.; Yokota, J.; Koeffler, H. P.

    1997-01-01

    Identification of the key genetic alterations leading to ovarian cancer is in its infancy. Polymerase chain reaction (PCR)-based analysis of loss of heterozygosity (LOH) is a powerful method for detecting regions of altered tumour-suppressor genes. Focusing on chromosome 12, we examined 23 ovarian cancer samples for LOH using 31 highly polymorphic microsatellite markers and found the chromosomal localization of two putative tumour-suppressor genes. Two commonly deleted regions were 12p12.3-13.1 in 6/23 (26%) and 12q23-ter in 7/23 (30%) samples. LOH on chromosome 12 was more common in late-stage ovarian carcinomas. The region of LOH at 12p12.3-13.1 includes the genes that code for the ETS-family transcriptional factor, known as TEL, and the cyclin-dependent kinase inhibitor, known as p27Kip1. Mutational analysis of both TEL and p27Kip1 using single-strand conformation polymorphism (SSCP) showed no abnormalities, suggesting that the altered gene in this region is neither of these genes. Taken together, our data suggest that new tumour-suppressor genes in the region of chromosomes 12p12.3-13.1 and 12q23-ter may be involved in the development of ovarian cancer. Images Figure 1 Figure 2 Figure 4 PMID:9155043

  7. Combined inhibition of EZH2 and histone deacetylases as a potential epigenetic therapy for non-small-cell lung cancer cells.

    PubMed

    Takashina, Taichi; Kinoshita, Ichiro; Kikuchi, Junko; Shimizu, Yasushi; Sakakibara-Konishi, Jun; Oizumi, Satoshi; Nishimura, Masaharu; Dosaka-Akita, Hirotoshi

    2016-07-01

    Recent discoveries have revealed that human cancer involves aberrant epigenetic alterations. We and others have previously shown that the histone methyltransferase EZH2, the catalytic subunit of polycomb repressive complex 2 (PRC2), is frequently overexpressed in non-small-cell lung cancer (NSCLC) and that an EZH2 inhibitor, 3-deazaneplanocin A, inhibits the proliferation of NSCLC cells. Transcriptional silencing by EZH2 was recently shown to be required for the activity of histone deacetylases (HDACs) that interact with another PRC2 protein, EED. To develop a more effective epigenetic therapy for NSCLC, we determined the effects of co-treatment with 3-deazaneplanocin A and the HDAC inhibitor vorinostat (SAHA) in NSCLC cells. The co-treatment synergistically suppressed the proliferation of all tested NSCLC cell lines, regardless of their epidermal growth factor receptor (EGFR) status. The synergistic effect was associated with slightly decreased histone H3 lysine 27 trimethylation, modestly increased histone acetylation, and the depletion of EZH2 and other PRC2 proteins. The co-treatment resulted in an accumulation of p27Kip1, decrease in cyclin A, and increased apoptotic fraction in an additive/synergistic manner. Interestingly, the co-treatment strongly suppressed EGFR signaling, not only in EGFR-wild-type NSCLC cells, but also in EGFR-mutant cells, mainly through dephosphorylation of EGFR. Furthermore, the co-treatment suppressed the in vivo tumor growth of EGFR-mutant, EGFR-tyrosine kinase-resistant H1975 cells more effectively than did each agent alone, without visible toxicity. These results suggest that the combined pharmacological targeting of EZH2 and HDACs may provide more effective epigenetic therapeutics for NSCLC. PMID:27116120

  8. Male breast cancer is not congruent with the female disease.

    PubMed

    Fentiman, Ian S

    2016-05-01

    It has become customary to extrapolate from the results of treatment trials for female breastcancer and apply them to males with the disease. In the absence of results from national and international randomised trials for male breast cancer (MBC) this appears superficially to be an appropriate response. Closer examination of available data reveals that aspects of the aetiology and treatment of MBC do not fit the simplistic model that men usually have endocrine sensitive tumours which behave like those in postmenopausal women. Most females and males with breast cancer have none of the recognised risk factors, indicating the gaps in our knowledge of the epidemiology of this disease. Several studies have compared epidemiological risk factors for MBC and female breast cancer (FBC) but many have been blighted by small numbers. In comparison with FBC there is a larger proportion of BRCA2 tumours, (occurring in 10% of MBC), and underrepresentation of BRCA1 tumours (found in only 1%), suggesting significant differences in the genetic aetiology of MBC and FBC. Genome-wide association studies in FBC reported single nucleotide polymorphisms (SNPs) in 12 novel independent loci were consistently associated with disease but for MBC 2 SNPs had a significantly increased risk. Molecular profiles of matched cancers in males and females showed a gender-associated modulation of major processes including energy metabolism, regulation of translation, matrix remodelling and immune recruitment. Immunohistochemistry for kinase inhibitor proteins (KIPs) p27Kip1 and p21Waf1 indicate a significant difference in the immunostaining of tumours from male patients compared with females. MBC is almost always estrogen receptor positive (ER+ve) and so systemic treatment is usually endocrine. With evidence in FBC that aromatase inhibitors are more effective than tamoxifen in the postmenopausal it was seemingly logical that the same would be true for MBC. Results however suggest less efficacy with AIs

  9. CRM1/Ran-mediated nuclear export of p27(Kip1) involves a nuclear export signal and links p27 export and proteolysis.

    PubMed

    Connor, Michael K; Kotchetkov, Rouslan; Cariou, Sandrine; Resch, Ansgar; Lupetti, Rafaella; Beniston, Richard G; Melchior, Frauke; Hengst, Ludger; Slingerland, Joyce M

    2003-01-01

    We show that p27 localization is cell cycle regulated and we suggest that active CRM1/RanGTP-mediated nuclear export of p27 may be linked to cytoplasmic p27 proteolysis in early G1. p27 is nuclear in G0 and early G1 and appears transiently in the cytoplasm at the G1/S transition. Association of p27 with the exportin CRM1 was minimal in G0 and increased markedly during G1-to-S phase progression. Proteasome inhibition in mid-G1 did not impair nuclear import of p27, but led to accumulation of p27 in the cytoplasm, suggesting that export precedes degradation for at least part of the cellular p27 pool. p27-CRM1 binding and nuclear export were inhibited by S10A mutation but not by T187A mutation. A putative nuclear export sequence in p27 is identified whose mutation reduced p27-CRM1 interaction, nuclear export, and p27 degradation. Leptomycin B (LMB) did not inhibit p27-CRM1 binding, nor did it prevent p27 export in vitro or in heterokaryon assays. Prebinding of CRM1 to the HIV-1 Rev nuclear export sequence did not inhibit p27-CRM1 interaction, suggesting that p27 binds CRM1 at a non-LMB-sensitive motif. LMB increased total cellular p27 and may do so indirectly, through effects on other p27 regulatory proteins. These data suggest a model in which p27 undergoes active, CRM1-dependent nuclear export and cytoplasmic degradation in early G1. This would permit the incremental activation of cyclin E-Cdk2 leading to cyclin E-Cdk2-mediated T187 phosphorylation and p27 proteolysis in late G1 and S phase. PMID:12529437

  10. Novel analogs targeting histone deacetylase suppress aggressive thyroid cancer cell growth and induce re-differentiation.

    PubMed

    Jang, S; Yu, X-M; Odorico, S; Clark, M; Jaskula-Sztul, R; Schienebeck, C M; Kupcho, K R; Harrison, A D; Winston-McPherson, G N; Tang, W; Chen, H

    2015-08-01

    To develop novel therapies for aggressive thyroid cancers, we have synthesized a collection of histone deacetylase (HDAC) inhibitor analogs named AB1 to AB13, which have different linkers between a metal chelating group and a hydrophobic cap. The purpose of this study was to screen out the most effective compounds and evaluate the therapeutic efficacy. AB2, AB3 and AB10 demonstrated the lowest half-maximal inhibitory concentration (IC50) values in one metastatic follicular and two anaplastic thyroid cancer cell lines. Treatment with each of the three ABs resulted in an increase in apoptosis markers, including cleaved poly adenosine diphosphate ribose polymerase (PARP) and cleaved caspase 3. Additionally, the expression of cell-cycle regulatory proteins p21(WAF1) and p27(Kip1) increased with the treatment of ABs while cyclin D1 decreased. Furthermore, AB2, AB3 and AB10 were able to induce thyrocyte-specific genes in the three thyroid cancer cell lines indicated by increased expression levels of sodium iodide symporter, paired box gene 8, thyroid transcription factor 1 (TTF1), TTF2 and thyroid-stimulating hormone receptors. AB2, AB3 and AB10 suppress thyroid cancer cell growth via cell-cycle arrest and apoptosis. They also induce cell re-differentiation, which could make aggressive cancer cells more susceptible to radioactive iodine therapy. PMID:26251030

  11. Fangchinoline induces G1 arrest in breast cancer cells through cell-cycle regulation.

    PubMed

    Xing, Zhibo; Zhang, Youxue; Zhang, Xianyu; Yang, Yanmei; Ma, Yuyan; Pang, Da

    2013-12-01

    Fangchinoline, an alkaloid derived from the dry roots of Stephaniae tetrandrine S. Moore (Menispermaceae), has been shown to possess cytotoxic, anti-inflammatory, and antioxidant properties. In this study, we used Fangchinoline to inhibit breast cancer cell proliferation and to investigate its underlying molecular mechanisms. Human breast cancer cell lines, MCF-7 and MDA-MB-231, were both used in this study. We found that Fangchinoline significantly decreased cell proliferation in a dose-dependent manner and induced G1-phase arrest in both cell lines. In addition, upon analysis of expression of cell cycle-related proteins, we found that Fangchinoline reduced expression of cyclin D1, cyclin D3, and cyclin E, and increased expression of the cyclin-dependent kinase (CDK) inhibitors, p21/WAF1, and p27/KIP1. Moreover, Fangchinoline also inhibited the kinase activities of CDK2, CDK4, and CDK6. These results suggest that Fangchinoline can inhibit human breast cancer cell proliferation and thus may have potential applications in cancer therapy. PMID:23401195

  12. Casticin induces ovarian cancer cell apoptosis by repressing FoxM1 through the activation of FOXO3a

    PubMed Central

    JIANG, LING; CAO, XIAO-CHENG; CAO, JIAN-GUO; LIU, FEI; QUAN, MEI-FANG; SHENG, XI-FENG; REN, KAI-QUN

    2013-01-01

    Casticin, a polymethoxyflavone, is reported to have anticancer activities. The aim of the present study was to examine the molecular mechanisms by which casticin induces apoptosis in ovarian cancer cells. The human ovarian cancer cell lines SKOV3 and A2780 were cultured in vitro. Various molecular techniques, including histone/DNA enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), western blot analysis and gene transfection, were used to assess the expression of FOXO3a and forkhead box protein M1 (FoxM1) in casticin-treated ovarian cancer cell lines. Casticin-induced apoptotic cell death was accompanied by the activation of transcription factor FOXO3a, with a concomitant decrease in the expression levels of FoxM1 and its downstream target factors, namely survivin and polo-like kinase 1 (PLK1), and an increase in p27KIP1. A small inhibitory RNA (siRNA) knockout of FoxM1 potentiated casticin-induced apoptosis in ovarian cancer cells. Silencing FOXO3a expression using siRNA increased FoxM1 expression levels and clearly attenuated the induction of apoptosis by casticin treatment. These results show that casticin-induced apoptosis in ovarian cancer may be caused by the activation of FOXO3a, leading to FoxM1 inhibition. PMID:23761826

  13. Euphol from Euphorbia tirucalli selectively inhibits human gastric cancer cell growth through the induction of ERK1/2-mediated apoptosis.

    PubMed

    Lin, Ming-Wei; Lin, An-Shen; Wu, Deng-Chyang; Wang, Sophie S W; Chang, Fang-Rong; Wu, Yang-Chang; Huang, Yaw-Bin

    2012-12-01

    Gastric cancer is one of the most common malignancies worldwide, and the main cause of cancer-related death in Asia. The present study assessed the anticancer effects of euphol, a triterpene alcohol with anti-inflammatory and antiviral activities on human gastric cancer cells. Euphol showed higher cytotoxicity activity against human gastric CS12 cancer cells than against noncancer CSN cells. In addition, it up-regulated the pro-apoptotic protein BAX and down-regulated the prosurvival protein Bcl-2, causing mitochondrial dysfunction, possibly by caspase-3 activation. The anti-proliferative effects of euphol were associated with the increased p27(kip1) levels and decreased cyclin B1 levels. Inhibition of ERK1/2 activation by PD98059 reversed euphol-induced pro-apoptotic protein expression and cell death. Taken together, these findings suggest that euphol selectively induced gastric cancer cells apoptosis by modulation of ERK signaling, and could thus be of value for cancer therapy. PMID:22634261

  14. Inhibitory effect of mimosine on proliferation of human lung cancer cells is mediated by multiple mechanisms.

    PubMed

    Chang, H C; Lee, T H; Chuang, L Y; Yen, M H; Hung, W C

    1999-10-18

    The plant amino acid mimosine has been reported to block cell cycle progression in the late G1 phase. A recent study showed that mimosine might induce growth arrest by activating the expression of p21CIP1, a cyclin-dependent kinase inhibitor (CDKI), and by inhibiting the activity of cyclin E-associated kinases in human breast cancer cells. However, mimosine at higher concentrations also blocked proliferation of p21-/- cells by unknown mechanisms. In this study, we investigated the effect of mimosine on the expression of cyclins and CDKIs in human lung cancer cells. We found that mimosine specifically inhibited cyclin D1 expression in H226 cells. The expression of another G1 cyclin, cyclin E, was not regulated by mimosine in all lung cancer cell lines examined. Moreover, mimosine induced p21CIP1 expression in H226 and H358 cells, while it activated p27KIP1 expression in H322 cells. However, mimosine does not affect transcription of these genes directly because significant changes in cyclin D1 or CDKI expression were observed at 12-24 h after drug addition. Our results indicate that mimosine may block cell proliferation by multiple mechanisms and this amino acid is a useful agent for the study of cell cycle control. PMID:10530763

  15. Pterostilbine, an active component of blueberries, sensitizes colon cancer cells to 5-fluorouracil cytotoxicity.

    PubMed

    Tolba, Mai F; Abdel-Rahman, Sherif Z

    2015-01-01

    Although colorectal cancer (CRC) treatment with 5-fluorouracil (5-FU) is the first line of therapy for this debilitating disease, treatment effectiveness is often hampered by the development of drug resistance and toxicity at high doses. ER-β can play an important role in CRC development and possibly in its response to therapy. Pterostilbene (PT) possesses antioxidant and anticancer effects that are mediated by ER-β. In the current study, we test the hypothesis that PT sensitizes colon cancer cells to 5-FU and we examine the underlying mechanism(s) by which PT exerts its cytotoxic effects in CRC cells. Our data indicate that PT exhibited a more potent cytotoxic effect in Caco-2 compared to HCT-116 cells. PT/5-FU co-treatment was more effective in Caco-2 cells. Our data indicate that ER-β is expressed at higher levels in Caco-2 cells and its levels are further boosted with PT treatment. PT significantly suppressed Akt and ERK phosphorylations, and enhanced FOXO-1 and p27(kip1) levels in Caco-2 cells. PT also induced a significant increase in Caco-2 cells at pre-G phase coupled with increased Bax/Bcl-2 ratio and PARP cleavage. These results provide a rationale for novel combination treatment strategies, especially for patients with 5-FU-resistant tumors expressing ER-β protein. PMID:26472352

  16. Pterostilbine, an active component of blueberries, sensitizes colon cancer cells to 5-fluorouracil cytotoxicity

    PubMed Central

    Tolba, Mai F.; Abdel-Rahman, Sherif Z.

    2015-01-01

    Although colorectal cancer (CRC) treatment with 5-fluorouracil (5-FU) is the first line of therapy for this debilitating disease, treatment effectiveness is often hampered by the development of drug resistance and toxicity at high doses. ER-β can play an important role in CRC development and possibly in its response to therapy. Pterostilbene (PT) possesses antioxidant and anticancer effects that are mediated by ER-β. In the current study, we test the hypothesis that PT sensitizes colon cancer cells to 5-FU and we examine the underlying mechanism(s) by which PT exerts its cytotoxic effects in CRC cells. Our data indicate that PT exhibited a more potent cytotoxic effect in Caco-2 compared to HCT-116 cells. PT/5-FU co-treatment was more effective in Caco-2 cells. Our data indicate that ER-β is expressed at higher levels in Caco-2 cells and its levels are further boosted with PT treatment. PT significantly suppressed Akt and ERK phosphorylations, and enhanced FOXO-1 and p27kip1 levels in Caco-2 cells. PT also induced a significant increase in Caco-2 cells at pre-G phase coupled with increased Bax/Bcl-2 ratio and PARP cleavage. These results provide a rationale for novel combination treatment strategies, especially for patients with 5-FU-resistant tumors expressing ER-β protein. PMID:26472352

  17. Synthesis, Characterization and Anti-Cancer Activity of Hydrazide Derivatives Incorporating a Quinoline Moiety.

    PubMed

    Bingul, Murat; Tan, Owen; Gardner, Christopher R; Sutton, Selina K; Arndt, Greg M; Marshall, Glenn M; Cheung, Belamy B; Kumar, Naresh; Black, David StC

    2016-01-01

    Identification of the novel (E)-N'-((2-chloro-7-methoxyquinolin-3-yl)methylene)-3-(phenylthio)propanehydrazide scaffold 18 has led to the development of a new series of biologically active hydrazide compounds. The parent compound 18 and new quinoline derivatives 19-26 were prepared from the corresponding quinoline hydrazones and substituted carboxylic acids using EDC-mediated peptide coupling reactions. Further modification of the parent compound 18 was achieved by replacement of the quinoline moiety with other aromatic systems. All the newly synthesized compounds were evaluated for their anti-cancer activity against the SH-SY5Y and Kelly neuroblastoma cell lines, as well as the MDA-MB-231 and MCF-7 breast adenocarcinoma cell lines. Analogues 19 and 22 significantly reduced the cell viability of neuroblastoma cancer cells with micromolar potency and significant selectivity over normal cells. The quinoline hydrazide 22 also induced G₁ cell cycle arrest, as well as upregulation of the p27(kip1) cell cycle regulating protein. PMID:27428941

  18. Induction of cell cycle arrest in prostate cancer cells by the dietary compound isoliquiritigenin.

    PubMed

    Lee, Yeo Myeong; Lim, Do Young; Choi, Hyun Ju; Jung, Jae In; Chung, Won-Yoon; Park, Jung Han Yoon

    2009-02-01

    Isoliquiritigenin (ISL), a flavonoid chalcone that is present in licorice, shallot, and bean sprouts, is known to have antitumorigenic activities. The present study examined whether ISL alters prostate cancer cell cycle progression. DU145 human and MatLyLu (MLL) rat prostate cancer cells were cultured with various concentrations of ISL. In both DU145 and MLL cells treated with ISL, the percentage of cells in the G1 phase increased, and the incorporation of [(3)H]thymidine decreased. ISL decreased the protein levels of cyclin D1, cyclin E, and cyclin-dependent kinase (CDK) 4, whereas cyclin A and CDK2 expressions were unaltered in cells treated with ISL. The expression of the CDK inhibitor p27(KIP1) was increased in cells treated with 20 micromol/L ISL. In addition, treatment of cells with 20 micromol/L ISL for 24 hours led to G2/M cell cycle arrest. Cell division control (CDC) 2 protein levels remained unchanged. The protein levels of phospho-CDC2 (Tyr15) and cyclin B1 were increased, and the CDC25C level was decreased by ISL dose-dependently. We demonstrate that ISL promotes cell cycle arrest in DU145 and MLL cells, thereby providing insights into the mechanisms underlying its antitumorigenic activities. PMID:19298190

  19. Testing the importance of p27 degradation by the SCFskp2 pathway in murine models of lung and colon cancer.

    PubMed

    Timmerbeul, Inke; Garrett-Engele, Carrie M; Kossatz, Uta; Chen, Xueyan; Firpo, Eduardo; Grünwald, Viktor; Kamino, Kenji; Wilkens, Ludwig; Lehmann, Ulrich; Buer, Jan; Geffers, Robert; Kubicka, Stefan; Manns, Michael P; Porter, Peggy L; Roberts, James M; Malek, Nisar P

    2006-09-19

    Decreased expression of the CDK inhibitor p27kip1 in human tumors directly correlates with increased resistance to chemotherapies, increased rates of metastasis, and an overall increased rate of patient mortality. It is thought that decreased p27 expression in tumors is caused by increased proteasomal turnover, in particular activation of the pathway governed by the SCFskp2 E3 ubiquitin protein ligase. We have directly tested the importance of the SCFskp-mediated degradation of p27 in tumorigenesis by analyzing the tumor susceptibility of mice that express a form of p27 that cannot be ubiquitinated and degraded by this pathway (p27T187A). In mouse models of both lung and colon cancer down-regulation of p27 promotes tumorigenesis. However, we found that preventing p27 degradation by the SCFskp2 pathway had no impact on tumor incidence or overall survival in either tumor model. Our study unveiled a previously unrecognized role for the control of p27 mRNA abundance in the development of non-small cell lung cancers. In the colon cancer model, the frequency of intestinal adenomas was similarly unaffected by the p27T187A mutation, but, unexpectedly, we found that it inhibited progression of intestinal adenomas to carcinomas. These studies may guide the choice of clinical settings in which pharmacologic inhibitors of the Skp2 pathway might be of therapeutic value. PMID:16966613

  20. The enhancer of zeste homolog 2 (EZH2), a potential therapeutic target, is regulated by miR-101 in renal cancer cells

    SciTech Connect

    Sakurai, Toshihiko; Bilim, Vladimir N.; Ugolkov, Andrey V.; Yuuki, Kaori; Tsukigi, Masaaki; Motoyama, Teiichi; Tomita, Yoshihiko

    2012-06-15

    Highlights: Black-Right-Pointing-Pointer EZH2 is overexpressed in the nuclei of renal cancer cells. Black-Right-Pointing-Pointer Nuclear EZH2 is associated with advanced stage and worse survival of RCC patients. Black-Right-Pointing-Pointer EZH2 is negatively regulated by miR-101 in renal cancer cells. Black-Right-Pointing-Pointer Depletion of EZH2 leads to re-expression of p27Kip1. Black-Right-Pointing-Pointer Reintroduction of miR-101 results in suppression of cell proliferation. -- Abstract: We investigated a prognostic significance and the mechanism of aberrant nuclear expression of EZH2, a histone methyltransferase, in human renal cell carcinoma (RCC). We found nuclear EZH2 in 48 of 100 RCCs and it was significantly correlated with worse survival in RCC patients. We detected a decreased expression of miR-101 in 15 of 54 RCCs. We found that re-expression of miR-101 resulted in EZH2 depletion and decreased renal cancer cell proliferation. Our results show nuclear EZH2 as a prognostic marker of worse survival in human RCC, and identify miR-101 as a negative regulator of EZH2 expression and renal cancer cell proliferation.

  1. Biological mechanisms of action of novel C-10 non-acetal trioxane dimers in prostate cancer cell lines.

    PubMed

    Alagbala, Adebusola A; McRiner, Andrew J; Borstnik, Kristina; Labonte, Tanzina; Chang, Wonsuk; D'Angelo, John G; Posner, Gary H; Foster, Barbara A

    2006-12-28

    The mechanisms of action of three C-10 non-acetal trioxane dimers (TDs) were examined in human (LNCaP) and mouse (TRAMP-C1A and -C2H) prostate cancer cell lines. 1 (AJM3/23), 2 (GHP-TM-III-07w), and 3 (GHP-KB-06) inhibited cell growth with 3 being the most potent in C1A (GI50 = 18.0 nM), C2H (GI50 = 17.0 nM), and LNCaP (GI50 = 17.9 nM) cells. In comparison to a standard cytotoxic agent such as doxorubicin (GI50 = 45.3 nM), 3 (GI50 = 17.9 nM) inhibited LNCaP cell growth more potently. TDs induced G0/G1 cell cycle arrest in LNCaP cells and decreased cells in the S phase. These changes correlated with modulation of G1 phase cell cycle proteins including decreased cyclin D1, cyclin E, and cdk2 and increased p21waf1 and p27Kip1. TDs also promoted apoptosis in LNCaP cells with increased expression of proapoptotic bax. These results demonstrate that TDs are potentially useful agents that warrant further preclinical development for treatment of prostate cancer. PMID:17181166

  2. TLE4 promotes colorectal cancer progression through activation of JNK/c-Jun signaling pathway

    PubMed Central

    Deng, Dan-Ling; Cai, Juan-Juan; Xiao, Zhi-Yuan; He, Liu-Qing; Jiao, Hong-Li; Ye, Ya-Ping; Yang, Run-Wei; Li, Ting-Ting; Liang, Li; Liao, Wen-Ting; Ding, Yan-Qing

    2016-01-01

    The Groucho transcriptional co-repressor TLE4 protein has been shown to be a tumor suppressor in a subset of acute myeloid leukemia. However, little is known about its role in development and progression of solid tumor. In this study, we found that the expression of TLE4 in colorectal cancer (CRC) tissues was significantly higher than that in their matched adjacent intestine epithelial tissues. In addition, high expression of TLE4 was significantly correlated with advanced Dukes stage, lymph node metastasis and poor prognosis of CRC. Moreover, enforced expression of TLE4 in CRC cell lines significantly enhanced proliferation, invasion and tumor growth. On the contrary, knock down of TLE4 repressed cell proliferation, invasion and tumor growth. Furthermore, our study exhibited that the TLE4 promoted cell proliferation and invasion partially via activation of JNK-c-Jun pathway and subsequently increased cyclinD1 and decreased P27Kip1 expression. In conclusion, these results suggested that TLE4, a potential prognostic biomarker for CRC, plays an important role in the development and progression of human CRC. PMID:26701208

  3. Overexpression of the human ZNF300 gene enhances growth and metastasis of cancer cells through activating NF-kB pathway

    PubMed Central

    Wang, Tao; Wang, Xian-guo; Xu, Jun-hua; Wu, Xiang-Peng; Qiu, Hong-ling; Yi, Hong; Li, Wen-Xin

    2012-01-01

    Abstract Zinc finger proteins (ZNF) play important roles in various physiological processes. Here we report that ZNF300, a novel zinc finger protein, identified specifically in humans, promotes tumour development by modulating the NF-κB pathway. Inflammatory factors were found to induce ZNF300 expression in HeLa cell line, and ZNF300 expression further enhanced NF-κB signalling by activating TRAF2 and physically interacting with IKKβ. Furthermore, ZNF300 overexpression increased ERK1/2 phosphorylation and the expression of c-myc, IL-6, and IL-8 but decreased the expression of p21waf-1 and p27Kip1; whose down-regulation led to the opposite effect. Most importantly, ZNF300 overexpression stimulated cancer cell proliferation in vitro and significantly enhanced tumour development and metastasis in mouse xenograft model, while knocking down ZNF300 led to the opposite effects. We have identified a novel function for ZNF300 in tumour development that may uniquely link inflammation and NF-κB to tumourigenesis in humans but not in mice. PMID:21777376

  4. SIAH ubiquitin ligases regulate breast cancer cell migration and invasion independent of the oxygen status

    PubMed Central

    Adam, M Gordian; Matt, Sonja; Christian, Sven; Hess-Stumpp, Holger; Haegebarth, Andrea; Hofmann, Thomas G; Algire, Carolyn

    2015-01-01

    Seven-in-absentia homolog (SIAH) proteins are evolutionary conserved RING type E3 ubiquitin ligases responsible for the degradation of key molecules regulating DNA damage response, hypoxic adaptation, apoptosis, angiogenesis, and cell proliferation. Many studies suggest a tumorigenic role for SIAH2. In breast cancer patients SIAH2 expression levels correlate with cancer aggressiveness and overall patient survival. In addition, SIAH inhibition reduced metastasis in melanoma. The role of SIAH1 in breast cancer is still ambiguous; both tumorigenic and tumor suppressive functions have been reported. Other studies categorized SIAH ligases as either pro- or antimigratory, while the significance for metastasis is largely unknown. Here, we re-evaluated the effects of SIAH1 and SIAH2 depletion in breast cancer cell lines, focusing on migration and invasion. We successfully knocked down SIAH1 and SIAH2 in several breast cancer cell lines. In luminal type MCF7 cells, this led to stabilization of the SIAH substrate Prolyl Hydroxylase Domain protein 3 (PHD3) and reduced Hypoxia-Inducible Factor 1α (HIF1α) protein levels. Both the knockdown of SIAH1 or SIAH2 led to increased apoptosis and reduced proliferation, with comparable effects. These results point to a tumor promoting role for SIAH1 in breast cancer similar to SIAH2. In addition, depletion of SIAH1 or SIAH2 also led to decreased cell migration and invasion in breast cancer cells. SIAH knockdown also controlled microtubule dynamics by markedly decreasing the protein levels of stathmin, most likely via p27Kip1. Collectively, these results suggest that both SIAH ligases promote a migratory cancer cell phenotype and could contribute to metastasis in breast cancer. PMID:26654769

  5. Overexpressed CISD2 has prognostic value in human gastric cancer and promotes gastric cancer cell proliferation and tumorigenesis via AKT signaling pathway

    PubMed Central

    Wang, Lan; Ouyang, Fei; Liu, Xiaobo; Wu, Shu; Wu, Hong-mei; Xu, Yuandong; Wang, Bin; Zhu, Jinrong; Xu, Xuehu; Zhang, Liang

    2016-01-01

    CDGSH iron sulfur domain 2 (CISD2) is localized in the outer mitochondrial membrane and mediates mitochondrial integrity and lifespan in mammals, but its role in cancer is unknown. In the current study, we reported that CISD2 mRNA and protein expression levels were significantly upregulated in gastric cancer cells compared to normal gastric epithelial cells (P < 0.001). Immunohistochemical analysis of 261 paraffin-embedded archived gastric cancer tissues showed that high CISD2 expression was significantly associated with clinical stage, TNM classifications, venous invasion and lymphatic invasion. Univariate and multivariate analysis indicated that high CISD2 expression was an independent prognostic factor for poorer overall survival in the entire cohort. Overexpressing CISD2 promoted, while silencing CISD2 inhibited, the proliferation of gastric cancer cells. Furthermore, we found that silencing endogenous CISD2 also significantly inhibited the proliferation and tumorigenicity of MGC-803 and SGC-7901 cells not only in vitro but also in vivo in NOD/SCID mice (P < 0.05). Furthermore, we found that CISD2 affected cell proliferation and tumorigenicity of gastric cancer cells through mediating the G1-to-S phase transition. Moreover, we demonstrated that the pro-proliferative effect of CISD2 on gastric cancer cells was associated with downregulation of cyclin-dependent kinase inhibitor p21Cip1 and p27Kip1, and activation of AKT signaling. The findings of this study indicate that CISD2 may promote proliferation and tumorigenicity, potentially representing a novel prognostic marker for overall survival in gastric cancer. PMID:26565812

  6. Overexpressed CISD2 has prognostic value in human gastric cancer and promotes gastric cancer cell proliferation and tumorigenesis via AKT signaling pathway.

    PubMed

    Wang, Lan; Ouyang, Fei; Liu, Xiaobo; Wu, Shu; Wu, Hong-Mei; Xu, Yuandong; Wang, Bin; Zhu, Jinrong; Xu, Xuehu; Zhang, Liang

    2016-01-26

    CDGSH iron sulfur domain 2 (CISD2) is localized in the outer mitochondrial membrane and mediates mitochondrial integrity and lifespan in mammals, but its role in cancer is unknown. In the current study, we reported that CISD2 mRNA and protein expression levels were significantly upregulated in gastric cancer cells compared to normal gastric epithelial cells (P < 0.001). Immunohistochemical analysis of 261 paraffin-embedded archived gastric cancer tissues showed that high CISD2 expression was significantly associated with clinical stage, TNM classifications, venous invasion and lymphatic invasion. Univariate and multivariate analysis indicated that high CISD2 expression was an independent prognostic factor for poorer overall survival in the entire cohort. Overexpressing CISD2 promoted, while silencing CISD2 inhibited, the proliferation of gastric cancer cells. Furthermore, we found that silencing endogenous CISD2 also significantly inhibited the proliferation and tumorigenicity of MGC-803 and SGC-7901 cells not only in vitro but also in vivo in NOD/SCID mice (P < 0.05). Furthermore, we found that CISD2 affected cell proliferation and tumorigenicity of gastric cancer cells through mediating the G1-to-S phase transition. Moreover, we demonstrated that the pro-proliferative effect of CISD2 on gastric cancer cells was associated with downregulation of cyclin-dependent kinase inhibitor p21Cip1 and p27Kip1, and activation of AKT signaling. The findings of this study indicate that CISD2 may promote proliferation and tumorigenicity, potentially representing a novel prognostic marker for overall survival in gastric cancer. PMID:26565812

  7. Nuclear export of proteins and drug resistance in cancer

    PubMed Central

    Turner, Joel G.; Dawson, Jana; Sullivan, Daniel M.

    2015-01-01

    The intracellular location of a protein is crucial to its normal functioning in a cell. Cancer cells utilize the normal processes of nuclear-cytoplasmic transport through the nuclear pore complex of a cell to effectively evade anti-neoplastic mechanisms. CRM1-mediated export is increased in various cancers. Proteins that are exported in cancer include tumor-suppressive proteins such as retinoblastoma, APC, p53, BRAC1, FOXO proteins, INI1/hSNF5, galectin-3, Bok, nucleophosmin, RASSF2, Merlin, p21CIP, p27KIP1, N-WASP/FAK, estradiol receptor and Tob, drug targets topoisomerase I and IIα and BCR-ABL, and the molecular chaperone protein Hsp90. Here, we review in detail the current processes and known structures involved in the export of a protein through the nuclear pore complex. We also discuss the export receptor molecule CRM1 and its binding to the leucine-rich nuclear export signal of the cargo protein and the formation of a nuclear export trimer with RanGTP. The therapeutic potential of various CRM1 inhibitors will be addressed, including leptomycin B, ratjadone, KOS-2464, and specific small molecule inhibitors of CRM1, N-azolylacrylate analogs, FOXO export inhibitors, valtrate, acetoxychavicol acetate, CBS9106, and SINE inhibitors. We will also discuss examples of how drug resistance may be reversed by targeting the exported proteins topoisomerase IIα, BCR-ABL, and galectin-3. As effective and less toxic CRM1 export inhibitors become available, they may be used as both single agents and in combination with current chemotherapeutic drugs. We believe that the future development of low-toxicity, small-molecule CRM1 inhibitors may provide a new approach to treating cancer. PMID:22209898

  8. Down-regulation of microRNA-135b inhibited growth of cervical cancer cells by targeting FOXO1

    PubMed Central

    Xu, Yue; Zhao, Shuhua; Cui, Manhua; Wang, Qiang

    2015-01-01

    More and more evidence has confirmed that dysregulation of microRNAs (miRNAs) can conduce to the progression of human cancers. Previous studied have shown that dysregulation of miR-135b is in varieties of tumors. However, the roles of miR-135b in cervical cancer remain unknown. Therefore, our aim of this study was to explore the biological function and molecular mechanism of miR-135b in cervical cancer cell lines, discussing whether it could be a therapeutic biomarker of cervical cancer in the future. The MTT assay and ELISA-Brdu assay were used to assess cell proliferation. Cell cycle was detected by flow cytometry. Real-time quantitative polymerase chain reaction (PCR) and Western blot analyses were used to detect expressions of cyclin D1, p21, p27 and FOXO1. In our study, we found that miR-135b is up-regulated in cervical cancer cell lines. Down-regulation of miR-135b evidently inhibited proliferation and arrested cell cycle in cervical cancer cells. Bioinformatics analysis predicted that the FOXO1 was a potential target gene of miR-135b. Besides, miR-135b inhibition significantly increased expressions of the cyclin-dependent kinase inhibitors, p21/CIP1 and p27/KIP1, and decreased expression of cyclin D1. However, the high level of miR-135b was associated with increased expression of FOXO1 in cervical cancer cells. Further study by luciferase reporter assay demonstrated that miR-135b could directly target FOXO1. Down-regulation of FOXO1 in cervical cancer cells transfected with miR-135b inhibitor partially reversed its inhibitory effects. In conclusion, down-regulation of miR-135b inhibited cell growth in cervical cancer cells by up-regulation of FOXO1. PMID:26617737

  9. Dasatinib Induces Autophagic Cell Death in Human Ovarian Cancer*

    PubMed Central

    Le, Xiao-Feng; Mao, Weiqun; Lu, Zhen; Carter, Bing Z.; Bast, Robert C.

    2010-01-01

    BACKGROUND Dasatinib, an inhibitor of Src/Abl family kinases, can inhibit tumor growth of a number of solid tumors. However, the effect and mechanism of action of dasatinib in human ovarian cancer cells remains unknown. METHODS Dasatinib-induced autophagy was determined by acridine orange staining, punctate localization of GFP-LC3, LC3 protein blotting and electron microscopy. Significance of Beclin-1, AKT and Bcl-2 in dasatinib-induced autophagy and growth inhibition was assayed by small interfering RNA silencing and/or overexpression of gene of interest. RESULTS Dasatinib inhibited cell growth by inducing little apoptosis, but substantial autophagy in SKOv3 and HEY ovarian cancer cells. In vivo studies showed dasatinib inhibited tumor growth and induced both autophagy and apoptosis in a HEY xenograft model. Knockdown of Beclin 1 and Atg12 expression with their respective siRNAs diminished dasatinib-induced autophagy, whereas knockdown of p27Kip1 with specific siRNAs did not. shRNA knockdown of Beclin-1 expression reduced dasatinib-induced autophagy and growth inhibition. Dasatinib reduced the phosphorylation of AKT, mTOR, p70S6K and S6 kinase expression. Constitutive expression of AKT1 and AKT2 inhibited dasatinib-induced autophagy in both HEY and SKOv3 cells. Dasatinib also reduced Bcl-2 expression and activity. Overexpression of Bcl-2 partially prevented dasatinib-induced autophagy. CONCLUSIONS We conclude that dasatinib induces autophagic cell death in ovarian cancer that partially depends on Beclin-1, AKT and Bcl-2. These results may have implications for clinical use of dasatinib. PMID:20629079

  10. Down-regulation of phospho-non-receptor Src tyrosine kinases contributes to growth inhibition of cervical cancer cells.

    PubMed

    Kong, Lu; Deng, Zhihong; Zhao, Yanzhong; Wang, Yamei; Sarkar, Fazlul H; Zhang, Yuxiang

    2011-12-01

    Non-receptor Src tyrosine kinases (nrTKs) are overexpressed in a variety of human tumors, including cancer of the colon, breast, and pancreas, and their inhibitors are under intensive investigations as novel anti-tumor agents. However, these studies are not clear in the case of cervical cancer. Therefore, we studied the role of nrTKs and their inhibitors in the cervical cancer. The expression level of phospho-SrcY416 (pSrcY416) in several cervical cancer cell lines, normal cervical tissues, and cervical cancer tissues have been examined, and it has also been done whether PP2, an inhibitor of Src kinase, can inhibit the growth of cervical cells in vitro and in vivo. Immunohistochemical and confocal microscope studies suggested that pSrcY416 is overexpressed in HeLa and SiHa cells, as well as in the clinical cervical cancer tissues, compared to the normal cervical tissues. Down-regulation of pSrcY416 inhibited the cell proliferation by increasing the HeLa or SiHa cell population in the G0-G1 phase or S phase, respectively. Down-regulation of pSrcY416 led to up-regulation of p21Cip1 and p27Kip1 in both HeLa and SiHa cells and decreased the expression of cyclin A1, cyclin E, and cyclin-dependent kinase-2,-6 (CDK-2,-6) in HeLa cells and of cyclin B and CDK-2 in SiHa cells. Nude mice xenograft data showed that PP2 inhibited subcutaneous tumor growth significantly (P<0.05, compared with the control). Down-regulation of pSrcY416 contributes to cell growth inhibition in cervical cancer cells. nrTKs could therefore be a novel therapeutic targets for the treatment of cervical cancer. PMID:20532678

  11. Translation initiation factor eIF3b expression in human cancer and its role in tumor growth and lung colonization

    PubMed Central

    Wang, Hong; Ru, Yuanbin; Sanchez-Carbayo, Marta; Wang, Xuejiao; Kieft, Jeffrey S.; Theodorescu, Dan

    2013-01-01

    Purpose Discovery transcriptomic analyses suggest eukaryotic initiation factor 3b (eIF3b) is elevated in human bladder and prostate cancer, yet its role as a prognostic factor or its requirement in the maintenance or progression of human cancer is not established. Here we determine the therapeutic potential of eIF3b by examining the clinical relevance of its expression in human cancer tissues and its role in experimental tumor models. Experimental Design We examined mRNA expression of eIF3b in bladder (N=317) and prostate (N=566) tissue samples and protein expression by immunohistochemistry in 143 bladder tumor samples as a function of clinicopathologic features. The impact of eIF3b depletion by siRNA in human cancer lines was evaluated in regards to in vitro cell growth, cell cycle, migration, in vivo subcutaneous tumor growth and lung colonization. Results eIF3b mRNA expression correlated to tumor grade, stage and survival in human bladder and prostate cancer. eIF3b protein expression stratified survival in human bladder cancer. eIF3b depletion reduced in vitro cancer cell growth; inhibited G1/S cell cycle transition by changing protein but not RNA expression of Cyclin A, E, Rb and p27Kip1; inhibited migration and disrupted actin cytoskeleton and focal adhesions. These changes were associated with decreased protein expression of integrin α5. Integrin α5 depletion phenocopied effects observed with eIF3b. eIF3b depleted bladder cancer cells formed fewer subcutaneous tumors that grew more slowly and had reduced lung colonization. Conclusion eIF3b expression relates to human bladder and prostate cancer prognosis, is required for tumor growth and thus a candidate therapeutic target. PMID:23575475

  12. The pentacyclic triterpenoid, plectranthoic acid, a novel activator of AMPK induces apoptotic death in prostate cancer cells.

    PubMed

    Akhtar, Nosheen; Syed, Deeba N; Khan, Mohammad Imran; Adhami, Vaqar M; Mirza, Bushra; Mukhtar, Hasan

    2016-01-26

    Epidemiologic studies indicated that diabetics treated with metformin had a lower incidence of cancer than those taking other anti-diabetes drugs. This led to a surge in the efforts for identification of safer and more effective metformin mimetic compounds. The plant Ficus microcarpa is widely used for the treatment of type 2 diabetes in traditional medicine in South Asia. We obtained extracts from this plant and identified a small molecule, plectranthoic acid (PA), with potent 5'AMP-activated kinase (AMPK) activating properties far superior than metformin. AMPK is the central hub of metabolic regulation and a well-studied therapeutic target for metabolic syndrome, type-2 diabetes and cancer. We observed that treatment of prostate cancer (PCa) cells with PA inhibited proliferation and induced G0/G1 phase cell cycle arrest that was associated with up-regulation of cyclin kinase inhibitors p21/CIP1 and p27/KIP1. PA treatment suppressed mTOR/S6K signaling and induced apoptosis in PCa cells in an AMPK-dependent manner. Interestingly, PA-induced autophagy in PCa cells was found to be independent of AMPK activation. Combination studies of PA and metformin demonstrated that metformin had an inhibitory effect on PA-induced AMPK activation and suppressed PA-mediated apoptosis. Given the anti-proliferative role of PA in cancer and its potent anti-hyperglycemic activity, we suggest that PA should be explored further as a novel activator of AMPK for its ultimate use for the prevention of cancers and treatment of type 2 diabetes. PMID:26683363

  13. The goya mouse mutant reveals distinct newly identified roles for MAP3K1 in the development and survival of cochlear sensory hair cells

    PubMed Central

    Parker, Andrew; Cross, Sally H.; Jackson, Ian J.; Hardisty-Hughes, Rachel; Morse, Susan; Nicholson, George; Coghill, Emma; Bowl, Michael R.; Brown, Steve D. M.

    2015-01-01

    ABSTRACT Mitogen-activated protein kinase, MAP3K1, plays an important role in a number of cellular processes, including epithelial migration during eye organogenesis. In addition, studies in keratinocytes indicate that MAP3K1 signalling through JNK is important for actin stress fibre formation and cell migration. However, MAP3K1 can also act independently of JNK in the regulation of cell proliferation and apoptosis. We have identified a mouse mutant, goya, which exhibits the eyes-open-at-birth and microphthalmia phenotypes. In addition, these mice also have hearing loss. The goya mice carry a splice site mutation in the Map3k1 gene. We show that goya and kinase-deficient Map3k1 homozygotes initially develop supernumerary cochlear outer hair cells (OHCs) that subsequently degenerate, and a progressive profound hearing loss is observed by 9 weeks of age. Heterozygote mice also develop supernumerary OHCs, but no cellular degeneration or hearing loss is observed. MAP3K1 is expressed in a number of inner-ear cell types, including outer and inner hair cells, stria vascularis and spiral ganglion. Investigation of targets downstream of MAP3K1 identified an increase in p38 phosphorylation (Thr180/Tyr182) in multiple cochlear tissues. We also show that the extra OHCs do not arise from aberrant control of proliferation via p27KIP1. The identification of the goya mutant reveals a signalling molecule involved with hair-cell development and survival. Mammalian hair cells do not have the ability to regenerate after damage, which can lead to irreversible sensorineural hearing loss. Given the observed goya phenotype, and the many diverse cellular processes that MAP3K1 is known to act upon, further investigation of this model might help to elaborate upon the mechanisms underlying sensory hair cell specification, and pathways important for their survival. In addition, MAP3K1 is revealed as a new candidate gene for human sensorineural hearing loss. PMID:26542706

  14. GNA13 as a prognostic factor and mediator of gastric cancer progression

    PubMed Central

    Chen, Jie-Wei; Weng, Hui-Wen; Zheng, Zou-San; Chen, Cui; Xie, Dan; Ye, Sheng

    2016-01-01

    Guanine nucleotide binding protein (G protein), alpha 13 (GNA13) has been implicated as an oncogenic protein in several human cancers. In this study, GNA13 was characterized for its role in gastric cancer (GC) progression and underlying molecular mechanisms. The expression dynamics of GNA13 were examined by immunohistochemistry (IHC) in two independent cohorts of GC samples. A series of in-vivo and in-vitro assays was performed to elucidate the function of GNA13 in GC and its underlying mechanisms. In both two cohorts of GC samples, we observed that GNA13 was markedly overexpressed in GC tissues and associated closely with aggressive magnitude of GC progression and poor patients' survival. Further study showed that upregulation of GNA13 expression increased the proliferation and tumorigenicity of GC cells in vitro and in vivo, by promoting cell growth rate, colony formation, and tumor formation in nude mice. By contrast, knockdown of GNA13 effectively suppressed the proliferation and tumorigenicity of GC cells in vitro and in vivo. Our results also demonstrated that the molecular mechanisms of the effect of GNA13 in GC included promotion of G1/S cell cycle transition through upregulation of c-Myc, activation of AKT and ERK activity, suppression of FOXO1 activity, upregulation of cyclin-dependent kinase (CDK) regulator cyclin D1 and downregulation of CDK inhibitor p21Cip1 and p27Kip1. Our present study illustrated that GNA13 has an important role in promoting proliferation and tumorigenicity of GC, and may represent a novel prognostic biomarker and therapeutic target for this disease. PMID:26735177

  15. KRAS mutational subtype and copy number predict in vitro response of human pancreatic cancer cell lines to MEK inhibition

    PubMed Central

    Hamidi, H; Lu, M; Chau, K; Anderson, L; Fejzo, M; Ginther, C; Linnartz, R; Zubel, A; Slamon, D J; Finn, R S

    2014-01-01

    Background: To study the molecular mechanism regulating sensitivity to MEK inhibition in pancreatic cancer cell lines. Methods: A growth inhibition assay determined sensitivity to MEK162 in a panel of 29 pancreatic cancer cell lines. For the same panel, KRAS mutational status and copy-number variation (CNV) was determine using PCR, array CGH and FISH. Two sensitive and two resistant cell lines were further interrogated for difference in baseline and MEK162-induced gene expression, as well as signal transduction using microarray and western blotting. Cell cycle and apoptosis analysis was measured by flow cytometry. Results: We report a strong correlation between both specific KRAS mutational subtype and CNV, and sensitivity to MEK inhibition. Cell lines with a KRAS (V12) mutation and KRAS gains or loss (n=7) are ∼10 times more resistant than those having neither a KRAS (V12) mutation nor KRAS CNV (n=14). Significant differences in baseline and MEK162-induced gene expression exist between the sensitive and resistant lines, especially in genes involved in RAS, EGF receptor and PI3K pathways. This was further supported by difference in signal transduction. MEK 162 blocked ERK1/2, as well as inhibited PI3K and S6 and increased p27KIP1 levels in the sensitive lines. Conclusions: Given the potency of MEK162, it may be a promising new therapy for patients with pancreatic cancer and KRAS mutational subtypes, and CNV may serve as important biomarkers for selecting patients that benefit from MEK-targeting based on these preclinical data. PMID:25167228

  16. Cavin-2 in oral cancer: A potential predictor for tumor progression.

    PubMed

    Unozawa, Motoharu; Kasamatsu, Atsushi; Higo, Morihiro; Fukumoto, Chonji; Koyama, Tomoyoshi; Sakazume, Tomomi; Nakashima, Dai; Ogawara, Katsunori; Yokoe, Hidetaka; Shiiba, Masashi; Tanzawa, Hideki; Uzawa, Katsuhiro

    2016-06-01

    Cavin-2 (CVN2) affects formation of large caveolae, which are membrane-rich cholesterol domains associated with several functions in signal transduction. Accumulating evidence suggests that CVN2 is present in many cellular types; however, the molecular mechanisms of CVN2 in cancers and its clinical relevance are unknown. We proposed a mechanism by which CVN2 regulates caveolin-1 expression leading to slow cellular proliferation by inactivation of the extracellular regulated kinase (ERK) pathway. Quantitative reverse transcriptase-polymerase chain reaction and immunoblot analyses were used to assess the CVN2 regulation mechanism in oral squamous cell carcinoma (OSCC). Immunohistochemistry (IHC) was performed to analyze the correlation between CVN2 expression and clinical behavior in 115 patients with OSCC. A CVN2 overexpressed model of OSCC cells (oeCVN2 cells) was used for functional experiments. CVN2 expression was down-regulated significantly (P < 0.05) in OSCCs compared with normal counterparts in vitro and in vivo. In addition to the findings that a serum deprivation culture induced up-regulation of CVN2 and slowed cellular proliferation, oeCVN2 cell growth decreased because of cell-cycle arrest at the G1 phase resulting from up-regulated cyclin-dependent kinase inhibitors (p21(Cip1) and p27(Kip1) ) and down-regulated cyclins (cyclin D1, cyclin E) and cyclin-dependent kinases (CDK2, CDK4, and CDK6). Interestingly, CVN2 overexpression facilitated caveolin-1 recruitment and colocalization with each other. We also found decreased ERK phosphorylation levels, an upstream event in cell-cycle arrest. Clinically, IHC data from primary OSCCs showed high tumoral progression in CVN2-negative patients with OSCC. CVN2 may be a possible key regulator of OSCC progression via the CVN2/caveolin-1/ERK pathway and a potential therapeutic target for developing new treatments for OSCCs. © 2015 Wiley Periodicals, Inc. PMID:26086332

  17. Piperine, an alkaloid from black pepper, inhibits growth of human colon cancer cells via G1 arrest and apoptosis triggered by endoplasmic reticulum stress.

    PubMed

    Yaffe, Paul B; Power Coombs, Melanie R; Doucette, Carolyn D; Walsh, Mark; Hoskin, David W

    2015-10-01

    Piperine, a piperidine alkaloid present in black pepper, inhibits the growth of cancer cells, although the mechanism of action is not well understood. In this study, we show that piperine (75-150 µM) inhibited the growth of several colon cancer cell lines but had little effect on the growth of normal fibroblasts and epithelial cells. Piperine inhibited HT-29 colon carcinoma cell proliferation by causing G1 phase cell cycle arrest that was associated with decreased expression of cyclins D1 and D3 and their activating partner cyclin-dependent kinases 4 and 6, as well as reduced phosphorylation of the retinoblastoma protein and up-regulation of p21/WAF1 and p27/KIP1 expression. In addition, piperine caused hydroxyl radical production and apoptosis that was partially dependent on the production of reactive oxygen species. Piperine-treated HT-29 cells showed loss of mitochondrial membrane integrity and cleavage of poly (ADP-ribose) polymerase-1, as well as caspase activation and reduced apoptosis in the presence of the pan-caspase inhibitor zVAD-FMK. Increased expression of the endoplasmic reticulum stress-associated proteins inositol-requiring 1α protein, C/EBP homologous protein, and binding immunoglobulin protein, and activation of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase, as well as decreased phosphorylation of Akt and reduced survivin expression were also observed in piperine-treated HT-29 cells. Furthermore, piperine inhibited colony formation by HT-29 cells, as well as the growth of HT-29 spheroids. Cell cycle arrest and endoplasmic reticulum stress-associated apoptosis following piperine treatment of HT-29 cells provides the first evidence that piperine may be useful in the treatment of colon cancer. PMID:24819444

  18. Regulation of the cell cycle and PI3K/Akt/mTOR signaling pathway by tanshinone I in human breast cancer cell lines

    PubMed Central

    WANG, LI; WU, JIANZHONG; LU, JIANWEI; MA, RONG; SUN, DAWEI; TANG, JINHAI

    2015-01-01

    Breast cancer is the second leading cause of cancer-related mortality in females worldwide. Therefore, identifying alternative strategies to combat the disease mortality is important. The aim of the present study was to investigate the effect of tanshinone I (Tan I) on the tumorigenicity of estrogen-responsive MCF-7 and estrogen-independent MDA-MB-453 human breast cancer cells. The cytotoxicity of Tan I was evaluated using a Cell Counting Kit-8 assay, the apoptosis and cell cycle distribution were detected using flow cytometry and the cell morphology was observed using a fluorescence microscope. In addition, the cell cycle regulatory proteins and apoptosis-associated proteins involved in the phosphatidylinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway were detected using western blot analysis using specific protein antibodies. The MCF-7 and MDA-MB-453 cells were equally sensitive to Tan I regardless of their responsiveness to estrogen. Tan I exerted similar antiproliferative activities and induction of apoptosis, resulting in S phase arrest accompanied by decreases in cyclin B and increases in cyclin E and cyclin A proteins, which may have been associated with the upregulation of cyclin-dependent kinase inhibitors p21Cip1 and p27Kip1. In addition, Tan I was found to downregulate anti-apoptotic and upregulate associated apoptotic components of the PI3K/Akt/mTOR signaling pathway. Notably, treatment with the PI3K inhibitor, LY294002, decreased the levels of phosphorylated (p)-PI3K, p-Akt and p-mTOR. These results clearly indicated that the mechanism of action of Tan I involved, at least partially, an effect on the PI3K/Akt/mTOR signaling pathway, providing new information for anticancer drug design and development. PMID:25355053

  19. Vitamin E δ-tocotrienol prolongs survival in the LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx-1-Cre (KPC) transgenic mouse model of pancreatic cancer.

    PubMed

    Husain, Kazim; Centeno, Barbara A; Chen, Dung-Tsa; Hingorani, Sunil R; Sebti, Said M; Malafa, Mokenge P

    2013-10-01

    Previous work has shown that vitamin E δ-tocotrienol (VEDT) prolongs survival and delays progression of pancreatic cancer in the LSL-Kras(G12D)(/+);Pdx-1-Cre mouse model of pancreatic cancer. However, the effect of VEDT alone or in combination with gemcitabine in the more aggressive LSL-Kras(G12D)(/+);LSL-Trp53(R172H)(/+);Pdx-1-Cre (KPC) mouse model is unknown. Here, we studied the effects of VEDT and the combination of VEDT and gemcitabine in the KPC mice. KPC mice were randomized into four groups: (i) vehicle [olive oil, 1.0 mL/kg per os twice a day and PBS 1.0 mL/kg intrapertoneally (i.p.) twice a week], (ii) gemcitabine (100 mg/kg i.p. twice a week), (iii) VEDT (200 mg/kg per os twice a day), and (iv) gemcitabine + VEDT. Mice received treatment until they displayed symptoms of impending death from pancreatic cancer, at which point animals were euthanized. At 16 weeks, survival was 10% in the vehicle group, 30% in the gemcitabine group, 70% in the VEDT group (P < 0.01), and 90% in the VEDT combined with gemcitabine group (P < 0.05). VEDT alone and combined with gemcitabine resulted in reversal of epithelial-to-mesenchymal transition in tumors. Biomarkers of apoptosis (plasma CK18), PARP1 cleavage, and Bax expression were more greatly induced in tumors subjected to combined treatment versus individual treatment. Combined treatment induced cell-cycle inhibitors (p27(Kip1) and p21(Cip1)) and inhibited VEGF, vascularity (CD31), and oncogenic signaling (pAKT, pMEK, and pERK) greater than individual drugs. No significant differences in body weight gain between drug treatment and control mice were observed. These results strongly support further investigation of VEDT alone and in combination with gemcitabine for pancreatic cancer prevention and treatment. PMID:23963802

  20. The Topoisomerase 1 Inhibitor Austrobailignan-1 Isolated from Koelreuteria henryi Induces a G2/M-Phase Arrest and Cell Death Independently of p53 in Non-Small Cell Lung Cancer Cells.

    PubMed

    Wu, Chun-Chi; Huang, Keh-Feng; Yang, Tsung-Ying; Li, Ya-Ling; Wen, Chi-Luan; Hsu, Shih-Lan; Chen, Tzu-Hsiu

    2015-01-01

    Koelreuteria henryi Dummer, an endemic plant of Taiwan, has been used as a folk medicine for the treatment of hepatitis, enteritis, cough, pharyngitis, allergy, hypertension, hyperlipidemia, and cancer. Austrobailignan-1, a natural lignan derivative isolated from Koelreuteria henryi Dummer, has anti-oxidative and anti-cancer properties. However, the effects of austrobailignan-1 on human cancer cells have not been studied yet. Here, we showed that austrobailignan-1 inhibited cell growth of human non-small cell lung cancer A549 and H1299 cell lines in both dose- and time-dependent manners, the IC50 value (48 h) of austrobailignan-1 were 41 and 22 nM, respectively. Data from flow cytometric analysis indicated that treatment with austrobailignan-1 for 24 h retarded the cell cycle at the G2/M phase. The molecular event of austrobailignan-1-mediated G2/M phase arrest was associated with the increase of p21Waf1/Cip1 and p27Kip1 expression, and decrease of Cdc25C expression. Moreover, treatment with 100 nM austrobailignan-1 for 48 h resulted in a pronounced release of cytochrome c followed by the activation of caspase-2, -3, and -9, and consequently induced apoptosis. These events were accompanied by the increase of PUMA and Bax, and the decrease of Mcl-1 and Bcl-2. Furthermore, our study also showed that austrobailignan-1 was a topoisomerase 1 inhibitor, as evidenced by a relaxation assay and induction of a DNA damage response signaling pathway, including ATM, and Chk1, Chk2, γH2AX phosphorylated activation. Overall, our results suggest that austrobailignan-1 is a novel DNA damaging agent and displays a topoisomerase I inhibitory activity, causes DNA strand breaks, and consequently induces DNA damage response signaling for cell cycle G2/M arrest and apoptosis in a p53 independent manner. PMID:26147394

  1. The Topoisomerase 1 Inhibitor Austrobailignan-1 Isolated from Koelreuteria henryi Induces a G2/M-Phase Arrest and Cell Death Independently of p53 in Non-Small Cell Lung Cancer Cells

    PubMed Central

    Yang, Tsung-Ying; Li, Ya-Ling; Wen, Chi-Luan; Hsu, Shih-Lan; Chen, Tzu-Hsiu

    2015-01-01

    Koelreuteria henryi Dummer, an endemic plant of Taiwan, has been used as a folk medicine for the treatment of hepatitis, enteritis, cough, pharyngitis, allergy, hypertension, hyperlipidemia, and cancer. Austrobailignan-1, a natural lignan derivative isolated from Koelreuteria henryi Dummer, has anti-oxidative and anti-cancer properties. However, the effects of austrobailignan-1 on human cancer cells have not been studied yet. Here, we showed that austrobailignan-1 inhibited cell growth of human non-small cell lung cancer A549 and H1299 cell lines in both dose- and time-dependent manners, the IC50 value (48 h) of austrobailignan-1 were 41 and 22 nM, respectively. Data from flow cytometric analysis indicated that treatment with austrobailignan-1 for 24 h retarded the cell cycle at the G2/M phase. The molecular event of austrobailignan-1-mediated G2/M phase arrest was associated with the increase of p21Waf1/Cip1 and p27Kip1 expression, and decrease of Cdc25C expression. Moreover, treatment with 100 nM austrobailignan-1 for 48 h resulted in a pronounced release of cytochrome c followed by the activation of caspase-2, -3, and -9, and consequently induced apoptosis. These events were accompanied by the increase of PUMA and Bax, and the decrease of Mcl-1 and Bcl-2. Furthermore, our study also showed that austrobailignan-1 was a topoisomerase 1 inhibitor, as evidenced by a relaxation assay and induction of a DNA damage response signaling pathway, including ATM, and Chk1, Chk2, γH2AX phosphorylated activation. Overall, our results suggest that austrobailignan-1 is a novel DNA damaging agent and displays a topoisomerase I inhibitory activity, causes DNA strand breaks, and consequently induces DNA damage response signaling for cell cycle G2/M arrest and apoptosis in a p53 independent manner. PMID:26147394

  2. Cks1 is required for tumor cell proliferation but not sufficient to induce hematopoietic malignancies.

    PubMed

    Kratzat, Susanne; Nikolova, Viktoriya; Miething, Cornelius; Hoellein, Alexander; Schoeffmann, Stephanie; Gorka, Oliver; Pietschmann, Elke; Illert, Anna-Lena; Ruland, Jürgen; Peschel, Christian; Nilsson, Jonas; Duyster, Justus; Keller, Ulrich

    2012-01-01

    The Cks1 component of the SCF(Skp2) complex is necessary for p27(Kip1) ubiquitylation and degradation. Cks1 expression is elevated in various B cell malignancies including Burkitt lymphoma and multiple myeloma. We have previously shown that loss of Cks1 results in elevated p27(Kip1) levels and delayed tumor development in a mouse model of Myc-induced B cell lymphoma. Surprisingly, loss of Skp2 in the same mouse model also resulted in elevated p27(Kip1) levels but exhibited no impact on tumor onset. This raises the possibility that Cks1 could have other oncogenic activities than suppressing p27(Kip1). To challenge this notion we have targeted overexpression of Cks1 to B cells using a conditional retroviral bone marrow transduction-transplantation system. Despite potent ectopic overexpression, Cks1 was unable to promote B cell hyperproliferation or B cell malignancies, indicating that Cks1 is not oncogenic when overexpressed in B cells. Since Skp2 overexpression can drive T-cell tumorigenesis or other cancers we also widened the quest for oncogenic activity of Cks1 by ubiquitously expressing Cks1 in hematopoetic progenitors. At variance with c-Myc overexpression, which caused acute myeloid leukemia, Cks1 overexpression did not induce myeloproliferation or leukemia. Therefore, despite being associated with a poor prognosis in various malignancies, sole Cks1 expression is insufficient to induce lymphoma or a myeloproliferative disease in vivo. PMID:22624029

  3. Hypoxia-mediated cancer stem cells in pseudopalisades with activation of hypoxia-inducible factor-1α/Akt axis in glioblastoma.

    PubMed

    Inukai, Madoka; Hara, Atsuko; Yasui, Yoshie; Kumabe, Toshihiro; Matsumoto, Toshihide; Saegusa, Makoto

    2015-10-01

    Pseudopalisades (Ps) around necrotic foci are severely hypoxic and overexpress hypoxia-inducible factor (HIF) in glioblastoma (GBM). Hypoxic regions have been proposed as one of several distinct niches for cancer stem cells (CSCs) in GBM, but little is known about the association between Ps features and CSC properties. Herein, we focused on the biological role of Ps lesions. In clinical cases of GBM, expression of hypoxia-related molecules including HIF-1α, Glut-1, p27(Kip1), and pAkt was significantly increased in perinecrotic Ps lesions compared with nonnecrotic areas and perinecrotic lesions lacking Ps features. Significantly higher expression levels of several CSC-related markers, including CD133, Sox2, CD44s, and aldehyde dehydrogenase (ALDH) 1, were also observed in Ps lesions, which were positively correlated with expression of hypoxia-related molecules and pAkt. Ps lesions also showed increased number of apoptotic cells and decreased bcl-2 and survivin expression compared with the surrounding tissue. Short-term exposure of astrocytoma cell lines to cobalt chloride, which is known to mimic the effect of hypoxia, caused an increase in expression of both hypoxia- and CSC-related markers, in line with increases in the ALDH(high) cell population and number of spheroids. Inhibition of endogenous Akt by LY294002 resulted in decreased expression of Sox2, ALDH1, and CD133, leading to enhancement of cobalt chloride-mediated apoptotic events due to altered ratio of bcl-2 to bax expression. These findings suggest that Ps lesions within GBM may serve as a specialized hypoxic niche, in which the HIF-1α/pAkt axis is activated, in response to severe hypoxia. PMID:26256949

  4. Combination of carmustine and selenite effectively inhibits tumor growth by targeting androgen receptor, androgen receptor-variants, and Akt in preclinical models: New hope for patients with castration resistant prostate cancer.

    PubMed

    Thamilselvan, Vijayalakshmi; Menon, Mani; Thamilselvan, Sivagnanam

    2016-10-01

    Despite established androgen receptor (AR) antagonists, AR/AR-variants signaling remain a major obstacle for the successful treatment of castration resistant prostate cancer (CRPC). In addition, CRPC cells adapt to survive via AR-independent pathways to escape next generation therapies. Therefore, there is an urgent need for drugs that can target these signaling pathways in CRPC. In this study, we sought to determine whether carmustine and selenite in combination could induce apoptosis and inhibit growth of CRPC in-vitro and in-vivo. CRPC (22Rv1, VCaP, and PC-3) cell lines in culture and xenograft mouse were used. Combination of carmustine and selenite treatment significantly increased reactive oxygen species, apoptosis and growth inhibition in CRPC cells with down regulation of anti-apoptotic (Bcl-2 and Mcl-1) and proliferative proteins (c-Myc and cyclin-D1). This effect was associated with complete reduction of AR/AR-variants, AR-V7, PSA and significant induction of p27Kip1. Combination treatment substantially abolished phospho-Akt, phospho-GSK-3β, and anchorage-independent growth in AR-positive and AR-negative cells. Consistent with in-vitro results, combination treatment effectively induced apoptosis and completely inhibited xenograft tumor growth and markedly reduced AR/AR-variants, AR-V7, PSA, and Bcl-2 in xenograft tumors without causing genotoxicity in host mice. Individual agent treatment showed only partial effect. The combination treatment showed a significant synergistic effect. The present study is the first to demonstrate that the combination of carmustine and selenite treatment completely suppressed CRPC tumor growth by reducing AR/AR-variants and Akt signaling. Our findings suggest that the combination of carmustine and selenite could constitute a promising next-generation therapy for successful treatment of patients with CRPC. PMID:27198552

  5. Cathepsin B promotes colorectal tumorigenesis, cell invasion, and metastasis

    PubMed Central

    Bian, Benjamin; Mongrain, Sébastien; Cagnol, Sébastien; Langlois, Marie‐Josée; Boulanger, Jim; Bernatchez, Gérald; Carrier, Julie C.; Boudreau, François

    2015-01-01

    Cathepsin B is a cysteine proteinase that primarily functions as an endopeptidase within endolysosomal compartments in normal cells. However, during tumoral expansion, the regulation of cathepsin B can be altered at multiple levels, thereby resulting in its overexpression and export outside of the cell. This may suggest a possible role of cathepsin B in alterations leading to cancer progression. The aim of this study was to determine the contribution of intracellular and extracellular cathepsin B in growth, tumorigenesis, and invasion of colorectal cancer (CRC) cells. Results show that mRNA and activated levels of cathepsin B were both increased in human adenomas and in CRCs of all stages. Treatment of CRC cells with the highly selective and non‐permeant cathepsin B inhibitor Ca074 revealed that extracellular cathepsin B actively contributed to the invasiveness of human CRC cells while not essential for their growth in soft agar. Cathepsin B silencing by RNAi in human CRC cells inhibited their growth in soft agar, as well as their invasion capacity, tumoral expansion, and metastatic spread in immunodeficient mice. Higher levels of the cell cycle inhibitor p27Kip1 were observed in cathepsin B‐deficient tumors as well as an increase in cyclin B1. Finally, cathepsin B colocalized with p27Kip1 within the lysosomes and efficiently degraded the inhibitor. In conclusion, the present data demonstrate that cathepsin B is a significant factor in colorectal tumor development, invasion, and metastatic spreading and may, therefore, represent a potential pharmacological target for colorectal tumor therapy. © 2015 The Authors. Molecular Carcinogenesis, published by Wiley Periodicals, Inc. PMID:25808857

  6. 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces expression of p27(kip¹) and FoxO3a in female rat cerebral cortex and PC12 cells.

    PubMed

    Xu, Guangfei; Liu, Jiao; Yoshimoto, Katsuhiko; Chen, Gang; Iwata, Takeo; Mizusawa, Noriko; Duan, Zhiqing; Wan, Chunhua; Jiang, Junkang

    2014-05-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent toxin that alters normal brain development, producing cognitive disability and motor dysfunction. Previous studies in rats have proved that female rats are more sensitive to TCDD lethality than male ones. Recent studies have shown that TCDD induces cell cycle arrest and apoptosis, but the regulatory proteins involved in these processes have yet to be elucidated. In this study, we constructed an acute TCDD injury female rat model, and investigated the effects of TCDD on apoptosis and expression of cell cycle regulators, forkhead box class O 3a (FoxO3a) and p27(kip1), in the central nervous system (CNS). Increased levels of active caspase-3 were observed in the cerebral cortex of female rats treated with TCDD, suggesting that TCDD-induced apoptosis occurs in the CNS. The terminal deoxynucleotidyl transferase-mediated biotinylated-dUTP nick-end labeling assay showed that apoptosis primarily occurred in neurons. Furthermore, Western blot analysis, reverse transcription-polymerase chain reaction, and immunohistochemistry showed a significant up-regulation of FoxO3a and p27(kip1) in the cerebral cortex. Immunofluorescent labeling indicated that FoxO3a and p27(kip1) were predominantly localized in apoptotic neurons, but not in astrocytes. In vitro experiments using PC12, a rat neuron-like pheochromocytoma cell line, also revealed that TCDD induced apoptosis and an increase in FoxO3a and p27(kip1) expression. Furthermore, knockdown of FoxO3a expression inhibited p27(kip1) transcription and TCDD-induced apoptosis. Based on our data, induction of FoxO3a may play an important role in TCDD-induced neurotoxicity. PMID:24594276

  7. Phosphoproteome Integration Reveals Patient-Specific Networks in Prostate Cancer.

    PubMed

    Drake, Justin M; Paull, Evan O; Graham, Nicholas A; Lee, John K; Smith, Bryan A; Titz, Bjoern; Stoyanova, Tanya; Faltermeier, Claire M; Uzunangelov, Vladislav; Carlin, Daniel E; Fleming, Daniel Teo; Wong, Christopher K; Newton, Yulia; Sudha, Sud; Vashisht, Ajay A; Huang, Jiaoti; Wohlschlegel, James A; Graeber, Thomas G; Witte, Owen N; Stuart, Joshua M

    2016-08-11

    We used clinical tissue from lethal metastatic castration-resistant prostate cancer (CRPC) patients obtained at rapid autopsy to evaluate diverse genomic, transcriptomic, and phosphoproteomic datasets for pathway analysis. Using Tied Diffusion through Interacting Events (TieDIE), we integrated differentially expressed master transcriptional regulators, functionally mutated genes, and differentially activated kinases in CRPC tissues to synthesize a robust signaling network consisting of druggable kinase pathways. Using MSigDB hallmark gene sets, six major signaling pathways with phosphorylation of several key residues were significantly enriched in CRPC tumors after incorporation of phosphoproteomic data. Individual autopsy profiles developed using these hallmarks revealed clinically relevant pathway information potentially suitable for patient stratification and targeted therapies in late stage prostate cancer. Here, we describe phosphorylation-based cancer hallmarks using integrated personalized signatures (pCHIPS) that shed light on the diversity of activated signaling pathways in metastatic CRPC while providing an integrative, pathway-based reference for drug prioritization in individual patients. PMID:27499020

  8. G1-checkpoint function including a cyclin-dependent kinase 2 regulatory pathway as potential determinant of 7-hydroxystaurosporine (UCN-01)-induced apoptosis and G1-phase accumulation.

    PubMed

    Akiyama, T; Sugiyama, K; Shimizu, M; Tamaoki, T; Akinaga, S

    1999-12-01

    7-Hydroxystaurosporine (UCN-01), which was originally identified as a protein kinase C selective inhibitor, is currently in clinical trials as an anti-cancer drug. We previously showed that UCN-01 induced preferential G1-phase accumulation in tumor cells and this effect was associated with the retinoblastoma (Rb) protein and its regulatory factors, such as cyclin-dependent kinase 2 (CDK2) and CDK inhibitors p21Cip1/WAF1 and p27Kip1. We demonstrate here that G1-phase accumulation was induced by UCN-01 in Rb-proficient cell lines (WiDr and HCT116 human colon carcinomas and WI-38 human lung fibroblast), and it was accompanied by dephosphorylation of Rb. In addition, UCN-01-induced G1-phase accumulation was also demonstrated in a Rb-defective cell line (Saos-2 human osteosarcoma), but not in a simian virus 40 (SV40)-transformed cell line (WI-38 VA13). Apoptosis was induced by UCN-01 in the two Rb-deficient cell lines, but not in the other Rb-proficient cell lines. These observations suggest that G1-checkpoint function might be important for cell survival during UCN-01 treatment. In addition, there may be a UCN-01-responsive factor in the G1-checkpoint machinery other than Rb which is targeted by SV40. Further studies revealed a correlation between UCN-01-induced G1-phase accumulation and reduction of cellular CDK2 kinase activity. This reduction was strictly dependent on down-regulation of the Thr160-phosphorylated form of CDK2 protein, and coincided in part with up-regulation of p27Kip1, but it was independent of the level of the p21Cip1/WAF1 protein. These results suggest that G1-checkpoint function, including a CDK2-regulatory pathway, may be a significant determinant of the sensitivity of tumor cells to UCN-01. PMID:10665655

  9. Resistance to cisplatin and paclitaxel does not affect the sensitivity of human ovarian cancer cells to antiprogestin-induced cytotoxicity

    PubMed Central

    2014-01-01

    Background Antiprogestin compounds have been shown to be effective in blocking the growth of ovarian cancer cells of different genetic backgrounds. Herein we studied the anti-ovarian cancer effect of a series of antiprogestins sharing the chemical backbone of the most characterized antiprogestin, mifepristone, but with unique modifications in position C-17 of the steroid ring. We assessed the effect of mifepristone-like antiprogestins on the growth of ovarian cancer cells sensitive to the standard combination therapy cisplatin-paclitaxel or made double-resistant upon six cycles of pulse-selection with the drugs used at clinically relevant concentrations and exposure times. Methods IGROV-1 and SKOV-3 cells were pulsed with 20 μM cisplatin for 1 h followed by 100 nM paclitaxel for 3 h once a week for six weeks. The cells that did not die and repopulate the culture after the chemotherapies were termed Platinum-Taxane-EScape cells (PTES). Parental cells were compared against their PTES derivatives in their responses to further platinum-taxane treatments. Moreover, both ovarian cancer cells and their PTES siblings were exposed to escalating doses of the various antiprogestin derivatives. We assessed cell growth, viability and sub-G1 DNA content using microcapillary cytometry. Cyclin-dependent kinase inhibitors p21cip1 and p27kip1 and cleavage of downstream caspase-3 substrate PARP were used to assess whether cell fate, as a consequence of treatment, was limited to cytostasis or progressed to lethality. Results Cells subjected to six pulse-selection cycles of cisplatin-paclitaxel gave rise to sibling derivatives that displayed ~2-7 fold reduction in their sensitivities to further chemotherapy. However, regardless of the sensitivity the cells developed to the combination cisplatin-paclitaxel, they displayed similar sensitivity to the antiprogestins, which blocked their growth in a dose-related manner, with lower concentrations causing cytostasis, and higher

  10. Most small bowel cancers are revealed by a complication

    PubMed Central

    Negoi, Ionut; Paun, Sorin; Hostiuc, Sorin; Stoica, Bodgan; Tanase, Ioan; Negoi, Ruxandra Irina; Beuran, Mircea

    2015-01-01

    ABSTRACT Objective To characterize the pattern of primary small bowel cancers in a tertiary East-European hospital. Methods A retrospective study of patients with small bowel cancers admitted to a tertiary emergency center, over the past 15 years. Results There were 57 patients with small bowel cancer, representing 0.039% of admissions and 0.059% of laparotomies. There were 37 (64.9%) men, mean age of 58 years; and 72 years for females. Out of 57 patients, 48 (84.2%) were admitted due to an emergency situation: obstruction in 21 (38.9%), perforation in 17 (31.5%), upper gastrointestinal bleeding in 8 (14.8%), and lower gastrointestinal bleeding in 2 (3.7%). There were 10 (17.5%) duodenal tumors, 21 (36.8%) jejunal tumors and 26 (45.6%) ileal tumors. The most frequent neoplasms were gastrointestinal stromal tumor in 24 patients (42.1%), adenocarcinoma in 19 (33.3%), lymphoma in 8 (14%), and carcinoids in 2 (3.5%). The prevalence of duodenal adenocarcinoma was 14.55 times greater than that of the small bowel, and the prevalence of duodenal stromal tumors was 1.818 time greater than that of the small bowel. Obstruction was the complication in adenocarcinoma in 57.9% of cases, and perforation was the major local complication (47.8%) in stromal tumors. Conclusion Primary small bowel cancers are usually diagnosed at advanced stages, and revealed by a local complication of the tumor. Their surgical management in emergency setting is associated to significant morbidity and mortality rates. PMID:26676271

  11. A negative genetic interaction map in isogenic cancer cell lines reveals cancer cell vulnerabilities

    PubMed Central

    Vizeacoumar, Franco J; Arnold, Roland; Vizeacoumar, Frederick S; Chandrashekhar, Megha; Buzina, Alla; Young, Jordan T F; Kwan, Julian H M; Sayad, Azin; Mero, Patricia; Lawo, Steffen; Tanaka, Hiromasa; Brown, Kevin R; Baryshnikova, Anastasia; Mak, Anthony B; Fedyshyn, Yaroslav; Wang, Yadong; Brito, Glauber C; Kasimer, Dahlia; Makhnevych, Taras; Ketela, Troy; Datti, Alessandro; Babu, Mohan; Emili, Andrew; Pelletier, Laurence; Wrana, Jeff; Wainberg, Zev; Kim, Philip M; Rottapel, Robert; O'Brien, Catherine A; Andrews, Brenda; Boone, Charles; Moffat, Jason

    2013-01-01

    Improved efforts are necessary to define the functional product of cancer mutations currently being revealed through large-scale sequencing efforts. Using genome-scale pooled shRNA screening technology, we mapped negative genetic interactions across a set of isogenic cancer cell lines and confirmed hundreds of these interactions in orthogonal co-culture competition assays to generate a high-confidence genetic interaction network of differentially essential or differential essentiality (DiE) genes. The network uncovered examples of conserved genetic interactions, densely connected functional modules derived from comparative genomics with model systems data, functions for uncharacterized genes in the human genome and targetable vulnerabilities. Finally, we demonstrate a general applicability of DiE gene signatures in determining genetic dependencies of other non-isogenic cancer cell lines. For example, the PTEN−/− DiE genes reveal a signature that can preferentially classify PTEN-dependent genotypes across a series of non-isogenic cell lines derived from the breast, pancreas and ovarian cancers. Our reference network suggests that many cancer vulnerabilities remain to be discovered through systematic derivation of a network of differentially essential genes in an isogenic cancer cell model. PMID:24104479

  12. miR-3646 promotes cell proliferation, migration, and invasion via regulating G2/M transition in human breast cancer cells

    PubMed Central

    Tao, Shuang; Liu, Yao-Bang; Zhou, Zhi-Wei; Lian, Bin; Li, Hong; Li, Jin-Ping; Zhou, Shu-Feng

    2016-01-01

    MicroRNAs (miRNAs) are small non-coding RNAs that are often located in genomic breakpoint regions and play a critical role in regulating a variety of the cellular processes in human cancer. miR-3646 has been reported to take part in tumorigenic progression in breast and bladder cancer, but its potential functions and exact mechanistic roles in breast cancer are still unclear. The objective of this study was to investigate the role of miR-3646 in breast cancer growth and metastasis using both bioinformatic and experimental approaches. Before starting the bench work, we conducted a bioinformatic study to predict the target genes regulated by miR-3646 using a panel of different algorithms. The results showed that miR-3646 might regulate a large number of genes that are related to cell growth, proliferation, metabolis, transport, and apoptosis and some were cancer-related genes. We found that the expression level of miR-3646 was significantly upregulated in breast cancer cells and tissues compared with normal breast cells and no tumor tissues. Subsequently, the MTT and colony formation assay results showed that up-regulation of miR-3646 promoted the cell viability and proliferation. Our results also showed that down-regulation of miR-3646 arrested the cells in G2/M phase in MCF7 and MDA-MB-231 cells which was accompanied by the down-regulation of CDK1/CDC2 and cyclin B1 and upregulation of p21Waf1/Cip1, p27 Kip1, and p53, suggesting that down-regulation of miR-3646 induces G2/M arrest through activation of the p53/p21/CDC2/cyclin B1 pathway. In addition, overexpression of miR-3646 promoted migration and invasion of MCF7 and MDA-MB-231 cells. Taken together, miR-3646 is a potential oncogene in breast cancer and it may represent a new niomarker in the diagnosis and prediction of prognosis and therapeutic response. PMID:27186291

  13. The anti-erbB3 antibody MM-121/SAR256212 in combination with trastuzumab exerts potent antitumor activity against trastuzumab-resistant breast cancer cells

    PubMed Central

    2013-01-01

    Background Elevated expression of erbB3 receptor has been reported to induce resistance to therapeutic agents, including trastuzumab in erbB2-overexpressing breast cancer. Our recent studies indicate that erbB3 interacts with both erbB2 and IGF-1 receptor to form a heterotrimeric complex in trastuzumab-resistant breast cancer cells. Herein, we investigate the antitumor activity of MM-121/SAR256212, a fully human anti-erbB3 antibody (Ab), against two erbB2-overexpressing breast cancer cell lines resistant to trastuzumab. Methods MTS-based proliferation assays were used to determine cell viability upon treatment of trastuzumab and/or MM-121/SAR256212. Cell cycle progression was examined by flow cytometric analysis. Western blot analyses were performed to determine the expression and activation of proteins. Tumor xenografts were established by inoculation of the trastuzumab-resistant BT474-HR20 cells into nude mice. The tumor-bearing mice were treated with trastuzumab and/or MM-121/SAR256212 via i.p injection to determine the Abs’ antitumor activity. Immunohistochemical analyses were carried out to study the Abs’ inhibitory effects on tumor cell proliferation and induction of apoptosis in vivo. Results MM-121 significantly enhanced trastuzumab-induced growth inhibition in two sensitive and two resistant breast cancer cell lines. MM-121 in combination with trastuzumab resulted in a dramatic reduction of phosphorylated erbB3 (P-erbB3) and Akt (P-Akt) in the in vitro studies. MM-121 combined with trastuzumab did not induce apoptosis in the trastuzumab-resistant cell lines under our cell culture condition, rather induced cell cycle G1 arrest mainly associated with the upregulation of p27kip1. Interestingly, in the tumor xenograft model established from the trastuzumab-resistant cells, MM-121 in combination with trastuzumab as compared to either agent alone dramatically inhibited tumor growth correlated with a significant reduction of Ki67 staining and increase of

  14. Tumor and host factors that may limit efficacy of chemotherapy in non-small cell and small cell lung cancer.

    PubMed

    Stewart, David J

    2010-09-01

    and expression of GST-pi, RB or p27kip1 may possibly confer resistance. While limited clinical data suggest that p53 mutations are associated with resistance to platinum-based therapies in NSCLC, data on p53 IHC positivity are equivocal. To date, resistance-modulating strategies have generally not proven clinically useful in lung cancer, although small randomized trials suggest a modest benefit of verapamil and related agents in NSCLC. PMID:20047843

  15. TUMOR AND HOST FACTORS THAT MAY LIMIT EFFICACY OF CHEMOTHERAPY IN NON-SMALL CELL AND SMALL CELL LUNG CANCER

    PubMed Central

    Stewart, David J.

    2010-01-01

    expression of GST-pi, RB or p27kip1 may possibly confer resistance. While limited clinical data suggest that p53 mutations are associated with resistance to platinum-based therapies in NSCLC, data on p53 IHC positivity are equivocal. To date, resistance-modulating strategies have generally not proven clinically useful in lung cancer, although small randomized trials suggest a modest benefit of verapamil and related agents in NSCLC. PMID:20047843

  16. Alisertib induces cell cycle arrest and autophagy and suppresses epithelial-to-mesenchymal transition involving PI3K/Akt/mTOR and sirtuin 1-mediated signaling pathways in human pancreatic cancer cells.

    PubMed

    Wang, Feng; Li, Hai; Yan, Xiao-Gang; Zhou, Zhi-Wei; Yi, Zhi-Gang; He, Zhi-Xu; Pan, Shu-Ting; Yang, Yin-Xue; Wang, Zuo-Zheng; Zhang, Xueji; Yang, Tianxing; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    Pancreatic cancer is the most aggressive cancer worldwide with poor response to current therapeutics. Alisertib (ALS), a potent and selective Aurora kinase A inhibitor, exhibits potent anticancer effects in preclinical and clinical studies; however, the effect and underlying mechanism of ALS in the pancreatic cancer treatment remain elusive. This study aimed to examine the effects of ALS on cell growth, autophagy, and epithelial-to-mesenchymal transition (EMT) and to delineate the possible molecular mechanisms in human pancreatic cancer PANC-1 and BxPC-3 cells. The results showed that ALS exerted potent cell growth inhibitory, pro-autophagic, and EMT-suppressing effects in PANC-1 and BxPC-3 cells. ALS remarkably arrested PANC-1 and BxPC-3 cells in G2/M phase via regulating the expression of cyclin-dependent kinases 1 and 2, cyclin B1, cyclin D1, p21 Waf1/Cip1, p27 Kip1, and p53. ALS concentration-dependently induced autophagy in PANC-1 and BxPC-3 cells, which may be attributed to the inhibition of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR), p38 mitogen-activated protein kinase (p38 MAPK), and extracellular signal-regulated kinases 1 and 2 (Erk1/2) but activation of 5'-AMP-dependent kinase signaling pathways. ALS significantly inhibited EMT in PANC-1 and BxPC-3 cells with an increase in the expression of E-cadherin and a decrease in N-cadherin. In addition, ALS suppressed the expression of sirtuin 1 (Sirt1) and pre-B cell colony-enhancing factor/visfatin in both cell lines with a rise in the level of acetylated p53. These findings show that ALS induces cell cycle arrest and promotes autophagic cell death but inhibits EMT in pancreatic cancer cells with the involvement of PI3K/Akt/mTOR, p38 MAPK, Erk1/2, and Sirt1-mediated signaling pathways. Taken together, ALS may represent a promising anticancer drug for pancreatic cancer treatment. More studies are warranted to investigate other molecular targets and

  17. Analysis of cancer genomes reveals basic features of human aging and its role in cancer development

    PubMed Central

    Podolskiy, Dmitriy I.; Lobanov, Alexei V.; Kryukov, Gregory V.; Gladyshev, Vadim N.

    2016-01-01

    Somatic mutations have long been implicated in aging and disease, but their impact on fitness and function is difficult to assess. Here by analysing human cancer genomes we identify mutational patterns associated with aging. Our analyses suggest that age-associated mutation load and burden double approximately every 8 years, similar to the all-cause mortality doubling time. This analysis further reveals variance in the rate of aging among different human tissues, for example, slightly accelerated aging of the reproductive system. Age-adjusted mutation load and burden correlate with the corresponding cancer incidence and precede it on average by 15 years, pointing to pre-clinical cancer development times. Behaviour of mutation load also exhibits gender differences and late-life reversals, explaining some gender-specific and late-life patterns in cancer incidence rates. Overall, this study characterizes some features of human aging and offers a mechanism for age being a risk factor for the onset of cancer. PMID:27515585

  18. Analysis of cancer genomes reveals basic features of human aging and its role in cancer development.

    PubMed

    Podolskiy, Dmitriy I; Lobanov, Alexei V; Kryukov, Gregory V; Gladyshev, Vadim N

    2016-01-01

    Somatic mutations have long been implicated in aging and disease, but their impact on fitness and function is difficult to assess. Here by analysing human cancer genomes we identify mutational patterns associated with aging. Our analyses suggest that age-associated mutation load and burden double approximately every 8 years, similar to the all-cause mortality doubling time. This analysis further reveals variance in the rate of aging among different human tissues, for example, slightly accelerated aging of the reproductive system. Age-adjusted mutation load and burden correlate with the corresponding cancer incidence and precede it on average by 15 years, pointing to pre-clinical cancer development times. Behaviour of mutation load also exhibits gender differences and late-life reversals, explaining some gender-specific and late-life patterns in cancer incidence rates. Overall, this study characterizes some features of human aging and offers a mechanism for age being a risk factor for the onset of cancer. PMID:27515585

  19. Hierarchical Clustering of Breast Cancer Methylomes Revealed Differentially Methylated and Expressed Breast Cancer Genes

    PubMed Central

    Lin, I-Hsuan; Chen, Dow-Tien; Chang, Yi-Feng; Lee, Yu-Ling; Su, Chia-Hsin; Cheng, Ching; Tsai, Yi-Chien; Ng, Swee-Chuan; Chen, Hsiao-Tan; Lee, Mei-Chen; Chen, Hong-Wei; Suen, Shih-Hui; Chen, Yu-Cheng; Liu, Tze-Tze; Chang, Chuan-Hsiung; Hsu, Ming-Ta

    2015-01-01

    Oncogenic transformation of normal cells often involves epigenetic alterations, including histone modification and DNA methylation. We conducted whole-genome bisulfite sequencing to determine the DNA methylomes of normal breast, fibroadenoma, invasive ductal carcinomas and MCF7. The emergence, disappearance, expansion and contraction of kilobase-sized hypomethylated regions (HMRs) and the hypomethylation of the megabase-sized partially methylated domains (PMDs) are the major forms of methylation changes observed in breast tumor samples. Hierarchical clustering of HMR revealed tumor-specific hypermethylated clusters and differential methylated enhancers specific to normal or breast cancer cell lines. Joint analysis of gene expression and DNA methylation data of normal breast and breast cancer cells identified differentially methylated and expressed genes associated with breast and/or ovarian cancers in cancer-specific HMR clusters. Furthermore, aberrant patterns of X-chromosome inactivation (XCI) was found in breast cancer cell lines as well as breast tumor samples in the TCGA BRCA (breast invasive carcinoma) dataset. They were characterized with differentially hypermethylated XIST promoter, reduced expression of XIST, and over-expression of hypomethylated X-linked genes. High expressions of these genes were significantly associated with lower survival rates in breast cancer patients. Comprehensive analysis of the normal and breast tumor methylomes suggests selective targeting of DNA methylation changes during breast cancer progression. The weak causal relationship between DNA methylation and gene expression observed in this study is evident of more complex role of DNA methylation in the regulation of gene expression in human epigenetics that deserves further investigation. PMID:25706888

  20. Cancer of Unknown Primary Finally Revealed to Be a Metastatic Prostate Cancer: A Case Report

    PubMed Central

    Cho, Jung Yeon; Shim, Eun Jin; Kim, In Seon; Nam, Eun Mi; Choi, Moon Young; Lee, Kyung Eun; Mun, Yeung Chul; Seoung, Chu Myoung; Song, Dong Eun; Han, Woon Sup

    2009-01-01

    The vast majority of patients with metastatic prostate cancer present with bone metastases and high prostate specific antigen (PSA) level. Rarely, prostate cancer can develop in patients with normal PSA level. Here, we report a patient who presented with a periureteral tumor of unknown primary site that was confirmed as prostate adenocarcinoma after three years with using specific immunohistochemical examination. A 64-year old man was admitted to our hospital with left flank pain associated with masses on the left pelvic cavity with left hydronephrosis. All tumor markers including CEA, CA19-9, and PSA were within the normal range. After an exploratory mass excision and left nephrectomy, the pelvic mass was diagnosed as poorly differentiated carcinoma without specific positive immunohistochemical markers. At that time, we treated him as having a cancer of unknown primary site. After approximately three years later, he revisited the hospital with a complaint of right shoulder pain. A right scapular mass was newly detected with a high serum PSA level (101.7 ng/ml). Tissues from the scapular mass and prostate revealed prostate cancer with positive immunoreactivity for P504S, a new prostate cancer-specific gene. The histological findings were the same as the previous pelvic mass; however, positive staining for PSA was observed only in the prostate mass. This case demonstrates a patient with prostate cancer and negative serological test and tissue staining that turned out to be positive during progression. We suggest the usefulness of newly developed immunohistochemical markers such as P504S to determine the specific primary site of metastatic poorly differentiated adenocarcinoma in men. PMID:19688071

  1. miR-340 inhibits tumor cell proliferation and induces apoptosis by targeting multiple negative regulators of p27 in non-small cell lung cancer

    PubMed Central

    Fernandez, Serena; Risolino, Maurizio; Mandia, Nadia; Talotta, Francesco; Soini, Ylermi; Incoronato, Mariarosaria; Condorelli, Gerolama; Banfi, Sandro; Verde, Pasquale

    2014-01-01

    MicroRNAs (miRNAs) control cell cycle progression by targeting the transcripts encoding for cyclins, CDKs and CDK inhibitors, such as p27KIP1 (p27). p27 expression is controlled by multiple transcriptional and posttranscriptional mechanisms, including translational inhibition by miR-221/222 and posttranslational regulation by the SCFSKP2 complex. The oncosuppressor activity of miR-340 has been recently characterized in breast, colorectal and osteosarcoma tumor cells. However, the mechanisms underlying miR-340-induced cell growth arrest have not been elucidated. Here we describe miR-340 as a novel tumor suppressor in non-small cell lung cancer (NSCLC). Starting from the observation that the growth-inhibitory and proapoptotic effects of miR-340 correlate with the accumulation of p27 in lung adenocarcinoma and glioblastoma cells, we have analyzed the functional relationship between miR-340 and p27 expression. miR-340 targets three key negative regulators of p27. The miR-340-mediated inhibition of both Pumilio-family RNA-binding proteins (PUM1 and PUM2), required for the miR-221/222 interaction with the p27 3′UTR, antagonizes the miRNA-dependent downregulation of p27. At the same time, miR-340 induces the stabilization of p27 by targeting SKP2, the key posttranslational regulator of p27. Therefore, miR-340 controls p27 at both translational and posttranslational levels. Accordingly, the inhibition of either PUM1 or SKP2 partially recapitulates the miR-340 effect on cell proliferation and apoptosis. In addition to the effect on tumor cell proliferation, miR-340 also inhibits intercellular adhesion and motility in lung cancer cells. These changes correlate with the miR-340-mediated inhibition of previously validated (MET and ROCK1) and potentially novel (RHOA and CDH1) miR-340 target transcripts. Finally, we show that in a small cohort of NSCLC patients (n=23), representative of all four stages of lung cancer, miR-340 expression inversely correlates with clinical

  2. miR-340 inhibits tumor cell proliferation and induces apoptosis by targeting multiple negative regulators of p27 in non-small cell lung cancer.

    PubMed

    Fernandez, S; Risolino, M; Mandia, N; Talotta, F; Soini, Y; Incoronato, M; Condorelli, G; Banfi, S; Verde, P

    2015-06-01

    MicroRNAs (miRNAs) control cell cycle progression by targeting the transcripts encoding for cyclins, CDKs and CDK inhibitors, such as p27(KIP1) (p27). p27 expression is controlled by multiple transcriptional and posttranscriptional mechanisms, including translational inhibition by miR-221/222 and posttranslational regulation by the SCF(SKP2) complex. The oncosuppressor activity of miR-340 has been recently characterized in breast, colorectal and osteosarcoma tumor cells. However, the mechanisms underlying miR-340-induced cell growth arrest have not been elucidated. Here, we describe miR-340 as a novel tumor suppressor in non-small cell lung cancer (NSCLC). Starting from the observation that the growth-inhibitory and proapoptotic effects of miR-340 correlate with the accumulation of p27 in lung adenocarcinoma and glioblastoma cells, we have analyzed the functional relationship between miR-340 and p27 expression. miR-340 targets three key negative regulators of p27. The miR-340-mediated inhibition of both Pumilio family RNA-binding proteins (PUM1 and PUM2), required for the miR-221/222 interaction with the p27 3'-UTR, antagonizes the miRNA-dependent downregulation of p27. At the same time, miR-340 induces the stabilization of p27 by targeting SKP2, the key posttranslational regulator of p27. Therefore, miR-340 controls p27 at both translational and posttranslational levels. Accordingly, the inhibition of either PUM1 or SKP2 partially recapitulates the miR-340 effect on cell proliferation and apoptosis. In addition to the effect on tumor cell proliferation, miR-340 also inhibits intercellular adhesion and motility in lung cancer cells. These changes correlate with the miR-340-mediated inhibition of previously validated (MET and ROCK1) and potentially novel (RHOA and CDH1) miR-340 target transcripts. Finally, we show that in a small cohort of NSCLC patients (n=23), representative of all four stages of lung cancer, miR-340 expression inversely correlates with clinical

  3. TCGA bladder cancer study reveals potential drug targets

    Cancer.gov

    Investigators with TCGA have identified new potential therapeutic targets for a major form of bladder cancer, including important genes and pathways that are disrupted in the disease. They also discovered that, at the molecular level, some subtypes of bla

  4. Revealing the Complexity of Breast Cancer by Next Generation Sequencing

    PubMed Central

    Verigos, John; Magklara, Angeliki

    2015-01-01

    Over the last few years the increasing usage of “-omic” platforms, supported by next-generation sequencing, in the analysis of breast cancer samples has tremendously advanced our understanding of the disease. New driver and passenger mutations, rare chromosomal rearrangements and other genomic aberrations identified by whole genome and exome sequencing are providing missing pieces of the genomic architecture of breast cancer. High resolution maps of breast cancer methylomes and sequencing of the miRNA microworld are beginning to paint the epigenomic landscape of the disease. Transcriptomic profiling is giving us a glimpse into the gene regulatory networks that govern the fate of the breast cancer cell. At the same time, integrative analysis of sequencing data confirms an extensive intertumor and intratumor heterogeneity and plasticity in breast cancer arguing for a new approach to the problem. In this review, we report on the latest findings on the molecular characterization of breast cancer using NGS technologies, and we discuss their potential implications for the improvement of existing therapies. PMID:26561834

  5. Biomolecular Events in Cancer Revealed by Attractor Metagenes

    PubMed Central

    Cheng, Wei-Yi; Yang, Tai-Hsien Ou; Anastassiou, Dimitris

    2013-01-01

    Mining gene expression profiles has proven valuable for identifying signatures serving as surrogates of cancer phenotypes. However, the similarities of such signatures across different cancer types have not been strong enough to conclude that they represent a universal biological mechanism shared among multiple cancer types. Here we present a computational method for generating signatures using an iterative process that converges to one of several precise attractors defining signatures representing biomolecular events, such as cell transdifferentiation or the presence of an amplicon. By analyzing rich gene expression datasets from different cancer types, we identified several such biomolecular events, some of which are universally present in all tested cancer types in nearly identical form. Although the method is unsupervised, we show that it often leads to attractors with strong phenotypic associations. We present several such multi-cancer attractors, focusing on three that are prominent and sharply defined in all cases: a mesenchymal transition attractor strongly associated with tumor stage, a mitotic chromosomal instability attractor strongly associated with tumor grade, and a lymphocyte-specific attractor. PMID:23468608

  6. Genomic Interaction Profiles in Breast Cancer Reveal Altered Chromatin Architecture

    PubMed Central

    Zeitz, Michael J.; Ay, Ferhat; Heidmann, Julia D.; Lerner, Paula L.

    2013-01-01

    Gene transcription can be regulated by remote enhancer regions through chromosome looping either in cis or in trans. Cancer cells are characterized by wholesale changes in long-range gene interactions, but the role that these long-range interactions play in cancer progression and metastasis is not well understood. In this study, we used IGFBP3, a gene involved in breast cancer pathogenesis, as bait in a 4C-seq experiment comparing normal breast cells (HMEC) with two breast cancer cell lines (MCF7, an ER positive cell line, and MDA-MB-231, a triple negative cell line). The IGFBP3 long-range interaction profile was substantially altered in breast cancer. Many interactions seen in normal breast cells are lost and novel interactions appear in cancer lines. We found that in HMEC, the breast carcinoma amplified sequence gene family (BCAS) 1–4 were among the top 10 most significantly enriched regions of interaction with IGFBP3. 3D-FISH analysis indicated that the translocation-prone BCAS genes, which are located on chromosomes 1, 17, and 20, are in close physical proximity with IGFBP3 and each other in normal breast cells. We also found that epidermal growth factor receptor (EGFR), a gene implicated in tumorigenesis, interacts significantly with IGFBP3 and that this interaction may play a role in their regulation. Breakpoint analysis suggests that when an IGFBP3 interacting region undergoes a translocation an additional interaction detectable by 4C is gained. Overall, our data from multiple lines of evidence suggest an important role for long-range chromosomal interactions in the pathogenesis of cancer. PMID:24019942

  7. Subclonal diversification of primary breast cancer revealed by multiregion sequencing

    DOE PAGESBeta

    Yates, Lucy R.; Gerstung, Moritz; Knappskog, Stian; Desmedt, Christine; Gundem, Gunes; Van Loo, Peter; Aas, Turid; Alexandrov, Ludmil B.; Larsimont, Denis; Davies, Helen; et al

    2015-06-22

    Sequencing cancer genomes may enable tailoring of therapeutics to the underlying biological abnormalities driving a particular patient's tumor. However, sequencing-based strategies rely heavily on representative sampling of tumors. To understand the subclonal structure of primary breast cancer, we applied whole-genome and targeted sequencing to multiple samples from each of 50 patients' tumors (303 samples in total). The extent of subclonal diversification varied among cases and followed spatial patterns. No strict temporal order was evident, with point mutations and rearrangements affecting the most common breast cancer genes, including PIK3CA, TP53, PTEN, BRCA2 and MYC, occurring early in some tumors and latemore » in others. In 13 out of 50 cancers, potentially targetable mutations were subclonal. Landmarks of disease progression, such as resistance to chemotherapy and the acquisition of invasive or metastatic potential, arose within detectable subclones of antecedent lesions. These findings highlight the importance of including analyses of subclonal structure and tumor evolution in clinical trials of primary breast cancer.« less

  8. Subclonal diversification of primary breast cancer revealed by multiregion sequencing

    SciTech Connect

    Yates, Lucy R.; Gerstung, Moritz; Knappskog, Stian; Desmedt, Christine; Gundem, Gunes; Van Loo, Peter; Aas, Turid; Alexandrov, Ludmil B.; Larsimont, Denis; Davies, Helen; Li, Yilong; Ju, Young Seok; Ramakrishna, Manasa; Haugland, Hans Kristian; Lilleng, Peer Kaare; Nik-Zainal, Serena; McLaren, Stuart; Butler, Adam; Martin, Sancha; Glodzik, Dominic; Menzies, Andrew; Raine, Keiran; Hinton, Jonathan; Jones, David; Mudie, Laura J.; Jiang, Bing; Vincent, Delphine; Greene-Colozzi, April; Adnet, Pierre -Yves; Fatima, Aquila; Maetens, Marion; Ignatiadis, Michail; Stratton, Michael R.; Sotiriou, Christos; Richardson, Andrea L.; Lønning, Per Eystein; Wedge, David C.; Campbell, Peter J.

    2015-06-22

    Sequencing cancer genomes may enable tailoring of therapeutics to the underlying biological abnormalities driving a particular patient's tumor. However, sequencing-based strategies rely heavily on representative sampling of tumors. To understand the subclonal structure of primary breast cancer, we applied whole-genome and targeted sequencing to multiple samples from each of 50 patients' tumors (303 samples in total). The extent of subclonal diversification varied among cases and followed spatial patterns. No strict temporal order was evident, with point mutations and rearrangements affecting the most common breast cancer genes, including PIK3CA, TP53, PTEN, BRCA2 and MYC, occurring early in some tumors and late in others. In 13 out of 50 cancers, potentially targetable mutations were subclonal. Landmarks of disease progression, such as resistance to chemotherapy and the acquisition of invasive or metastatic potential, arose within detectable subclones of antecedent lesions. These findings highlight the importance of including analyses of subclonal structure and tumor evolution in clinical trials of primary breast cancer.

  9. Danusertib, a potent pan-Aurora kinase and ABL kinase inhibitor, induces cell cycle arrest and programmed cell death and inhibits epithelial to mesenchymal transition involving the PI3K/Akt/mTOR-mediated signaling pathway in human gastric cancer AGS and NCI-N78 cells

    PubMed Central

    Yuan, Chun-Xiu; Zhou, Zhi-Wei; Yang, Yin-Xue; He, Zhi-Xu; Zhang, Xueji; Wang, Dong; Yang, Tianxing; Pan, Si-Yuan; Chen, Xiao-Wu; Zhou, Shu-Feng

    2015-01-01

    Gastric cancer is the second leading cause of cancer-related death worldwide, with a poor response to current chemotherapy. Danusertib is a pan-inhibitor of the Aurora kinases and a third-generation Bcr-Abl tyrosine kinase inhibitor with potent anticancer effects, but its antitumor effect and underlying mechanisms in the treatment of human gastric cancer are unknown. This study aimed to investigate the effects of danusertib on cell growth, apoptosis, autophagy, and epithelial to mesenchymal transition and the molecular mechanisms involved in human gastric cancer AGS and NCI-N78 cells. The results showed that danusertib had potent growth-inhibitory, apoptosis-inducing, and autophagy-inducing effects on AGS and NCI-N78 cells. Danusertib arrested AGS and NCI-N78 cells in G2/M phase, with downregulation of expression of cyclin B1 and cyclin-dependent kinase 1 and upregulation of expression of p21 Waf1/Cip1, p27 Kip1, and p53. Danusertib induced mitochondria-mediated apoptosis, with an increase in expression of proapoptotic protein and a decrease in antiapoptotic proteins in both cell lines. Danusertib induced release of cytochrome c from the mitochondria to the cytosol and triggered activation of caspase 9 and caspase 3 in AGS and NCI-N78 cells. Further, danusertib induced autophagy, with an increase in expression of beclin 1 and conversion of microtubule-associated protein 1A/1B-light chain 3 (LC3-I) to LC3-II in both cell lines. Inhibition of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) and p38 mitogen-activated protein kinase pathways as well as activation of 5′ AMP-activated protein kinase contributed to the proautophagic effect of danusertib in AGS and NCI-N78 cells. SB202191 and wortmannin enhanced the autophagy-inducing effect of danusertib in AGS and NCI-N78 cells. In addition, danusertib inhibited epithelial to mesenchymal transition with an increase in expression of E-cadherin and a decrease in expression

  10. Neurotropic and neuroprotective activities of the earthworm peptide Lumbricusin.

    PubMed

    Kim, Dae Hong; Lee, Ik Hwan; Nam, Seung Taek; Hong, Ji; Zhang, Peng; Hwang, Jae Sam; Seok, Heon; Choi, Hyemin; Lee, Dong Gun; Kim, Jae Il; Kim, Ho

    2014-06-01

    We recently isolated a polypeptide from the earthworm Lumbricus terrestris that is structurally similar to defensin, a well-known antibacterial peptide. An 11-mer antibacterial peptide (NH2-RNRRWCIDQQA), designated Lumbricusin, was synthesized based on the amino acid sequence of the isolated polypeptide. Since we previously reported that CopA3, a dung beetle peptide, enhanced neuronal cell proliferation, we here examined whether Lumbricusin exerted neurotropic and/or neuroprotective effects. Lumbricusin treatment induced a time-dependent increase (∼51%) in the proliferation of human neuroblastoma SH-SY5Y cells. Lumbricusin also significantly inhibited the apoptosis and decreased viability induced by treatment with 6-hydroxy dopamine, a Parkinson's disease-mimicking agent. Immunoblot analyses revealed that Lumbricusin treatment increased ubiquitination of p27(Kip1) protein, a negative regulator of cell-cycle progression, in SH-SY5Y cells, and markedly promoted its degradation. Notably, adenoviral-mediated over-expression of p27(Kip1) significantly blocked the antiapoptotic effect of Lumbricusin in 6-hydroxy dopamine-treated SH-SY5Y cells. These results suggest that promotion of p27(Kip1) degradation may be the main mechanism underlying the neuroprotective and neurotropic effects of Lumbricusin. PMID:24796676

  11. Pancreatic cancer genomes reveal aberrations in axon guidance pathway genes

    PubMed Central

    Biankin, Andrew V.; Waddell, Nicola; Kassahn, Karin S.; Gingras, Marie-Claude; Muthuswamy, Lakshmi B.; Johns, Amber L.; Miller, David K.; Wilson, Peter J.; Patch, Ann-Marie; Wu, Jianmin; Chang, David K.; Cowley, Mark J.; Gardiner, Brooke B.; Song, Sarah; Harliwong, Ivon; Idrisoglu, Senel; Nourse, Craig; Nourbakhsh, Ehsan; Manning, Suzanne; Wani, Shivangi; Gongora, Milena; Pajic, Marina; Scarlett, Christopher J.; Gill, Anthony J.; Pinho, Andreia V.; Rooman, Ilse; Anderson, Matthew; Holmes, Oliver; Leonard, Conrad; Taylor, Darrin; Wood, Scott; Xu, Qinying; Nones, Katia; Fink, J. Lynn; Christ, Angelika; Bruxner, Tim; Cloonan, Nicole; Kolle, Gabriel; Newell, Felicity; Pinese, Mark; Mead, R. Scott; Humphris, Jeremy L.; Kaplan, Warren; Jones, Marc D.; Colvin, Emily K.; Nagrial, Adnan M.; Humphrey, Emily S.; Chou, Angela; Chin, Venessa T.; Chantrill, Lorraine A.; Mawson, Amanda; Samra, Jaswinder S.; Kench, James G.; Lovell, Jessica A.; Daly, Roger J.; Merrett, Neil D.; Toon, Christopher; Epari, Krishna; Nguyen, Nam Q.; Barbour, Andrew; Zeps, Nikolajs; Kakkar, Nipun; Zhao, Fengmei; Wu, Yuan Qing; Wang, Min; Muzny, Donna M.; Fisher, William E.; Brunicardi, F. Charles; Hodges, Sally E.; Reid, Jeffrey G.; Drummond, Jennifer; Chang, Kyle; Han, Yi; Lewis, Lora R.; Dinh, Huyen; Buhay, Christian J.; Beck, Timothy; Timms, Lee; Sam, Michelle; Begley, Kimberly; Brown, Andrew; Pai, Deepa; Panchal, Ami; Buchner, Nicholas; De Borja, Richard; Denroche, Robert E.; Yung, Christina K.; Serra, Stefano; Onetto, Nicole; Mukhopadhyay, Debabrata; Tsao, Ming-Sound; Shaw, Patricia A.; Petersen, Gloria M.; Gallinger, Steven; Hruban, Ralph H.; Maitra, Anirban; Iacobuzio-Donahue, Christine A.; Schulick, Richard D.; Wolfgang, Christopher L.; Morgan, Richard A.; Lawlor, Rita T.; Capelli, Paola; Corbo, Vincenzo; Scardoni, Maria; Tortora, Giampaolo; Tempero, Margaret A.; Mann, Karen M.; Jenkins, Nancy A.; Perez-Mancera, Pedro A.; Adams, David J.; Largaespada, David A.; Wessels, Lodewyk F. A.; Rust, Alistair G.; Stein, Lincoln D.; Tuveson, David A.; Copeland, Neal G.; Musgrove, Elizabeth A.; Scarpa, Aldo; Eshleman, James R.; Hudson, Thomas J.; Sutherland, Robert L.; Wheeler, David A.; Pearson, John V.; McPherson, John D.; Gibbs, Richard A.; Grimmond, Sean M.

    2012-01-01

    Pancreatic cancer is a highly lethal malignancy with few effective therapies. We performed exome sequencing and copy number analysis to define genomic aberrations in a prospectively accrued clinical cohort (n = 142) of early (stage I and II) sporadic pancreatic ductal adenocarcinoma. Detailed analysis of 99 informative tumours identified substantial heterogeneity with 2,016 non-silent mutations and 1,628 copy-number variations. We define 16 significantly mutated genes, reaffirming known mutations (KRAS, TP53, CDKN2A, SMAD4, MLL3, TGFBR2, ARID1A and SF3B1), and uncover novel mutated genes including additional genes involved in chromatin modification (EPC1 and ARID2), DNA damage repair (ATM) and other mechanisms (ZIM2, MAP2K4, NALCN, SLC16A4 and MAGEA6). Integrative analysis with in vitro functional data and animal models provided supportive evidence for potential roles for these genetic aberrations in carcinogenesis. Pathway-based analysis of recurrently mutated genes recapitulated clustering in core signalling pathways in pancreatic ductal adenocarcinoma, and identified new mutated genes in each pathway. We also identified frequent and diverse somatic aberrations in genes described traditionally as embryonic regulators of axon guidance, particularly SLIT/ROBO signalling, which was also evident in murine Sleeping Beauty transposon-mediated somatic mutagenesis models of pancreatic cancer, providing further supportive evidence for the potential involvement of axon guidance genes in pancreatic carcinogenesis. PMID:23103869

  12. Sleeping Beauty screen reveals Pparg activation in metastatic prostate cancer.

    PubMed

    Ahmad, Imran; Mui, Ernest; Galbraith, Laura; Patel, Rachana; Tan, Ee Hong; Salji, Mark; Rust, Alistair G; Repiscak, Peter; Hedley, Ann; Markert, Elke; Loveridge, Carolyn; van der Weyden, Louise; Edwards, Joanne; Sansom, Owen J; Adams, David J; Leung, Hing Y

    2016-07-19

    Prostate cancer (CaP) is the most common adult male cancer in the developed world. The paucity of biomarkers to predict prostate tumor biology makes it important to identify key pathways that confer poor prognosis and guide potential targeted therapy. Using a murine forward mutagenesis screen in a Pten-null background, we identified peroxisome proliferator-activated receptor gamma (Pparg), encoding a ligand-activated transcription factor, as a promoter of metastatic CaP through activation of lipid signaling pathways, including up-regulation of lipid synthesis enzymes [fatty acid synthase (FASN), acetyl-CoA carboxylase (ACC), ATP citrate lyase (ACLY)]. Importantly, inhibition of PPARG suppressed tumor growth in vivo, with down-regulation of the lipid synthesis program. We show that elevated levels of PPARG strongly correlate with elevation of FASN in human CaP and that high levels of PPARG/FASN and PI3K/pAKT pathway activation confer a poor prognosis. These data suggest that CaP patients could be stratified in terms of PPARG/FASN and PTEN levels to identify patients with aggressive CaP who may respond favorably to PPARG/FASN inhibition. PMID:27357679

  13. Sleeping Beauty screen reveals Pparg activation in metastatic prostate cancer

    PubMed Central

    Ahmad, Imran; Mui, Ernest; Galbraith, Laura; Patel, Rachana; Tan, Ee Hong; Salji, Mark; Rust, Alistair G.; Repiscak, Peter; Hedley, Ann; Markert, Elke; Loveridge, Carolyn; van der Weyden, Louise; Edwards, Joanne; Sansom, Owen J.; Adams, David J.; Leung, Hing Y.

    2016-01-01

    Prostate cancer (CaP) is the most common adult male cancer in the developed world. The paucity of biomarkers to predict prostate tumor biology makes it important to identify key pathways that confer poor prognosis and guide potential targeted therapy. Using a murine forward mutagenesis screen in a Pten-null background, we identified peroxisome proliferator-activated receptor gamma (Pparg), encoding a ligand-activated transcription factor, as a promoter of metastatic CaP through activation of lipid signaling pathways, including up-regulation of lipid synthesis enzymes [fatty acid synthase (FASN), acetyl-CoA carboxylase (ACC), ATP citrate lyase (ACLY)]. Importantly, inhibition of PPARG suppressed tumor growth in vivo, with down-regulation of the lipid synthesis program. We show that elevated levels of PPARG strongly correlate with elevation of FASN in human CaP and that high levels of PPARG/FASN and PI3K/pAKT pathway activation confer a poor prognosis. These data suggest that CaP patients could be stratified in terms of PPARG/FASN and PTEN levels to identify patients with aggressive CaP who may respond favorably to PPARG/FASN inhibition. PMID:27357679

  14. Intravital imaging reveals new ancillary mechanisms co-opted by cancer cells to drive tumor progression.

    PubMed

    Vennin, Claire; Herrmann, David; Lucas, Morghan C; Timpson, Paul

    2016-01-01

    Intravital imaging is providing new insights into the dynamics of tumor progression in native tissues and has started to reveal the layers of complexity found in cancer. Recent advances in intravital imaging have allowed us to look deeper into cancer behavior and to dissect the interactions between tumor cells and the ancillary host niche that promote cancer development. In this review, we provide an insight into the latest advances in cancer biology achieved by intravital imaging, focusing on recently discovered mechanisms by which tumor cells manipulate normal tissue to facilitate disease progression. PMID:27239290

  15. Intravital imaging reveals new ancillary mechanisms co-opted by cancer cells to drive tumor progression

    PubMed Central

    Lucas, Morghan C.; Timpson, Paul

    2016-01-01

    Intravital imaging is providing new insights into the dynamics of tumor progression in native tissues and has started to reveal the layers of complexity found in cancer. Recent advances in intravital imaging have allowed us to look deeper into cancer behavior and to dissect the interactions between tumor cells and the ancillary host niche that promote cancer development. In this review, we provide an insight into the latest advances in cancer biology achieved by intravital imaging, focusing on recently discovered mechanisms by which tumor cells manipulate normal tissue to facilitate disease progression. PMID:27239290

  16. What can digital transcript profiling reveal about human cancers?

    PubMed

    Cerutti, J M; Riggins, G J; de Souza, S J

    2003-08-01

    Important biological and clinical features of malignancy are reflected in its transcript pattern. Recent advances in gene expression technology and informatics have provided a powerful new means to obtain and interpret these expression patterns. A comprehensive approach to expression profiling is serial analysis of gene expression (SAGE), which provides digital information on transcript levels. SAGE works by counting transcripts and storing these digital values electronically, providing absolute gene expression levels that make historical comparisons possible. SAGE produces a comprehensive profile of gene expression and can be used to search for candidate tumor markers or antigens in a limited number of samples. The Cancer Genome Anatomy Project has created a SAGE database of human gene expression levels for many different tumors and normal reference tissues and provides online tools for viewing, comparing, and downloading expression profiles. Digital expression profiling using SAGE and informatics have been useful for identifying genes that have a role in tumor invasion and other aspects of tumor progression. PMID:12886451

  17. Targeted cancer exome sequencing reveals recurrent mutations in myeloproliferative neoplasms.

    PubMed

    Tenedini, E; Bernardis, I; Artusi, V; Artuso, L; Roncaglia, E; Guglielmelli, P; Pieri, L; Bogani, C; Biamonte, F; Rotunno, G; Mannarelli, C; Bianchi, E; Pancrazzi, A; Fanelli, T; Malagoli Tagliazucchi, G; Ferrari, S; Manfredini, R; Vannucchi, A M; Tagliafico, E

    2014-05-01

    With the intent of dissecting the molecular complexity of Philadelphia-negative myeloproliferative neoplasms (MPN), we designed a target enrichment panel to explore, using next-generation sequencing (NGS), the mutational status of an extensive list of 2000 cancer-associated genes and microRNAs. The genomic DNA of granulocytes and in vitro-expanded CD3+T-lymphocytes, as a germline control, was target-enriched and sequenced in a learning cohort of 20 MPN patients using Roche 454 technology. We identified 141 genuine somatic mutations, most of which were not previously described. To test the frequency of the identified variants, a larger validation cohort of 189 MPN patients was additionally screened for these mutations using Ion Torrent AmpliSeq NGS. Excluding the genes already described in MPN, for 8 genes (SCRIB, MIR662, BARD1, TCF12, FAT4, DAP3, POLG and NRAS), we demonstrated a mutation frequency between 3 and 8%. We also found that mutations at codon 12 of NRAS (NRASG12V and NRASG12D) were significantly associated, for primary myelofibrosis (PMF), with highest dynamic international prognostic scoring system (DIPSS)-plus score categories. This association was then confirmed in 66 additional PMF patients composing a final dataset of 168 PMF showing a NRAS mutation frequency of 4.7%, which was associated with a worse outcome, as defined by the DIPSS plus score. PMID:24150215

  18. Integrative analyses reveal signaling pathways underlying familial breast cancer susceptibility.

    PubMed

    Piccolo, Stephen R; Hoffman, Laura M; Conner, Thomas; Shrestha, Gajendra; Cohen, Adam L; Marks, Jeffrey R; Neumayer, Leigh A; Agarwal, Cori A; Beckerle, Mary C; Andrulis, Irene L; Spira, Avrum E; Moos, Philip J; Buys, Saundra S; Johnson, William Evan; Bild, Andrea H

    2016-03-01

    The signaling events that drive familial breast cancer (FBC) risk remain poorly understood. While the majority of genomic studies have focused on genetic risk variants, known risk variants account for at most 30% of FBC cases. Considering that multiple genes may influence FBC risk, we hypothesized that a pathway-based strategy examining different data types from multiple tissues could elucidate the biological basis for FBC. In this study, we performed integrated analyses of gene expression and exome-sequencing data from peripheral blood mononuclear cells and showed that cell adhesion pathways are significantly and consistently dysregulated in women who develop FBC. The dysregulation of cell adhesion pathways in high-risk women was also identified by pathway-based profiling applied to normal breast tissue data from two independent cohorts. The results of our genomic analyses were validated in normal primary mammary epithelial cells from high-risk and control women, using cell-based functional assays, drug-response assays, fluorescence microscopy, and Western blotting assays. Both genomic and cell-based experiments indicate that cell-cell and cell-extracellular matrix adhesion processes seem to be disrupted in non-malignant cells of women at high risk for FBC and suggest a potential role for these processes in FBC development. PMID:26969729

  19. Integrative analysis reveals disease-associated genes and biomarkers for prostate cancer progression

    PubMed Central

    2014-01-01

    Background Prostate cancer is one of the most common complex diseases with high leading cause of death in men. Identifications of prostate cancer associated genes and biomarkers are thus essential as they can gain insights into the mechanisms underlying disease progression and advancing for early diagnosis and developing effective therapies. Methods In this study, we presented an integrative analysis of gene expression profiling and protein interaction network at a systematic level to reveal candidate disease-associated genes and biomarkers for prostate cancer progression. At first, we reconstructed the human prostate cancer protein-protein interaction network (HPC-PPIN) and the network was then integrated with the prostate cancer gene expression data to identify modules related to different phases in prostate cancer. At last, the candidate module biomarkers were validated by its predictive ability of prostate cancer progression. Results Different phases-specific modules were identified for prostate cancer. Among these modules, transcription Androgen Receptor (AR) nuclear signaling and Epidermal Growth Factor Receptor (EGFR) signalling pathway were shown to be the pathway targets for prostate cancer progression. The identified candidate disease-associated genes showed better predictive ability of prostate cancer progression than those of published biomarkers. In context of functional enrichment analysis, interestingly candidate disease-associated genes were enriched in the nucleus and different functions were encoded for potential transcription factors, for examples key players as AR, Myc, ESR1 and hidden player as Sp1 which was considered as a potential novel biomarker for prostate cancer. Conclusions The successful results on prostate cancer samples demonstrated that the integrative analysis is powerful and useful approach to detect candidate disease-associate genes and modules which can be used as the potential biomarkers for prostate cancer progression. The

  20. Potential microRNA-mediated oncogenic intercellular communication revealed by pan-cancer analysis

    NASA Astrophysics Data System (ADS)

    Li, Yue; Zhang, Zhaolei

    2014-11-01

    Carcinogenesis consists of oncogenesis and metastasis, and intriguingly microRNAs (miRNAs) are involved in both processes. Although aberrant miRNA activities are prevalent in diverse tumor types, the exact mechanisms for how they regulate cancerous processes are not always clear. To this end, we performed a large-scale pan-cancer analysis via a novel probabilistic approach to infer recurrent miRNA-target interactions implicated in 12 cancer types using data from The Cancer Genome Atlas. We discovered ~20,000 recurrent miRNA regulations, which are enriched for cancer-related miRNAs/genes. Notably, miRNA 200 family (miR-200/141/429) is among the most prominent miRNA regulators, which is known to be involved in metastasis. Importantly, the recurrent miRNA regulatory network is not only enriched for cancer pathways but also for extracellular matrix (ECM) organization and ECM-receptor interactions. The results suggest an intriguing cancer mechanism involving miRNA-mediated cell-to-cell communication, which possibly involves delivery of tumorigenic miRNA messengers to adjacent cells via exosomes. Finally, survival analysis revealed 414 recurrent-prognostic associations, where both gene and miRNA involved in each interaction conferred significant prognostic power in one or more cancer types. Together, our comprehensive pan-cancer analysis provided not only biological insights into metastasis but also brought to bear the clinical relevance of the proposed recurrent miRNA-gene associations.

  1. An international survey of physician attitudes and practice in regard to revealing the diagnosis of cancer.

    PubMed

    Holland, J C; Geary, N; Marchini, A; Tross, S

    1987-01-01

    In 1984, questionnaires were sent to members of the International Psycho-Oncology Society concerning the practice in their country with regard to revealing the diagnosis of cancer to patients, their opinion about the effect of their policy, and their impression of local trends and attitudes toward cancer. Data from 90 respondents from 20 countries revealed that use of the word "cancer" was often avoided in discussions with the patient. Words commonly substituted for cancer were those that implied a "swelling" (e.g., tumor, growth, lump), and "inflammation," or a pathophysiologic change (blood disease, precancerous, unclean tissue). Oncologists estimated that a low percentage (less than 40%) of their colleagues revealed the word cancer in Africa, France, Hungary, Italy, Japan, Panama, Portugal, and Spain. Oncologists from Austria, Denmark Finland, The Netherlands, New Zealand, Norway, Sweden, and Switzerland, estimated the percentage to be high (greater than 80%). However, in all countries, the majority of physicians tell the family the diagnosis. The majority (90%) reported a trend toward increased telling of the diagnosis, due to greater patient information and expectations, and increased physician openess in using the word cancer. Most (68%) felt that the overall effect of revealing the diagnosis was positive. While emotional distress was transiently greater when patients were told, there were positive effects concerning coping, compliance, tolerance of treatment, planning for future, communication with physicians and others, and improved prognosis. The transient negative effects were depression, anxiety, and anger. It is important to recognize that efforts to find the "correct" position about revealing or concealing cancer diagnosis must recognize that the language between doctor and patient is constrained by cultural norms. Communication is likely to be far less dependent upon the specific words used then upon the meaning that is conveyed by the doctor. PMID

  2. Mutational Strand Asymmetries in Cancer Genomes Reveal Mechanisms of DNA Damage and Repair.

    PubMed

    Haradhvala, Nicholas J; Polak, Paz; Stojanov, Petar; Covington, Kyle R; Shinbrot, Eve; Hess, Julian M; Rheinbay, Esther; Kim, Jaegil; Maruvka, Yosef E; Braunstein, Lior Z; Kamburov, Atanas; Hanawalt, Philip C; Wheeler, David A; Koren, Amnon; Lawrence, Michael S; Getz, Gad

    2016-01-28

    Mutational processes constantly shape the somatic genome, leading to immunity, aging, cancer, and other diseases. When cancer is the outcome, we are afforded a glimpse into these processes by the clonal expansion of the malignant cell. Here, we characterize a less explored layer of the mutational landscape of cancer: mutational asymmetries between the two DNA strands. Analyzing whole-genome sequences of 590 tumors from 14 different cancer types, we reveal widespread asymmetries across mutagenic processes, with transcriptional ("T-class") asymmetry dominating UV-, smoking-, and liver-cancer-associated mutations and replicative ("R-class") asymmetry dominating POLE-, APOBEC-, and MSI-associated mutations. We report a striking phenomenon of transcription-coupled damage (TCD) on the non-transcribed DNA strand and provide evidence that APOBEC mutagenesis occurs on the lagging-strand template during DNA replication. As more genomes are sequenced, studying and classifying their asymmetries will illuminate the underlying biological mechanisms of DNA damage and repair. PMID:26806129

  3. A Balanced Tissue Composition Reveals New Metabolic and Gene Expression Markers in Prostate Cancer

    PubMed Central

    Tessem, May-Britt; Bertilsson, Helena; Angelsen, Anders; Bathen, Tone F.; Drabløs, Finn; Rye, Morten Beck

    2016-01-01

    Molecular analysis of patient tissue samples is essential to characterize the in vivo variability in human cancers which are not accessible in cell-lines or animal models. This applies particularly to studies of tumor metabolism. The challenge is, however, the complex mixture of various tissue types within each sample, such as benign epithelium, stroma and cancer tissue, which can introduce systematic biases when cancers are compared to normal samples. In this study we apply a simple strategy to remove such biases using sample selections where the average content of stroma tissue is balanced between the sample groups. The strategy is applied to a prostate cancer patient cohort where data from MR spectroscopy and gene expression have been collected from and integrated on the exact same tissue samples. We reveal in vivo changes in cancer-relevant metabolic pathways which are otherwise hidden in the data due to tissue confounding. In particular, lowered levels of putrescine are connected to increased expression of SRM, reduced levels of citrate are attributed to upregulation of genes promoting fatty acid synthesis, and increased succinate levels coincide with reduced expression of SUCLA2 and SDHD. In addition, the strategy also highlights important metabolic differences between the stroma, epithelium and prostate cancer. These results show that important in vivo metabolic features of cancer can be revealed from patient data only if the heterogeneous tissue composition is properly accounted for in the analysis. PMID:27100877

  4. A Balanced Tissue Composition Reveals New Metabolic and Gene Expression Markers in Prostate Cancer.

    PubMed

    Tessem, May-Britt; Bertilsson, Helena; Angelsen, Anders; Bathen, Tone F; Drabløs, Finn; Rye, Morten Beck

    2016-01-01

    Molecular analysis of patient tissue samples is essential to characterize the in vivo variability in human cancers which are not accessible in cell-lines or animal models. This applies particularly to studies of tumor metabolism. The challenge is, however, the complex mixture of various tissue types within each sample, such as benign epithelium, stroma and cancer tissue, which can introduce systematic biases when cancers are compared to normal samples. In this study we apply a simple strategy to remove such biases using sample selections where the average content of stroma tissue is balanced between the sample groups. The strategy is applied to a prostate cancer patient cohort where data from MR spectroscopy and gene expression have been collected from and integrated on the exact same tissue samples. We reveal in vivo changes in cancer-relevant metabolic pathways which are otherwise hidden in the data due to tissue confounding. In particular, lowered levels of putrescine are connected to increased expression of SRM, reduced levels of citrate are attributed to upregulation of genes promoting fatty acid synthesis, and increased succinate levels coincide with reduced expression of SUCLA2 and SDHD. In addition, the strategy also highlights important metabolic differences between the stroma, epithelium and prostate cancer. These results show that important in vivo metabolic features of cancer can be revealed from patient data only if the heterogeneous tissue composition is properly accounted for in the analysis. PMID:27100877

  5. Whole-exome sequencing combined with functional genomics reveals novel candidate driver cancer genes in endometrial cancer.

    PubMed

    Liang, Han; Cheung, Lydia W T; Li, Jie; Ju, Zhenlin; Yu, Shuangxing; Stemke-Hale, Katherine; Dogruluk, Turgut; Lu, Yiling; Liu, Xiuping; Gu, Chao; Guo, Wei; Scherer, Steven E; Carter, Hannah; Westin, Shannon N; Dyer, Mary D; Verhaak, Roeland G W; Zhang, Fan; Karchin, Rachel; Liu, Chang-Gong; Lu, Karen H; Broaddus, Russell R; Scott, Kenneth L; Hennessy, Bryan T; Mills, Gordon B

    2012-11-01

    Endometrial cancer is the most common gynecological malignancy, with more than 280,000 cases occurring annually worldwide. Although previous studies have identified important common somatic mutations in endometrial cancer, they have primarily focused on a small set of known cancer genes and have thus provided a limited view of the molecular basis underlying this disease. Here we have developed an integrated systems-biology approach to identifying novel cancer genes contributing to endometrial tumorigenesis. We first performed whole-exome sequencing on 13 endometrial cancers and matched normal samples, systematically identifying somatic alterations with high precision and sensitivity. We then combined bioinformatics prioritization with high-throughput screening (including both shRNA-mediated knockdown and expression of wild-type and mutant constructs) in a highly sensitive cell viability assay. Our results revealed 12 potential driver cancer genes including 10 tumor-suppressor candidates (ARID1A, INHBA, KMO, TTLL5, GRM8, IGFBP3, AKTIP, PHKA2, TRPS1, and WNT11) and two oncogene candidates (ERBB3 and RPS6KC1). The results in the "sensor" cell line were recapitulated by siRNA-mediated knockdown in endometrial cancer cell lines. Focusing on ARID1A, we integrated mutation profiles with functional proteomics in 222 endometrial cancer samples, demonstrating that ARID1A mutations frequently co-occur with mutations in the phosphatidylinositol 3-kinase (PI3K) pathway and are associated with PI3K pathway activation. siRNA knockdown in endometrial cancer cell lines increased AKT phosphorylation supporting ARID1A as a novel regulator of PI3K pathway activity. Our study presents the first unbiased view of somatic coding mutations in endometrial cancer and provides functional evidence for diverse driver genes and mutations in this disease. PMID:23028188

  6. Clinical next-generation sequencing reveals aggressive cancer biology in adolescent and young adult patients

    PubMed Central

    Subbiah, Vivek; Bupathi, Manojkumar; Kato, Shumei; Livingston, Andrew; Slopis, John; Anderson, Pete M.; Hong, David S.

    2015-01-01

    Background The aggressive biology of cancers arising in adolescent and young adult (AYA; ages 15–39 years) patients is thought to contribute to poor survival outcomes. Methods We used clinical next-generation sequencing (NGS) results to examine the molecular alterations and diverse biology of cancer in AYA patients referred to the Phase 1 program at UT MD Anderson Cancer Center. Results Among the 28 patients analyzed (14 female and 14 male), 12 had pediatric-type cancers, six had adult-type cancers, and ten had orphan cancers. Unique, hitherto unreported aberrations were identified in all types of cancers. Aberrations in TP53, NKX2-1, KRAS, CDKN2A, MDM4, MCL1, MYC, BCL2L2, and RB1 were demonstrated across all tumor types. Five patients harbored TP53 aberrations; three patients harbored MYC, MCL1, and CDKN2A aberrations; and two patients harbored NKX2-1, KRAS, MDM4, BCL2L2, and RB1 alterations. Several patients had multiple aberrations; a patient with wild-type gastrointestinal stromal tumor harbored five alterations (MDM4, MCL1, KIT, AKT3, and PDGRFA). Conclusions This preliminary report of NGS of cancer in AYA patients reveals diverse and unique aberrations. Further molecular profiling and a deeper understanding of the biology of these unique aberrations are warranted and may lead to targeted therapeutic interventions. PMID:26328274

  7. Quantitative proteomics reveals middle infrared radiation-interfered networks in breast cancer cells.

    PubMed

    Chang, Hsin-Yi; Li, Ming-Hua; Huang, Tsui-Chin; Hsu, Chia-Lang; Tsai, Shang-Ru; Lee, Si-Chen; Huang, Hsuan-Cheng; Juan, Hsueh-Fen

    2015-02-01

    Breast cancer is one of the leading cancer-related causes of death worldwide. Treatment of triple-negative breast cancer (TNBC) is complex and challenging, especially when metastasis has developed. In this study, we applied infrared radiation as an alternative approach for the treatment of TNBC. We used middle infrared (MIR) with a wavelength range of 3-5 μm to irradiate breast cancer cells. MIR significantly inhibited cell proliferation in several breast cancer cells but did not affect the growth of normal breast epithelial cells. We performed iTRAQ-coupled LC-MS/MS analysis to investigate the MIR-triggered molecular mechanisms in breast cancer cells. A total of 1749 proteins were identified, quantified, and subjected to functional enrichment analysis. From the constructed functionally enriched network, we confirmed that MIR caused G2/M cell cycle arrest, remodeled the microtubule network to an astral pole arrangement, altered the actin filament formation and focal adhesion molecule localization, and reduced cell migration activity and invasion ability. Our results reveal the coordinative effects of MIR-regulated physiological responses in concentrated networks, demonstrating the potential implementation of infrared radiation in breast cancer therapy. PMID:25556991

  8. Exome Analysis Reveals Differentially Mutated Gene Signatures of Stage, Grade and Subtype in Breast Cancers

    PubMed Central

    Li, You; Wang, Xiaosheng; Vural, Suleyman; Mishra, Nitish K.; Cowan, Kenneth H.; Guda, Chittibabu

    2015-01-01

    Breast cancers exhibit highly heterogeneous molecular profiles. Although gene expression profiles have been used to predict the risks and prognostic outcomes of breast cancers, the high variability of gene expression limits its clinical application. In contrast, genetic mutation profiles would be more advantageous than gene expression profiles because genetic mutations can be stably detected and the mutational heterogeneity widely exists in breast cancer genomes. We analyzed 98 breast cancer whole exome samples that were sorted into three subtypes, two grades and two stages. The sum deleterious effect of all mutations in each gene was scored to identify differentially mutated genes (DMGs) for this case-control study. DMGs were corroborated using extensive published knowledge. Functional consequences of deleterious SNVs on protein structure and function were also investigated. Genes such as ERBB2, ESP8, PPP2R4, KIAA0922, SP4, CENPJ, PRCP and SELP that have been experimentally or clinically verified to be tightly associated with breast cancer prognosis are among the DMGs identified in this study. We also identified some genes such as ARL6IP5, RAET1E, and ANO7 that could be crucial for breast cancer development and prognosis. Further, SNVs such as rs1058808, rs2480452, rs61751507, rs79167802, rs11540666, and rs2229437 that potentially influence protein functions are observed at significantly different frequencies in different comparison groups. Protein structure modeling revealed that many non-synonymous SNVs have a deleterious effect on protein stability, structure and function. Mutational profiling at gene- and SNV-level revealed differential patterns within each breast cancer comparison group, and the gene signatures correlate with expected prognostic characteristics of breast cancer classes. Some of the genes and SNVs identified in this study show high promise and are worthy of further investigation by experimental studies. PMID:25803781

  9. Base changes in tumour DNA have the power to reveal the causes and evolution of cancer

    DOE PAGESBeta

    Hollstein, M.; Alexandrov, L. B.; Wild, C. P.; Ardin, M.; Zavadil, J.

    2016-06-06

    Next-generation sequencing (NGS) technology has demonstrated that the cancer genomes are peppered with mutations. Although most somatic tumour mutations are unlikely to have any role in the cancer process per se, the spectra of DNA sequence changes in tumour mutation catalogues have the potential to identify the mutagens, and to reveal the mutagenic processes responsible for human cancer. Very recently, a novel approach for data mining of the vast compilations of tumour NGS data succeeded in separating and precisely defining at least 30 distinct patterns of sequence change hidden in mutation databases. At least half of these mutational signatures canmore » be readily assigned to known human carcinogenic exposures or endogenous mechanisms of mutagenesis. A quantum leap in our knowledge of mutagenesis in human cancers has resulted, stimulating a flurry of research activity. We trace here the major findings leading first to the hypothesis that carcinogenic insults leave characteristic imprints on the DNA sequence of tumours, and culminating in empirical evidence from NGS data that well-defined carcinogen mutational signatures are indeed present in tumour genomic DNA from a variety of cancer types. The notion that tumour DNAs can divulge environmental sources of mutation is now a well-accepted fact. This approach to cancer aetiology has also incriminated various endogenous, enzyme-driven processes that increase the somatic mutation load in sporadic cancers. The tasks now confronting the field of molecular epidemiology are to assign mutagenic processes to orphan and newly discovered tumour mutation patterns, and to determine whether avoidable cancer risk factors influence signatures produced by endogenous enzymatic mechanisms. As a result, innovative research with experimental models and exploitation of the geographical heterogeneity in cancer incidence can address these challenges.« less

  10. The distinctive gastric fluid proteome in gastric cancer reveals a multi-biomarker diagnostic profile

    PubMed Central

    Kon, Oi Lian; Yip, Tai-Tung; Ho, Meng Fatt; Chan, Weng Hoong; Wong, Wai Keong; Tan, Soo Yong; Ng, Wai Har; Kam, Siok Yuen; Eng, Alvin KH; Ho, Patrick; Viner, Rosa; Ong, Hock Soo; Kumarasinghe, M Priyanthi

    2008-01-01

    Background Overall gastric cancer survival remains poor mainly because there are no reliable methods for identifying highly curable early stage disease. Multi-protein profiling of gastric fluids, obtained from the anatomic site of pathology, could reveal diagnostic proteomic fingerprints. Methods Protein profiles were generated from gastric fluid samples of 19 gastric cancer and 36 benign gastritides patients undergoing elective, clinically-indicated gastroscopy using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry on multiple ProteinChip arrays. Proteomic features were compared by significance analysis of microarray algorithm and two-way hierarchical clustering. A second blinded sample set (24 gastric cancers and 29 clinically benign gastritides) was used for validation. Results By significance analysyis of microarray, 60 proteomic features were up-regulated and 46 were down-regulated in gastric cancer samples (p < 0.01). Multimarker clustering showed two distinctive proteomic profiles independent of age and ethnicity. Eighteen of 19 cancer samples clustered together (sensitivity 95%) while 27/36 of non-cancer samples clustered in a second group. Nine non-cancer samples that clustered with cancer samples included 5 pre-malignant lesions (1 adenomatous polyp and 4 intestinal metaplasia). Validation using a second sample set showed the sensitivity and specificity to be 88% and 93%, respectively. Positive predictive value of the combined data was 0.80. Selected peptide sequencing identified pepsinogen C and pepsin A activation peptide as significantly down-regulated and alpha-defensin as significantly up-regulated. Conclusion This simple and reproducible multimarker proteomic assay could supplement clinical gastroscopic evaluation of symptomatic patients to enhance diagnostic accuracy for gastric cancer and pre-malignant lesions. PMID:18950519

  11. Suicide Gene-Engineered Stromal Cells Reveal a Dynamic Regulation of Cancer Metastasis

    NASA Astrophysics Data System (ADS)

    Shen, Keyue; Luk, Samantha; Elman, Jessica; Murray, Ryan; Mukundan, Shilpaa; Parekkadan, Biju

    2016-02-01

    Cancer-associated fibroblasts (CAFs) are a major cancer-promoting component in the tumor microenvironment (TME). The dynamic role of human CAFs in cancer progression has been ill-defined because human CAFs lack a unique marker needed for a cell-specific, promoter-driven knockout model. Here, we developed an engineered human CAF cell line with an inducible suicide gene to enable selective in vivo elimination of human CAFs at different stages of xenograft tumor development, effectively circumventing the challenge of targeting a cell-specific marker. Suicide-engineered CAFs were highly sensitive to apoptosis induction in vitro and in vivo by the addition of a simple small molecule inducer. Selection of timepoints for targeted CAF apoptosis in vivo during the progression of a human breast cancer xenograft model was guided by a bi-phasic host cytokine response that peaked at early timepoints after tumor implantation. Remarkably, we observed that the selective apoptosis of CAFs at these early timepoints did not affect primary tumor growth, but instead increased the presence of tumor-associated macrophages and the metastatic spread of breast cancer cells to the lung and bone. The study revealed a dynamic relationship between CAFs and cancer metastasis that has counter-intuitive ramifications for CAF-targeted therapy.

  12. Suicide Gene-Engineered Stromal Cells Reveal a Dynamic Regulation of Cancer Metastasis

    PubMed Central

    Shen, Keyue; Luk, Samantha; Elman, Jessica; Murray, Ryan; Mukundan, Shilpaa; Parekkadan, Biju

    2016-01-01

    Cancer-associated fibroblasts (CAFs) are a major cancer-promoting component in the tumor microenvironment (TME). The dynamic role of human CAFs in cancer progression has been ill-defined because human CAFs lack a unique marker needed for a cell-specific, promoter-driven knockout model. Here, we developed an engineered human CAF cell line with an inducible suicide gene to enable selective in vivo elimination of human CAFs at different stages of xenograft tumor development, effectively circumventing the challenge of targeting a cell-specific marker. Suicide-engineered CAFs were highly sensitive to apoptosis induction in vitro and in vivo by the addition of a simple small molecule inducer. Selection of timepoints for targeted CAF apoptosis in vivo during the progression of a human breast cancer xenograft model was guided by a bi-phasic host cytokine response that peaked at early timepoints after tumor implantation. Remarkably, we observed that the selective apoptosis of CAFs at these early timepoints did not affect primary tumor growth, but instead increased the presence of tumor-associated macrophages and the metastatic spread of breast cancer cells to the lung and bone. The study revealed a dynamic relationship between CAFs and cancer metastasis that has counter-intuitive ramifications for CAF-targeted therapy. PMID:26893143

  13. Chrysin suppresses the achaete-scute complex-like1 and alters the neuroendocrine phenotype of carcinoids

    PubMed Central

    Somnay, Yash R.; Dull, Barbara Zarebczan; Eide, Jacob; Jaskula-Sztul, Renata; Chen, Herbert

    2015-01-01

    Carcinoids are neuroendocrine neoplasms that cause significant morbidity and mortality, and for which few effective therapies are available. Given the recent identification of the anti-cancer flavonoid chrysin, we sought to investigate its therapeutic potential in carcinoids. Here, we report chrysin’s ability to modulate the achaete-scute complex-like1 (ASCL1), a neuroendocrine-specific transcription factor highly implicated in the malignant phenotype of carcinoids and other neuroendocrine cancers. Moreover, we elucidate the role of ASCL1 in carcinoid growth and bioactivity. Treatment of two carcinoid cell lines (BON and H727) with varying chrysin concentrations suppressed cell proliferation, while reducing expression of ASCL1 and the neuroendocrine biomarker chromogranin A (CgA), demonstrated by Western blotting. Propidium iodide and PE AnnexinV/7-AAD staining and sorting following chrysin treatment revealed S/G2 phase arrest and apoptosis, respectively. This was corroborated by chrysin-induced cleavage of caspase-3 and PARP and activation of p21Waf1/Cip1. Furthermore, direct ASCL1 knockdown with an ASCL1-specific small interfering RNA inhibited CgA and synaptophysin expression as well as carcinoid proliferation, while also reducing cyclin B1 and D1, and increasing p21Waf1/Cip1 and p27Kip1 expression, suggesting an arrest of the cell-cycle. Collectively, these findings warrant the deliberation of targeted ASCL1 suppression by chrysin or other agents as a therapeutic approach for carcinoid management. PMID:26403073

  14. Integrative analysis of breast cancer reveals prognostic haematopoietic activity and patient-specific immune response profiles

    PubMed Central

    Varn, Frederick S.; Andrews, Erik H.; Mullins, David W.; Cheng, Chao

    2016-01-01

    Transcriptional programmes active in haematopoietic cells enable a variety of functions including dedifferentiation, innate immunity and adaptive immunity. Understanding how these programmes function in the context of cancer can provide valuable insights into host immune response, cancer severity and potential therapy response. Here we present a method that uses the transcriptomes of over 200 murine haematopoietic cells, to infer the lineage-specific haematopoietic activity present in human breast tumours. Correlating this activity with patient survival and tumour purity reveals that the transcriptional programmes of many cell types influence patient prognosis and are found in environments of high lymphocytic infiltration. Collectively, these results allow for a detailed and personalized assessment of the patient immune response to a tumour. When combined with routinely collected patient biopsy genomic data, this method can enable a richer understanding of the complex interplay between the host immune system and cancer. PMID:26725977

  15. Integrative analysis of breast cancer reveals prognostic haematopoietic activity and patient-specific immune response profiles.

    PubMed

    Varn, Frederick S; Andrews, Erik H; Mullins, David W; Cheng, Chao

    2016-01-01

    Transcriptional programmes active in haematopoietic cells enable a variety of functions including dedifferentiation, innate immunity and adaptive immunity. Understanding how these programmes function in the context of cancer can provide valuable insights into host immune response, cancer severity and potential therapy response. Here we present a method that uses the transcriptomes of over 200 murine haematopoietic cells, to infer the lineage-specific haematopoietic activity present in human breast tumours. Correlating this activity with patient survival and tumour purity reveals that the transcriptional programmes of many cell types influence patient prognosis and are found in environments of high lymphocytic infiltration. Collectively, these results allow for a detailed and personalized assessment of the patient immune response to a tumour. When combined with routinely collected patient biopsy genomic data, this method can enable a richer understanding of the complex interplay between the host immune system and cancer. PMID:26725977

  16. Proteome–Metabolome Profiling of Ovarian Cancer Ascites Reveals Novel Components Involved in Intercellular Communication*

    PubMed Central

    Shender, Victoria O.; Pavlyukov, Marat S.; Ziganshin, Rustam H.; Arapidi, Georgij P.; Kovalchuk, Sergey I.; Anikanov, Nikolay A.; Altukhov, Ilya A.; Alexeev, Dmitry G.; Butenko, Ivan O.; Shavarda, Alexey L.; Khomyakova, Elena B.; Evtushenko, Evgeniy; Ashrafyan, Lev A.; Antonova, Irina B.; Kuznetcov, Igor N.; Gorbachev, Alexey Yu.; Shakhparonov, Mikhail I.; Govorun, Vadim M.

    2014-01-01

    Ovarian cancer ascites is a native medium for cancer cells that allows investigation of their secretome in a natural environment. This medium is of interest as a promising source of potential biomarkers, and also as a medium for cell–cell communication. The aim of this study was to elucidate specific features of the malignant ascites metabolome and proteome. In order to omit components of the systemic response to ascites formation, we compared malignant ascites with cirrhosis ascites. Metabolome analysis revealed 41 components that differed significantly between malignant and cirrhosis ascites. Most of the identified cancer-specific metabolites are known to be important signaling molecules. Proteomic analysis identified 2096 and 1855 proteins in the ovarian cancer and cirrhosis ascites, respectively; 424 proteins were specific for the malignant ascites. Functional analysis of the proteome demonstrated that the major differences between cirrhosis and malignant ascites were observed for the cluster of spliceosomal proteins. Additionally, we demonstrate that several splicing RNAs were exclusively detected in malignant ascites, where they probably existed within protein complexes. This result was confirmed in vitro using an ovarian cancer cell line. Identification of spliceosomal proteins and RNAs in an extracellular medium is of particular interest; the finding suggests that they might play a role in the communication between cancer cells. In addition, malignant ascites contains a high number of exosomes that are known to play an important role in signal transduction. Thus our study reveals the specific features of malignant ascites that are associated with its function as a medium of intercellular communication. PMID:25271300

  17. Revealing Glycoproteins in the Secretome of MCF-7 Human Breast Cancer Cells

    PubMed Central

    Tan, Aik-Aun; Phang, Wai-Mei; Gopinath, Subash C. B.; Hashim, Onn H.; Kiew, Lik Voon; Chen, Yeng

    2015-01-01

    Breast cancer is one of the major issues in the field of oncology, reported with a higher prevalence rate in women worldwide. In attempt to reveal the potential biomarkers for breast cancer, the findings of differentially glycosylated haptoglobin and osteonectin in previous study have drawn our attention towards glycoproteins of secretome from the MCF-7 cancer cell line. In the present study, further analyses were performed on the medium of MCF-7 cells by subjecting it to two-dimensional analyses followed by image analysis in contrast to the medium of human mammary epithelial cells (HMEpC) as a negative control. Carboxypeptidase A4 (CPA4), alpha-1-antitrypsin (AAT), haptoglobin (HP), and HSC70 were detected in the medium of MCF-7, while only CPA4 and osteonectin (ON) were detected in HMEpC medium. In addition, CPA4 was detected as upregulated in the MCF-7 medium. Further analysis by lectin showed that CPA4, AAT, HP, and HSC70 were secreted as N-glycan in the medium of MCF-7, with HP also showing differentially N-glycosylated isoforms. For the HMEpC, only CPA4 was detected as N-glycan. No O-glycan was detected in the medium of HMEpC but MCF-7 expressed O-glycosylated CPA4 and HSC70. All these revealed that glycoproteins could be used as glycan-based biomarkers for the prognosis of breast cancer. PMID:26167486

  18. Solutions to Peto's paradox revealed by mathematical modelling and cross-species cancer gene analysis.

    PubMed

    Caulin, Aleah F; Graham, Trevor A; Wang, Li-San; Maley, Carlo C

    2015-07-19

    Whales have 1000-fold more cells than humans and mice have 1000-fold fewer; however, cancer risk across species does not increase with the number of somatic cells and the lifespan of the organism. This observation is known as Peto's paradox. How much would evolution have to change the parameters of somatic evolution in order to equalize the cancer risk between species that differ by orders of magnitude in size? Analysis of previously published models of colorectal cancer suggests that a two- to three-fold decrease in the mutation rate or stem cell division rate is enough to reduce a whale's cancer risk to that of a human. Similarly, the addition of one to two required tumour-suppressor gene mutations would also be sufficient. We surveyed mammalian genomes and did not find a positive correlation of tumour-suppressor genes with increasing body mass and longevity. However, we found evidence of the amplification of TP53 in elephants, MAL in horses and FBXO31 in microbats, which might explain Peto's paradox in those species. Exploring parameters that evolution may have fine-tuned in large, long-lived organisms will help guide future experiments to reveal the underlying biology responsible for Peto's paradox and guide cancer prevention in humans. PMID:26056366

  19. Solutions to Peto's paradox revealed by mathematical modelling and cross-species cancer gene analysis

    PubMed Central

    Caulin, Aleah F.; Graham, Trevor A.; Wang, Li-San; Maley, Carlo C.

    2015-01-01

    Whales have 1000-fold more cells than humans and mice have 1000-fold fewer; however, cancer risk across species does not increase with the number of somatic cells and the lifespan of the organism. This observation is known as Peto's paradox. How much would evolution have to change the parameters of somatic evolution in order to equalize the cancer risk between species that differ by orders of magnitude in size? Analysis of previously published models of colorectal cancer suggests that a two- to three-fold decrease in the mutation rate or stem cell division rate is enough to reduce a whale's cancer risk to that of a human. Similarly, the addition of one to two required tumour-suppressor gene mutations would also be sufficient. We surveyed mammalian genomes and did not find a positive correlation of tumour-suppressor genes with increasing body mass and longevity. However, we found evidence of the amplification of TP53 in elephants, MAL in horses and FBXO31 in microbats, which might explain Peto's paradox in those species. Exploring parameters that evolution may have fine-tuned in large, long-lived organisms will help guide future experiments to reveal the underlying biology responsible for Peto's paradox and guide cancer prevention in humans. PMID:26056366

  20. Methylome sequencing in triple-negative breast cancer reveals distinct methylation clusters with prognostic value.

    PubMed

    Stirzaker, Clare; Zotenko, Elena; Song, Jenny Z; Qu, Wenjia; Nair, Shalima S; Locke, Warwick J; Stone, Andrew; Armstong, Nicola J; Robinson, Mark D; Dobrovic, Alexander; Avery-Kiejda, Kelly A; Peters, Kate M; French, Juliet D; Stein, Sandra; Korbie, Darren J; Trau, Matt; Forbes, John F; Scott, Rodney J; Brown, Melissa A; Francis, Glenn D; Clark, Susan J

    2015-01-01

    Epigenetic alterations in the cancer methylome are common in breast cancer and provide novel options for tumour stratification. Here, we perform whole-genome methylation capture sequencing on small amounts of DNA isolated from formalin-fixed, paraffin-embedded tissue from triple-negative breast cancer (TNBC) and matched normal samples. We identify differentially methylated regions (DMRs) enriched with promoters associated with transcription factor binding sites and DNA hypersensitive sites. Importantly, we stratify TNBCs into three distinct methylation clusters associated with better or worse prognosis and identify 17 DMRs that show a strong association with overall survival, including DMRs located in the Wilms tumour 1 (WT1) gene, bi-directional-promoter and antisense WT1-AS. Our data reveal that coordinated hypermethylation can occur in oestrogen receptor-negative disease, and that characterizing the epigenetic framework provides a potential signature to stratify TNBCs. Together, our findings demonstrate the feasibility of profiling the cancer methylome with limited archival tissue to identify regulatory regions associated with cancer. PMID:25641231

  1. Multiplatform analysis of 12 cancer types reveals molecular classification within and across tissues of origin.

    PubMed

    Hoadley, Katherine A; Yau, Christina; Wolf, Denise M; Cherniack, Andrew D; Tamborero, David; Ng, Sam; Leiserson, Max D M; Niu, Beifang; McLellan, Michael D; Uzunangelov, Vladislav; Zhang, Jiashan; Kandoth, Cyriac; Akbani, Rehan; Shen, Hui; Omberg, Larsson; Chu, Andy; Margolin, Adam A; Van't Veer, Laura J; Lopez-Bigas, Nuria; Laird, Peter W; Raphael, Benjamin J; Ding, Li; Robertson, A Gordon; Byers, Lauren A; Mills, Gordon B; Weinstein, John N; Van Waes, Carter; Chen, Zhong; Collisson, Eric A; Benz, Christopher C; Perou, Charles M; Stuart, Joshua M

    2014-08-14

    Recent genomic analyses of pathologically defined tumor types identify "within-a-tissue" disease subtypes. However, the extent to which genomic signatures are shared across tissues is still unclear. We performed an integrative analysis using five genome-wide platforms and one proteomic platform on 3,527 specimens from 12 cancer types, revealing a unified classification into 11 major subtypes. Five subtypes were nearly identical to their tissue-of-origin counterparts, but several distinct cancer types were found to converge into common subtypes. Lung squamous, head and neck, and a subset of bladder cancers coalesced into one subtype typified by TP53 alterations, TP63 amplifications, and high expression of immune and proliferation pathway genes. Of note, bladder cancers split into three pan-cancer subtypes. The multiplatform classification, while correlated with tissue-of-origin, provides independent information for predicting clinical outcomes. All data sets are available for data-mining from a unified resource to support further biological discoveries and insights into novel therapeutic strategies. PMID:25109877

  2. Chrysin suppresses achaete-scute complex-like 1 and alters the neuroendocrine phenotype of carcinoids.

    PubMed

    Somnay, Y R; Dull, B Z; Eide, J; Jaskula-Sztul, R; Chen, H

    2015-10-01

    Carcinoids are neuroendocrine neoplasms that cause significant morbidity and mortality and for which few effective therapies are available. Given the recent identification of the anticancer flavonoid chrysin, we sought to investigate its therapeutic potential in carcinoids. Here we report chrysin's ability to modulate the achaete-scute complex-like 1 (ASCL1), a neuroendocrine-specific transcription factor highly implicated in the malignant phenotype of carcinoids and other neuroendocrine cancers. Moreover, we elucidate the role of ASCL1 in carcinoid growth and bioactivity. Treatment of two carcinoid cell lines (BON and H727) with varying chrysin concentrations suppressed cell proliferation, while reducing expression of ASCL1 and the neuroendocrine biomarker chromogranin A (CgA), demonstrated by western blotting. Propidium iodide and phycoerythrin AnnexinV/7-aminoactinomycin D staining and sorting following chrysin treatment revealed S/G2 phase arrest and apoptosis, respectively. This was corroborated by chrysin-induced cleavage of caspase-3 and poly ADP-ribose polymerase and activation of p21(Waf1/Cip1). Furthermore, direct ASCL1 knockdown with an ASCL1-specific small interfering RNA inhibited CgA and synaptophysin expression as well as carcinoid proliferation, while also reducing cyclin B1 and D1 and increasing p21(Waf1/Cip1) and p27(Kip1) expression, suggesting an arrest of the cell cycle. Collectively, these findings warrant the deliberation of targeted ASCL1 suppression by chrysin or other agents as a therapeutic approach for carcinoid management. PMID:26403073

  3. The pan-inhibitor of Aurora kinases danusertib induces apoptosis and autophagy and suppresses epithelial-to-mesenchymal transition in human breast cancer cells

    PubMed Central

    Li, Jin-Ping; Yang, Yin-Xue; Liu, Qi-Lun; Zhou, Zhi-Wei; Pan, Shu-Ting; He, Zhi-Xu; Zhang, Xueji; Yang, Tianxin; Pan, Si-Yuan; Duan, Wei; He, Shu-Ming; Chen, Xiao-Wu; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    Danusertib (Danu) is a pan-inhibitor of Aurora kinases and a third-generation breakpoint cluster region-Abelson murine leukemia viral oncogene homolog 1 (Bcr-Abl) tyrosine kinase inhibitor, but its antitumor effect and underlying mechanisms in the treatment of human breast cancer remain elusive. This study aimed to investigate the effects of Danu on the growth, apoptosis, autophagy, and epithelial-to-mesenchymal transition (EMT) and the molecular mechanisms in human breast cancer MCF7 and MDA-MB-231 cells. The results demonstrated that Danu remarkably inhibited cell proliferation, induced apoptosis and autophagy, and suppressed EMT in both breast cancer cell lines. Danu arrested MCF7 and MDA-MB-231 cells in G2/M phase, accompanied by the downregulation of cyclin-dependent kinase 1 and cyclin B1 and upregulation of p21 Waf1/Cip1, p27 Kip1, and p53. Danu significantly decreased the expression of B-cell lymphoma-extra-large (Bcl-xl) and B-cell lymphoma 2 (Bcl-2), but increased the expression of Bcl-2-associated X protein (Bax) and p53-upregulated modulator of apoptosis (PUMA), and promoted the cleavage of caspases 3 and 9. Furthermore, Danu significantly increased the expression levels of the membrane-bound microtubule-associated protein 1A/1B-light chain 3 (LC3-II) and beclin 1 in breast cancer cells, two markers for autophagy. Danu induced the activation of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinases 1 and 2 (Erk1/2) and inhibited the activation of protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathways in breast cancer cells. Treatment with wortmannin (a phosphatidylinositol 3-kinase inhibitor) markedly inhibited Danu-induced activation of p38 MAPK and conversion of cytosolic LC3-I to membrane-bound LC3-II. Pharmacological inhibition and small interfering RNA-mediated knockdown of p38 MAPK suppressed Akt activation, resulting in LC3-II accumulation and enhanced autophagy. Pharmacological inhibition

  4. Quantitative measures to reveal coordinated cytoskeleton-nucleus reorganization during in vitro invasion of cancer cells

    NASA Astrophysics Data System (ADS)

    Dvir, Liron; Nissim, Ronen; Alvarez-Elizondo, Martha B.; Weihs, Daphne

    2015-04-01

    Metastasis formation is a major cause of mortality in cancer patients and includes tumor cell relocation to distant organs. A metastatic cell invades through other cells and extracellular matrix by biochemical attachment and mechanical force application. Force is used to move on or through a 2- or 3-dimensional (3D) environment, respectively, or to penetrate a 2D substrate. We have previously shown that even when a gel substrate is impenetrable, metastatic breast cancer cells can still indent it by applying force. Cells typically apply force through the acto-myosin network, which is mechanically connected to the nucleus. We develop a 3D image-analysis to reveal relative locations of the cell elements, and show that as cells apply force to the gel, a coordinated process occurs that involves cytoskeletal remodeling and repositioning of the nucleus. Our approach shows that the actin and microtubules reorganize in the cell, bringing the actin to the leading edge of the cell. In parallel, the nucleus is transported behind the actin, likely by the cytoskeleton, into the indentation dimple formed in the gel. The nucleus volume below the gel surface correlates with indentation depth, when metastatic breast cancer cells indent gels deeply. However, the nucleus always remains above the gel in benign cells, even when small indentations are observed. Determining mechanical processes during metastatic cell invasion can reveal how cells disseminate in the body and can uncover targets for diagnosis and treatment.

  5. Population analysis of microsatellite genotypes reveals a signature associated with ovarian cancer.

    PubMed

    Fonville, Natalie C; Vaksman, Zalman; McIver, Lauren J; Garner, Harold R

    2015-05-10

    Ovarian cancer (OV) ranks fifth in cancer deaths among women, yet there remain few informative biomarkers for this disease. Microsatellites are repetitive genomic regions which we hypothesize could be a source of novel biomarkers for OV and have traditionally been under-appreciated relative to Single Nucleotide Polymorphisms (SNPs). In this study, we explore microsatellite variation as a potential novel source of genomic variation associated with OV. Exomes from 305 OV patient germline samples and 54 tumors, sequenced as part of The Cancer Genome Atlas, were analyzed for microsatellite variation and compared to healthy females sequenced as part of the 1,000 Genomes Project. We identified a subset of 60 microsatellite loci with genotypes that varied significantly between the OV and healthy female populations. Using these loci as a signature set, we classified germline genomes as 'at risk' for OV with a sensitivity of 90.1% and a specificity of 87.6%. Cross-analysis with a similar set of breast cancer associated loci identified individuals 'at risk' for both diseases. This study revealed a genotype-based microsatellite signature present in the germlines of individuals diagnosed with OV, and provides the basis for a potential novel risk assessment diagnostic for OV and new personal genomics targets in tumors. PMID:25779658

  6. Network modelling reveals the mechanism underlying colitis-associated colon cancer and identifies novel combinatorial anti-cancer targets.

    PubMed

    Lu, Junyan; Zeng, Hanlin; Liang, Zhongjie; Chen, Limin; Zhang, Liyi; Zhang, Hao; Liu, Hong; Jiang, Hualiang; Shen, Bairong; Huang, Ming; Geng, Meiyu; Spiegel, Sarah; Luo, Cheng

    2015-01-01

    The connection between inflammation and tumourigenesis has been well established. However, the detailed molecular mechanism underlying inflammation-associated tumourigenesis remains unknown because this process involves a complex interplay between immune microenvironments and epithelial cells. To obtain a more systematic understanding of inflammation-associated tumourigenesis as well as to identify novel therapeutic approaches, we constructed a knowledge-based network describing the development of colitis-associated colon cancer (CAC) by integrating the extracellular microenvironment and intracellular signalling pathways. Dynamic simulations of the CAC network revealed a core network module, including P53, MDM2, and AKT, that may govern the malignant transformation of colon epithelial cells in a pro-tumor inflammatory microenvironment. Furthermore, in silico mutation studies and experimental validations led to a novel finding that concurrently targeting ceramide and PI3K/AKT pathway by chemical probes or marketed drugs achieves synergistic anti-cancer effects. Overall, our network model can guide further mechanistic studies on CAC and provide new insights into the design of combinatorial cancer therapies in a rational manner. PMID:26446703

  7. Network modelling reveals the mechanism underlying colitis-associated colon cancer and identifies novel combinatorial anti-cancer targets

    PubMed Central

    Lu, Junyan; Zeng, Hanlin; Liang, Zhongjie; Chen, Limin; Zhang, Liyi; Zhang, Hao; Liu, Hong; Jiang, Hualiang; Shen, Bairong; Huang, Ming; Geng, Meiyu; Spiegel, Sarah; Luo, Cheng

    2015-01-01

    The connection between inflammation and tumourigenesis has been well established. However, the detailed molecular mechanism underlying inflammation-associated tumourigenesis remains unknown because this process involves a complex interplay between immune microenvironments and epithelial cells. To obtain a more systematic understanding of inflammation-associated tumourigenesis as well as to identify novel therapeutic approaches, we constructed a knowledge-based network describing the development of colitis-associated colon cancer (CAC) by integrating the extracellular microenvironment and intracellular signalling pathways. Dynamic simulations of the CAC network revealed a core network module, including P53, MDM2, and AKT, that may govern the malignant transformation of colon epithelial cells in a pro-tumor inflammatory microenvironment. Furthermore, in silico mutation studies and experimental validations led to a novel finding that concurrently targeting ceramide and PI3K/AKT pathway by chemical probes or marketed drugs achieves synergistic anti-cancer effects. Overall, our network model can guide further mechanistic studies on CAC and provide new insights into the design of combinatorial cancer therapies in a rational manner. PMID:26446703

  8. Structure-Based Analysis Reveals Cancer Missense Mutations Target Protein Interaction Interfaces.

    PubMed

    Engin, H Billur; Kreisberg, Jason F; Carter, Hannah

    2016-01-01

    Recently it has been shown that cancer mutations selectively target protein-protein interactions. We hypothesized that mutations affecting distinct protein interactions involving established cancer genes could contribute to tumor heterogeneity, and that novel mechanistic insights might be gained into tumorigenesis by investigating protein interactions under positive selection in cancer. To identify protein interactions under positive selection in cancer, we mapped over 1.2 million nonsynonymous somatic cancer mutations onto 4,896 experimentally determined protein structures and analyzed their spatial distribution. In total, 20% of mutations on the surface of known cancer genes perturbed protein-protein interactions (PPIs), and this enrichment for PPI interfaces was observed for both tumor suppressors (Odds Ratio 1.28, P-value < 10-4) and oncogenes (Odds Ratio 1.17, P-value < 10-3). To study this further, we constructed a bipartite network representing structurally resolved PPIs from all available human complexes in the Protein Data Bank (2,864 proteins, 3,072 PPIs). Analysis of frequently mutated cancer genes within this network revealed that tumor-suppressors, but not oncogenes, are significantly enriched with functional mutations in homo-oligomerization regions (Odds Ratio 3.68, P-Value < 10-8). We present two important examples, TP53 and beta-2-microglobulin, for which the patterns of somatic mutations at interfaces provide insights into specifically perturbed biological circuits. In patients with TP53 mutations, patient survival correlated with the specific interactions that were perturbed. Moreover, we investigated mutations at the interface of protein-nucleotide interactions and observed an unexpected number of missense mutations but not silent mutations occurring within DNA and RNA binding sites. Finally, we provide a resource of 3,072 PPI interfaces ranked according to their mutation rates. Analysis of this list highlights 282 novel candidate cancer genes

  9. Structure-Based Analysis Reveals Cancer Missense Mutations Target Protein Interaction Interfaces

    PubMed Central

    Engin, H. Billur; Kreisberg, Jason F.; Carter, Hannah

    2016-01-01

    Recently it has been shown that cancer mutations selectively target protein-protein interactions. We hypothesized that mutations affecting distinct protein interactions involving established cancer genes could contribute to tumor heterogeneity, and that novel mechanistic insights might be gained into tumorigenesis by investigating protein interactions under positive selection in cancer. To identify protein interactions under positive selection in cancer, we mapped over 1.2 million nonsynonymous somatic cancer mutations onto 4,896 experimentally determined protein structures and analyzed their spatial distribution. In total, 20% of mutations on the surface of known cancer genes perturbed protein-protein interactions (PPIs), and this enrichment for PPI interfaces was observed for both tumor suppressors (Odds Ratio 1.28, P-value < 10−4) and oncogenes (Odds Ratio 1.17, P-value < 10−3). To study this further, we constructed a bipartite network representing structurally resolved PPIs from all available human complexes in the Protein Data Bank (2,864 proteins, 3,072 PPIs). Analysis of frequently mutated cancer genes within this network revealed that tumor-suppressors, but not oncogenes, are significantly enriched with functional mutations in homo-oligomerization regions (Odds Ratio 3.68, P-Value < 10−8). We present two important examples, TP53 and beta-2-microglobulin, for which the patterns of somatic mutations at interfaces provide insights into specifically perturbed biological circuits. In patients with TP53 mutations, patient survival correlated with the specific interactions that were perturbed. Moreover, we investigated mutations at the interface of protein-nucleotide interactions and observed an unexpected number of missense mutations but not silent mutations occurring within DNA and RNA binding sites. Finally, we provide a resource of 3,072 PPI interfaces ranked according to their mutation rates. Analysis of this list highlights 282 novel candidate cancer

  10. Integrated analysis of the prostate cancer small-nucleolar transcriptome reveals SNORA55 as a driver of prostate cancer progression.

    PubMed

    Crea, Francesco; Quagliata, Luca; Michael, Agnieszka; Liu, Hui Hsuan; Frumento, Paolo; Azad, Arun A; Xue, Hui; Pikor, Larissa; Watahiki, Akira; Morant, Rudolf; Eppenberger-Castori, Serenella; Wang, Yuwei; Parolia, Abhijit; Lennox, Kim A; Lam, Wan L; Gleave, Martin; Chi, Kim N; Pandha, Hardev; Wang, Yuzhuo; Helgason, Cheryl D

    2016-05-01

    Metastasis is the primary cause of death in prostate cancer (PCa) patients. Small nucleolar RNAs (snoRNAs) have long been considered "housekeeping" genes with no relevance for cancer biology. Emerging evidence has challenged this assumption, suggesting that snoRNA expression is frequently modulated during cancer progression. Despite this, no study has systematically addressed the prognostic and functional significance of snoRNAs in PCa. We performed RNA Sequencing on paired metastatic/non-metastatic PCa xenografts derived from clinical specimens. The clinical significance of differentially expressed snoRNAs was further investigated in two independent primary PCa cohorts (131 and 43 patients, respectively). The snoRNA demonstrating the strongest association with clinical outcome was quantified in PCa patient-derived serum samples and its functional relevance was investigated in PCa cells via gene expression profiling, pathway analysis and gene silencing. Our comparison revealed 21 differentially expressed snoRNAs in the metastatic vs. non-metastatic xenografts. Of those, 12 were represented in clinical databases and were further analyzed. SNORA55 emerged as a predictor of shorter relapse-free survival (results confirmed in two independent databases). SNORA55 was reproducibly detectable in serum samples from PCa patients. SNORA55 silencing in PCa cell lines significantly inhibited cell proliferation and migration. Pathway analysis revealed that SNORA55 expression is significantly associated with growth factor signaling and pro-inflammatory cytokine expression in PCa. Our results demonstrate that SNORA55 up-regulation predicts PCa progression and that silencing this non-coding gene affects PCa cell proliferation and metastatic potential, thus positioning it as both a novel biomarker and therapeutic target. PMID:26809501

  11. Heterogeneity of pancreatic cancer metastases in a single patient revealed by quantitative proteomics.

    PubMed

    Kim, Min-Sik; Zhong, Yi; Yachida, Shinichi; Rajeshkumar, N V; Abel, Melissa L; Marimuthu, Arivusudar; Mudgal, Keshav; Hruban, Ralph H; Poling, Justin S; Tyner, Jeffrey W; Maitra, Anirban; Iacobuzio-Donahue, Christine A; Pandey, Akhilesh

    2014-11-01

    Many patients with pancreatic cancer have metastases to distant organs at the time of initial presentation. Recent studies examining the evolution of pancreatic cancer at the genetic level have shown that clonal complexity of metastatic pancreatic cancer is already initiated within primary tumors, and organ-specific metastases are derived from different subclones. However, we do not yet understand to what extent the evolution of pancreatic cancer contributes to proteomic and signaling alterations. We hypothesized that genetic heterogeneity of metastatic pancreatic cancer results in heterogeneity at the proteome level. To address this, we employed a model system in which cells isolated from three sites of metastasis (liver, lung, and peritoneum) from a single patient were compared. We used a SILAC-based accurate quantitative proteomic strategy combined with high-resolution mass spectrometry to analyze the total proteome and tyrosine phosphoproteome of each of the distal metastases. Our data revealed distinct patterns of both overall proteome expression and tyrosine kinase activities across the three different metastatic lesions. This heterogeneity was significant because it led to differential sensitivity of the neoplastic cells to small molecule inhibitors targeting various kinases and other pathways. For example, R428, a tyrosine kinase inhibitor that targets Axl receptor tyrosine kinase, was able to inhibit cells derived from lung and liver metastases much more effectively than cells from the peritoneal metastasis. Finally, we confirmed that administration of R428 in mice bearing xenografts of cells derived from the three different metastatic sites significantly diminished tumors formed from liver- and lung-metastasis-derived cell lines as compared with tumors derived from the peritoneal metastasis cell line. Overall, our data provide proof-of-principle support that personalized therapy of multiple organ metastases in a single patient should involve the

  12. A spontaneous metastasis model reveals the significance of claudin-9 overexpression in lung cancer metastasis.

    PubMed

    Sharma, Rajesh K; Chheda, Zinal S; Das Purkayastha, Biswa Pratim; Gomez-Gutierrez, Jorge G; Jala, Venkatakrishna R; Haribabu, Bodduluri

    2016-03-01

    Metastasis causes most cancer related mortality but the mechanisms governing metastatic dissemination are poorly defined. Metastasis involves egression of cancer cells from the primary tumors, their survival in circulation and colonization at the secondary sites. Cancer cell egression from the primary tumor is the least defined process of metastasis as experimental metastasis models directly seed cancer cells in circulation, thus bypassing this crucial step. Here, we developed a spontaneous metastasis model that retains the egression step of metastasis. By repeated in vivo passaging of the poorly metastatic Lewis lung carcinoma (3LL) cells, we generated a cell line (p-3LL) that readily metastasizes to lungs and liver from subcutaneous (s.c.) tumors. Interestingly, when injected intravenously, 3LL and p-3LL cells showed a similar frequency of metastasis. This suggests enhanced egression of p-3LL cells may underlie the enhanced metastatic spread from primary tumors. Microarray analysis of 3LL and p-3LL cells as well as the primary tumors derived from these cells revealed altered expression of several genes including significant upregulation of a tight junction protein, claudin-9. Increased expression of claudin-9 was confirmed in both p-3LL cells and tumors derived from these cells. Knockdown of claudin-9 expression in p-3LL cells by si-RNA significantly reduced their motility, invasiveness in vitro and metastasis in vivo. Conversely, transient overexpression of claudin-9 in 3LL cells enhanced their motility. These results suggest an essential role for claudin-9 in promoting lung cancer metastasis. PMID:26669782

  13. Meta-Analysis of Public Microarray Datasets Reveals Voltage-Gated Calcium Gene Signatures in Clinical Cancer Patients

    PubMed Central

    Wang, Chih-Yang; Lai, Ming-Derg; Phan, Nam Nhut; Sun, Zhengda; Lin, Yen-Chang

    2015-01-01

    Voltage-gated calcium channels (VGCCs) are well documented to play roles in cell proliferation, migration, and apoptosis; however, whether VGCCs regulate the onset and progression of cancer is still under investigation. The VGCC family consists of five members, which are L-type, N-type, T-type, R-type and P/Q type. To date, no holistic approach has been used to screen VGCC family genes in different types of cancer. We analyzed the transcript expression of VGCCs in clinical cancer tissue samples by accessing ONCOMINE (www.oncomine.org), a web-based microarray database, to perform a systematic analysis. Every member of the VGCCs was examined across 21 different types of cancer by comparing mRNA expression in cancer to that in normal tissue. A previous study showed that altered expression of mRNA in cancer tissue may play an oncogenic role and promote tumor development; therefore, in the present findings, we focus only on the overexpression of VGCCs in different types of cancer. This bioinformatics analysis revealed that different subtypes of VGCCs (CACNA1C, CACNA1D, CACNA1B, CACNA1G, and CACNA1I) are implicated in the development and progression of diverse types of cancer and show dramatic up-regulation in breast cancer. CACNA1F only showed high expression in testis cancer, whereas CACNA1A, CACNA1C, and CACNA1D were highly expressed in most types of cancer. The current analysis revealed that specific VGCCs likely play essential roles in specific types of cancer. Collectively, we identified several VGCC targets and classified them according to different cancer subtypes for prospective studies on the underlying carcinogenic mechanisms. The present findings suggest that VGCCs are possible targets for prospective investigation in cancer treatment. PMID:26147197

  14. Meta-Analysis of Public Microarray Datasets Reveals Voltage-Gated Calcium Gene Signatures in Clinical Cancer Patients.

    PubMed

    Wang, Chih-Yang; Lai, Ming-Derg; Phan, Nam Nhut; Sun, Zhengda; Lin, Yen-Chang

    2015-01-01

    Voltage-gated calcium channels (VGCCs) are well documented to play roles in cell proliferation, migration, and apoptosis; however, whether VGCCs regulate the onset and progression of cancer is still under investigation. The VGCC family consists of five members, which are L-type, N-type, T-type, R-type and P/Q type. To date, no holistic approach has been used to screen VGCC family genes in different types of cancer. We analyzed the transcript expression of VGCCs in clinical cancer tissue samples by accessing ONCOMINE (www.oncomine.org), a web-based microarray database, to perform a systematic analysis. Every member of the VGCCs was examined across 21 different types of cancer by comparing mRNA expression in cancer to that in normal tissue. A previous study showed that altered expression of mRNA in cancer tissue may play an oncogenic role and promote tumor development; therefore, in the present findings, we focus only on the overexpression of VGCCs in different types of cancer. This bioinformatics analysis revealed that different subtypes of VGCCs (CACNA1C, CACNA1D, CACNA1B, CACNA1G, and CACNA1I) are implicated in the development and progression of diverse types of cancer and show dramatic up-regulation in breast cancer. CACNA1F only showed high expression in testis cancer, whereas CACNA1A, CACNA1C, and CACNA1D were highly expressed in most types of cancer. The current analysis revealed that specific VGCCs likely play essential roles in specific types of cancer. Collectively, we identified several VGCC targets and classified them according to different cancer subtypes for prospective studies on the underlying carcinogenic mechanisms. The present findings suggest that VGCCs are possible targets for prospective investigation in cancer treatment. PMID:26147197

  15. Integrative Molecular Profiling Reveals Asparagine Synthetase Is a Target in Castration-Resistant Prostate Cancer

    PubMed Central

    Sircar, Kanishka; Huang, Heng; Hu, Limei; Cogdell, David; Dhillon, Jasreman; Tzelepi, Vassiliki; Efstathiou, Eleni; Koumakpayi, Ismaël H.; Saad, Fred; Luo, Dijun; Bismar, Tarek A.; Aparicio, Ana; Troncoso, Patricia; Navone, Nora; Zhang, Wei

    2013-01-01

    The identification of new and effective therapeutic targets for the lethal, castration-resistant stage of prostate cancer (CRPC) has been challenging because of both the paucity of adequate frozen tissues and a lack of integrated molecular analysis. Therefore, in this study, we performed a genome-wide analysis of DNA copy number alterations from 34 unique surgical CRPC specimens and 5 xenografts, with matched transcriptomic profiling of 25 specimens. An integrated analysis of these data revealed that the asparagine synthetase (ASNS) gene showed a gain in copy number and was overexpressed at the transcript level. The overexpression of ASNS was validated by analyzing other public CRPC data sets. ASNS protein expression, as detected by reverse-phase protein lysate array, was tightly correlated with gene copy number. In addition, ASNS protein expression, as determined by IHC analysis, was associated with progression to a therapy-resistant disease state in TMAs that included 77 castration-resistant and 40 untreated prostate cancer patient samples. Knockdown of ASNS by small-interfering RNAs in asparagine-deprived media led to growth inhibition in both androgen-responsive (ie, LNCaP) and castration-resistant (ie, C4-2B) prostate cancer cell lines and in cells isolated from a CRPC xenograft (ie, MDA PCa 180-30). Together, our results suggest that ASNS is up-regulated in cases of CRPC and that depletion of asparagine using ASNS inhibitors will be a novel strategy for targeting CRPC cells. PMID:22245216

  16. Microbial genomic analysis reveals the essential role of inflammation in bacteria-induced colorectal cancer

    PubMed Central

    Arthur, Janelle C.; Gharaibeh, Raad Z.; Mühlbauer, Marcus; Perez-Chanona, Ernesto; Uronis, Joshua M.; McCafferty, Jonathan; Fodor, Anthony A.; Jobin, Christian

    2014-01-01

    Enterobacteria, especially Escherichia coli, are abundant in patients with inflammatory bowel disease or colorectal cancer (CRC). However, it is unclear whether cancer is promoted by inflammation-induced expansion of E. coli and/or changes in expression of specific microbial genes. Here we use longitudinal (2, 12 and 20 weeks) 16S rRNA sequencing of luminal microbiota from ex-germ free mice to show that inflamed Il10−/− mice maintain a higher abundance of Enterobacteriaceae than healthy wild-type mice. Experiments with mono-colonized Il10−/− mice reveal that host inflammation is necessary for E. coli cancer-promoting activity. RNA-sequence analysis indicates significant changes in E. coli gene catalogue in Il10−/− mice, with changes mostly driven by adaptation to the intestinal environment. Expression of specific genes present in the tumor-promoting E. coli pks island are modulated by inflammation/CRC development. Thus, progression of inflammation in Il10−/− mice supports Enterobacteriaceae and alters a small subset of microbial genes important for tumor development. PMID:25182170

  17. Singularity analysis of the AKT signaling pathway reveals connections between cancer and metabolic diseases

    NASA Astrophysics Data System (ADS)

    Wang, Guanyu

    2010-12-01

    Connections between cancer and metabolic diseases may consist in the complex network of interactions among a common set of biomolecules. By applying singularity and bifurcation analysis, the phenotypes constrained by the AKT signaling pathway are identified and mapped onto the parameter space, which include cancer and certain metabolic diseases. By considering physiologic properties (sensitivity, robustness and adaptivity) the AKT pathway must possess in order to efficiently sense growth factors and nutrients, the region of normal responses is located. To optimize these properties, the intracellular concentration of the AKT protein must be sufficiently high to saturate its enzymes; the strength of the positive feedback must be stronger than that of the negative feedback. The analysis illuminates the parameter space and reveals system-level mechanisms in regulating biological functions (cell growth, survival, proliferation and metabolism) and how their deregulation may lead to the development of diseases. The analytical expressions summarize the synergistic interactions among many molecules, which provides valuable insights into therapeutic interventions. In particular, a strategy for overcoming the limitations of mTOR inhibition is proposed for cancer therapy.

  18. Endogenous molecular network reveals two mechanisms of heterogeneity within gastric cancer

    PubMed Central

    Li, Site; Zhu, Xiaomei; Liu, Bingya; Wang, Gaowei; Ao, Ping

    2015-01-01

    Intratumor heterogeneity is a common phenomenon and impedes cancer therapy and research. Gastric cancer (GC) cells have generally been classified into two heterogeneous cellular phenotypes, the gastric and intestinal types, yet the mechanisms of maintaining two phenotypes and controlling phenotypic transition are largely unknown. A qualitative systematic framework, the endogenous molecular network hypothesis, has recently been proposed to understand cancer genesis and progression. Here, a minimal network corresponding to such framework was found for GC and was quantified via a stochastic nonlinear dynamical system. We then further extended the framework to address the important question of intratumor heterogeneity quantitatively. The working network characterized main known features of normal gastric epithelial and GC cell phenotypes. Our results demonstrated that four positive feedback loops in the network are critical for GC cell phenotypes. Moreover, two mechanisms that contribute to GC cell heterogeneity were identified: particular positive feedback loops are responsible for the maintenance of intestinal and gastric phenotypes; GC cell progression routes that were revealed by the dynamical behaviors of individual key components are heterogeneous. In this work, we constructed an endogenous molecular network of GC that can be expanded in the future and would broaden the known mechanisms of intratumor heterogeneity. PMID:25962957

  19. Breast cancer revealed by a paraneoplastic cerebellar syndrome: about one case and literature review

    PubMed Central

    Adama, Dembélé; Moussa, Bambara; Emmanuel, Macoumi; Dennis, Ullmann

    2015-01-01

    To describe a case of breast cancer manifested by cerebellar syndrome and to establish a relationship between breast cancer and Paraneoplastic syndromes through the presence of anti- yo antibodies in serum and cerebrospinal fluid of a patient. Our patient was 52 years old, Multipara with 5 children alive. She had been 3 years post-menopausal under Hormonal Replacement Therapy. Weight: 46.7 Kg; Height: 1.60 m; Body Surface Area: 1.59 m2. Nil history of alcohol or tobacco smoking. Nilhistory suggestive of malignancies or autoimmune diseases. Her Blood group was oRhpositive, nil presence of irregular agglutinins. She was admitted to the neurology service for vertigo and it was determined an isolated cerebellar syndrome. All tests were negative including tumor markers and radiological imaging. The clinical gynecological examination was perfectly normal. The diagnosis hypothesis was “meningo-encephalocerebellitis of viral origin” but with persistence and aggravation of the cerebellar syndrome, despite treatment. We decided to search, antibodies, anti-Hu, anti-Yo, anti-Ri, and anti Ta. Anti Yo was positive + + + in the cerebrospinal fluid and serum of the patient. The search for a gynecological cancer included a mammography which revealed micro calcifications in the left breast + + +. A lumpectomy of the left breast accompanied with x-ray identification of the micro calcifications was done and the histology showed a High Grade Intraductal carcinoma of the left breast with two homes of 3mm and 1 mm, corresponding to an infiltrating carcinoma of the left breast, grade II tumor of Scarff and Bloom (SBRII, 21 N + / 26, RH +, low Ki 67) and Estrogen and progesterone receptor positive +: multifocal cancer. Following the lumpectomy, mastectomy withganglion clearing was done with adjuvant chemotherapy (FEC 6 Cycles): histology still showed Infiltrating Intraductal Carcinoma of the left breast, grade II tumor of Scarff and Bloom. Radiotherapy was followed and he patient was

  20. 1, 25(OH)2D3 Inhibits Hepatocellular Carcinoma Development Through Reducing Secretion of Inflammatory Cytokines from Immunocytes

    PubMed Central

    Guo, Jian; Ma, Zhenhua; Ma, Qingyong; Wu, Zheng; Fan, Ping; Zhou, Xiaojie; Chen, Lulu; Zhou, Shuang; Goltzman, David; Miao, Dengshun; Wu, Erxi

    2014-01-01

    Epidemiological and clinical studies have indicated that low vitamin D activity is not only associated with an increased cancer risk and a more aggressive tumor growth, but also connected with an aggravated liver damage caused by chronic inflammation. Meanwhile, increasing evidence has demonstrated that 1,25(OH)2D3 (the most biologically active metabolite of vitamin D) can inhibit inflammatory response in some chronic inflammatory associated cancer, which is considered to have the anti-tumor potency. However, the interaction between 1,25(OH)2D3 and inflammation during hepatocellular carcinoma (HCC) initiation and progression is not yet clear. Here, we report an anti-tumorigenesis effect of 1,25(OH)2D3 via decreasing inflammatory cytokine secretion in HCC and hypothesize the possible underlying mechanism. Firstly, we show that the enhanced tumor growth is associated with elevated inflammatory cytokine IL-6 and TNF-α in 1α(OH)ase gene-knockout mice. Secondly, 1,25(OH)2D3 can inhibit vitamin D receptor (VDR) shRNA interfered tumor cell growth through decreasing inflammatory cytokine secretion in vitro and in vivo. Finally, using p27kip1 gene knock-out mouse model, we demonstrate that the effect of 1,25(OH)2D3 in inhibiting immune cell related inflammatory cytokine secretion, exerts in a p27kip1 gene dependent way. Collectively, 1,25(OH)2D3 inhibits HCC development through up-regulating the expression of p27kip1 in immune cell and reducing inflammatory cytokine production. PMID:23992309

  1. Endogenous Voltage Potentials and the Microenvironment: Bioelectric Signals that Reveal, Induce and Normalize Cancer

    PubMed Central

    Chernet, Brook; Levin, Michael

    2014-01-01

    Cancer may be a disease of geometry: a misregulation of the field of information that orchestrates individual cells’ activities towards normal anatomy. Recent work identified molecular mechanisms underlying a novel system of developmental control: bioelectric gradients. Endogenous spatio-temporal differences in resting potential of non-neural cells provide instructive cues for cell regulation and complex patterning during embryogenesis and regeneration. It is now appreciated that these cues are an important layer of the dysregulation of cell: cell interactions that leads to cancer. Abnormal depolarization of resting potential (Vmem) is a convenient marker for neoplasia and activates a metastatic phenotype in genetically-normal cells in vivo. Moreover, oncogene expression depolarizes cells that form tumor-like structures, but is unable to form tumors if this depolarization is artificially prevented by misexpression of hyperpolarizing ion channels. Vmem triggers metastatic behaviors at considerable distance, mediated by transcriptional and epigenetic effects of electrically-modulated flows of serotonin and butyrate. While in vivo data on voltages in carcinogenesis comes mainly from the amphibian model, unbiased genetic screens and network profiling in rodents and human tissues reveal several ion channel proteins as bona fide oncogene and promising targets for cancer drug development. However, we propose that a focus on specific channel genes is just the tip of the iceberg. Bioelectric state is determined by post-translational gating of ion channels, not only from genetically-specified complements of ion translocators. A better model is a statistical dynamics view of spatial Vmem gradients. Cancer may not originate at the single cell level, since gap junctional coupling results in multi-cellular physiological networks with multiple stable attractors in bioelectrical state space. New medical applications await a detailed understanding of the mechanisms by which organ

  2. T-Cell-Specific Deletion of Map3k1 Reveals the Critical Role for Mekk1 and Jnks in Cdkn1b-Dependent Proliferative Expansion

    PubMed Central

    Suddason, Tesha; Anwar, Saba; Charlaftis, Nikolaos; Gallagher, Ewen

    2016-01-01

    Summary MAPK signaling is important for T lymphocyte development, homeostasis, and effector responses. To better understand the role of Mekk1 (encoded by Map3k1) in T cells, we conditionally deleted Map3k1 in LckCre/+Map3k1f/f mice, and these display larger iNKT cell populations within the liver, spleen, and bone marrow. Mekk1 signaling controls splenic and liver iNKT cell expansion in response to glycolipid antigen. LckCre/+Map3k1f/f mice have enhanced liver damage in response to glycolipid antigen. Mekk1 regulates Jnk activation in iNKT cells and binds and transfers Lys63-linked poly-ubiquitin onto Carma1. Map3k1 is critical for the regulation of p27Kip1 (encoded by Cdkn1b). PMID:26774476

  3. T-Cell-Specific Deletion of Map3k1 Reveals the Critical Role for Mekk1 and Jnks in Cdkn1b-Dependent Proliferative Expansion.

    PubMed

    Suddason, Tesha; Anwar, Saba; Charlaftis, Nikolaos; Gallagher, Ewen

    2016-01-26

    MAPK signaling is important for T lymphocyte development, homeostasis, and effector responses. To better understand the role of Mekk1 (encoded by Map3k1) in T cells, we conditionally deleted Map3k1 in Lck(Cre/+)Map3k1(f/f) mice, and these display larger iNKT cell populations within the liver, spleen, and bone marrow. Mekk1 signaling controls splenic and liver iNKT cell expansion in response to glycolipid antigen. Lck(Cre/+)Map3k1(f/f) mice have enhanced liver damage in response to glycolipid antigen. Mekk1 regulates Jnk activation in iNKT cells and binds and transfers Lys63-linked poly-ubiquitin onto Carma1. Map3k1 is critical for the regulation of p27(Kip1) (encoded by Cdkn1b). PMID:26774476

  4. Gene expression profiles of prostate cancer reveal involvement of multiple molecular pathways in the metastatic process

    PubMed Central

    Chandran, Uma R; Ma, Changqing; Dhir, Rajiv; Bisceglia, Michelle; Lyons-Weiler, Maureen; Liang, Wenjing; Michalopoulos, George; Becich, Michael; Monzon, Federico A

    2007-01-01

    Background Prostate cancer is characterized by heterogeneity in the clinical course that often does not correlate with morphologic features of the tumor. Metastasis reflects the most adverse outcome of prostate cancer, and to date there are no reliable morphologic features or serum biomarkers that can reliably predict which patients are at higher risk of developing metastatic disease. Understanding the differences in the biology of metastatic and organ confined primary tumors is essential for developing new prognostic markers and therapeutic targets. Methods Using Affymetrix oligonucleotide arrays, we analyzed gene expression profiles of 24 androgen-ablation resistant metastatic samples obtained from 4 patients and a previously published dataset of 64 primary prostate tumor samples. Differential gene expression was analyzed after removing potentially uninformative stromal genes, addressing the differences in cellular content between primary and metastatic tumors. Results The metastatic samples are highly heterogenous in expression; however, differential expression analysis shows that 415 genes are upregulated and 364 genes are downregulated at least 2 fold in every patient with metastasis. The expression profile of metastatic samples reveals changes in expression of a unique set of genes representing both the androgen ablation related pathways and other metastasis related gene networks such as cell adhesion, bone remodelling and cell cycle. The differentially expressed genes include metabolic enzymes, transcription factors such as Forkhead Box M1 (FoxM1) and cell adhesion molecules such as Osteopontin (SPP1). Conclusion We hypothesize that these genes have a role in the biology of metastatic disease and that they represent potential therapeutic targets for prostate cancer. PMID:17430594

  5. An acute adrenal insufficiency revealing pituitary metastases of lung cancer in an elderly patient.

    PubMed

    Marmouch, Hela; Arfa, Sondes; Mohamed, Saoussen Cheikh; Slim, Tensim; Khochtali, Ines

    2016-01-01

    Metastases of solid tumors to the pituitary gland are often asymptomatic or appereas as with diabetes insipid us. Pituitary metastases more commonly affect the posterior lobe and the infundibulum than the anterior lobe. The presentation with an acute adrenal insufficiency is a rare event. A 69-year-old men presented with vomiting, low blood pressure and hypoglycemia. Hormonal exploration confirmed a hypopituitarism. Appropriate therapy was initiated urgently. The hypothalamic-pituitary MRI showed a pituitary hypertrophy, a nodular thickening of the pituitary stalk. The chest X Rays revealed pulmonary opacity. Computed tomography scan of the chest showed a multiples tumors with mediastinal lymphadenopathy. Bronchoscopy and biopsy demonstrated a pulmonary adenocarcinoma. Hence we concluded to a lung cancer with multiple pituitary and adrenal gland metastases. This case emphasizes the need for an etiological investigation of acute adrenal insufficiency after treatment of acute phase. PMID:27200139

  6. Transcriptomic profiling of urine extracellular vesicles reveals alterations of CDH3 in prostate cancer

    PubMed Central

    Sanchez-Mosquera, Pilar; Ugalde-Olano, Aitziber; González, Esperanza; Cortazar, Ana R.; Palomo, Laura; Fernández-Ruiz, Sonia; Lacasa-Viscasillas, Isabel; Berdasco, Maria; Sutherland, James D.; Barrio, Rosa; Zabala-Letona, Amaia; Martín-Martín, Natalia; Arruabarrena-Aristorena, Amaia; Valcarcel-Jimenez, Lorea; Caro-Maldonado, Alfredo; Gonzalez-Tampan, Jorge; Cachi-Fuentes, Guido; Esteller, Manel; Aransay, Ana M.; Unda, Miguel

    2016-01-01

    Extracellular vesicles (EV) are emerging structures with promising properties for intercellular communication. In addition, the characterization of EV in biofluids is an attractive source of non-invasive diagnostic, prognostic and predictive biomarkers. Here we show that urinary EV (uEV) from prostate cancer (PCa) patients exhibit genuine and differential physical and biological properties compared to benign prostate hyperplasia (BPH). Importantly, transcriptomics characterization of uEVs led us to define the decreased abundance of Cadherin 3, type 1 (CDH3) transcript in uEV from PCa patients. Tissue and cell line analysis strongly suggested that the status of CDH3 in uEVs is a distal reflection of changes in the expression of this cadherin in the prostate tumor. CDH3 was negatively regulated at the genomic, transcriptional, and epigenetic level in PCa. Our results reveal that uEVs could represent a non-invasive tool to inform about the molecular alterations in PCa. PMID:26771841

  7. An acute adrenal insufficiency revealing pituitary metastases of lung cancer in an elderly patient

    PubMed Central

    Marmouch, Hela; Arfa, Sondes; Mohamed, Saoussen Cheikh; Slim, Tensim; Khochtali, Ines

    2016-01-01

    Metastases of solid tumors to the pituitary gland are often asymptomatic or appereas as with diabetes insipid us. Pituitary metastases more commonly affect the posterior lobe and the infundibulum than the anterior lobe. The presentation with an acute adrenal insufficiency is a rare event. A 69-year-old men presented with vomiting, low blood pressure and hypoglycemia. Hormonal exploration confirmed a hypopituitarism. Appropriate therapy was initiated urgently. The hypothalamic-pituitary MRI showed a pituitary hypertrophy, a nodular thickening of the pituitary stalk. The chest X Rays revealed pulmonary opacity. Computed tomography scan of the chest showed a multiples tumors with mediastinal lymphadenopathy. Bronchoscopy and biopsy demonstrated a pulmonary adenocarcinoma. Hence we concluded to a lung cancer with multiple pituitary and adrenal gland metastases. This case emphasizes the need for an etiological investigation of acute adrenal insufficiency after treatment of acute phase. PMID:27200139

  8. Model of fibrolamellar hepatocellular carcinomas reveals striking enrichment in cancer stem cells.

    PubMed

    Oikawa, Tsunekazu; Wauthier, Eliane; Dinh, Timothy A; Selitsky, Sara R; Reyna-Neyra, Andrea; Carpino, Guido; Levine, Ronald; Cardinale, Vincenzo; Klimstra, David; Gaudio, Eugenio; Alvaro, Domenico; Carrasco, Nancy; Sethupathy, Praveen; Reid, Lola M

    2015-01-01

    The aetiology of human fibrolamellar hepatocellular carcinomas (hFL-HCCs), cancers occurring increasingly in children to young adults, is poorly understood. We present a transplantable tumour line, maintained in immune-compromised mice, and validate it as a bona fide model of hFL-HCCs by multiple methods. RNA-seq analysis confirms the presence of a fusion transcript (DNAJB1-PRKACA) characteristic of hFL-HCC tumours. The hFL-HCC tumour line is highly enriched for cancer stem cells as indicated by limited dilution tumourigenicity assays, spheroid formation and flow cytometry. Immunohistochemistry on the hFL-HCC model, with parallel studies on 27 primary hFL-HCC tumours, provides robust evidence for expression of endodermal stem cell traits. Transcriptomic analyses of the tumour line and of multiple, normal hepatic lineage stages reveal a gene signature for hFL-HCCs closely resembling that of biliary tree stem cells--newly discovered precursors for liver and pancreas. This model offers unprecedented opportunities to investigate mechanisms underlying hFL-HCCs pathogenesis and potential therapies. PMID:26437858

  9. Spatiotemporal dynamics of intratumoral immune cells reveal the immune landscape in human cancer.

    PubMed

    Bindea, Gabriela; Mlecnik, Bernhard; Tosolini, Marie; Kirilovsky, Amos; Waldner, Maximilian; Obenauf, Anna C; Angell, Helen; Fredriksen, Tessa; Lafontaine, Lucie; Berger, Anne; Bruneval, Patrick; Fridman, Wolf Herman; Becker, Christoph; Pagès, Franck; Speicher, Michael R; Trajanoski, Zlatko; Galon, Jérôme

    2013-10-17

    The complex interactions between tumors and their microenvironment remain to be elucidated. Combining large-scale approaches, we examined the spatio-temporal dynamics of 28 different immune cell types (immunome) infiltrating tumors. We found that the immune infiltrate composition changed at each tumor stage and that particular cells had a major impact on survival. Densities of T follicular helper (Tfh) cells and innate cells increased, whereas most T cell densities decreased along with tumor progression. The number of B cells, which are key players in the core immune network and are associated with prolonged survival, increased at a late stage and showed a dual effect on recurrence and tumor progression. The immune control relevance was demonstrated in three endoscopic orthotopic colon-cancer mouse models. Genomic instability of the chemokine CXCL13 was a mechanism associated with Tfh and B cell infiltration. CXCL13 and IL21 were pivotal factors for the Tfh/B cell axis correlating with survival. This integrative study reveals the immune landscape in human colorectal cancer and the major hallmarks of the microenvironment associated with tumor progression and recurrence. PMID:24138885

  10. An Atlas of the Human Kinome Reveals the Mutational Landscape Underlying Dysregulated Phosphorylation Cascades in Cancer.

    PubMed

    Olow, Aleksandra; Chen, Zhongzhong; Niedner, R Hannes; Wolf, Denise M; Yau, Christina; Pankov, Aleksandr; Lee, Evelyn Pei Rong; Brown-Swigart, Lamorna; van 't Veer, Laura J; Coppé, Jean-Philippe

    2016-04-01

    Kinase inhibitors are used widely to treat various cancers, but adaptive reprogramming of kinase cascades and activation of feedback loop mechanisms often contribute to therapeutic resistance. Determining comprehensive, accurate maps of kinase circuits may therefore help elucidate mechanisms of response and resistance to kinase inhibitor therapies. In this study, we identified and validated phosphorylatable target sites across human cell and tissue types to generate PhosphoAtlas, a map of 1,733 functionally interconnected proteins comprising the human phospho-reactome. A systematic curation approach was used to distill protein phosphorylation data cross-referenced from 38 public resources. We demonstrated how a catalog of 2,617 stringently verified heptameric peptide regions at the catalytic interface of kinases and substrates could expose mutations that recurrently perturb specific phospho-hubs. In silico mapping of 2,896 nonsynonymous tumor variants identified from thousands of tumor tissues also revealed that normal and aberrant catalytic interactions co-occur frequently, showing how tumors systematically hijack, as well as spare, particular subnetworks. Overall, our work provides an important new resource for interrogating the human tumor kinome to strategically identify therapeutically actionable kinase networks that drive tumorigenesis. Cancer Res; 76(7); 1733-45. ©2016 AACR. PMID:26921330

  11. Genomic Profiling of Thyroid Cancer Reveals a Role for Thyroglobulin in Metastasis.

    PubMed

    Siraj, Abdul K; Masoodi, Tariq; Bu, Rong; Beg, Shaham; Al-Sobhi, Saif S; Al-Dayel, Fouad; Al-Dawish, Mohammed; Alkuraya, Fowzan S; Al-Kuraya, Khawla S

    2016-06-01

    Papillary thyroid carcinoma (PTC) has a wide geographic variation in incidence; it is most common in Saudi Arabia, where it is only second to breast cancer as the most common cancer among females. Genomic profiling of PTC from Saudi Arabia has not been attempted previously. We performed whole-exome sequencing of 101 PTC samples and the corresponding genomic DNA to identify genes with recurrent somatic mutations, then sequenced these genes by using a next-generation gene-panel approach in an additional 785 samples. In addition to BRAF, N-RAS, and H-RAS, which have previously been shown to be recurrently mutated in PTC, our analysis highlights additional genes, including thyroglobulin (TG), which harbored somatic mutations in 3% of the entire cohort. Surprisingly, although TG mutations were not exclusive to mutations in the RAS-MAP kinase pathway, their presence was associated with a significantly worse clinical outcome, which suggests a pathogenic role beyond driving initial oncogenesis. Analysis of metastatic PTC tissue revealed significant enrichment for TG mutations (p < 0.001), including events of apparent clonal expansion. Our results suggest a previously unknown role of TG somatic mutations in the pathogenesis of PTC and its malignant evolution. PMID:27236916

  12. An optimized isolation of biotinylated cell surface proteins reveals novel players in cancer metastasis

    PubMed Central

    Karhemo, Piia-Riitta; Ravela, Suvi; Laakso, Marko; Ritamo, Ilja; Tatti, Olga; Mäkinen, Selina; Goodison, Steve; Stenman, Ulf-Håkan; Hölttä, Erkki; Hautaniemi, Sampsa; Valmu, Leena; Lehti, Kaisa; Laakkonen, Pirjo

    2012-01-01

    Details of metastasis, the deadliest aspect of cancer, are unclear. Cell surface proteins play central roles in adhesive contacts between the tumor cell and the stroma during metastasis. We optimized a fast, small-scale isolation of biotinylated cell surface proteins to reveal novel metastasis-associated players froman isogenic pair of human MDA-MB-435 cancer cells with opposite metastatic phenotypes. Isolated proteins were trypsin digested and analyzed using LC–MS/MS followed by quantitation with the Progenesis LC–MS software. Sixteen proteins displayed over twofold expression differences between the metastatic and non-metastatic cells. Interestingly, overexpression of most of them (14/16) in the metastatic cells indicates a gain of novel surface protein profile as compared to the non-metastatic one. All five validated, differentially expressed proteins showed higher expression in the metastatic cells in culture, and four of these were further validated in vivo. Moreover, we analyzed the expression of two of the identified proteins, CD109 and ITGA6 in 3-dimensional cultures of six melanoma cell lines. Both proteins marked the surface of cells derived from melanoma metastasis over cells derived from primary melanoma. These unbiased identification and validation of both known and novel metastasis-associated proteins indicate a reliable approach for the identification of differentially expressed surface proteins. PMID:22813880

  13. Revealing potential molecular targets bridging colitis and colorectal cancer based on multidimensional integration strategy

    PubMed Central

    Hu, Yongfei; Li, Xiaobo; Wang, Xishan; Fan, Huihui; Wang, Guiyu; Wang, Dong

    2015-01-01

    Chronic inflammation may play a vital role in the pathogenesis of inflammation-associated tumors. However, the underlying mechanisms bridging ulcerative colitis (UC) and colorectal cancer (CRC) remain unclear. Here, we integrated multidimensional interaction resources, including gene expression profiling, protein-protein interactions (PPIs), transcriptional and post-transcriptional regulation data, and virus-host interactions, to tentatively explore potential molecular targets that functionally link UC and CRC at a systematic level. In this work, by deciphering the overlapping genes, crosstalking genes and pivotal regulators of both UC- and CRC-associated functional module pairs, we revealed a variety of genes (including FOS and DUSP1, etc.), transcription factors (including SMAD3 and ETS1, etc.) and miRNAs (including miR-155 and miR-196b, etc.) that may have the potential to complete the connections between UC and CRC. Interestingly, further analyses of the virus-host interaction network demonstrated that several virus proteins (including EBNA-LP of EBV and protein E7 of HPV) frequently inter-connected to UC- and CRC-associated module pairs with their validated targets significantly enriched in both modules of the host. Together, our results suggested that multidimensional integration strategy provides a novel approach to discover potential molecular targets that bridge the connections between UC and CRC, which could also be extensively applied to studies on other inflammation-related cancers. PMID:26461477

  14. Model of fibrolamellar hepatocellular carcinomas reveals striking enrichment in cancer stem cells

    PubMed Central

    Oikawa, Tsunekazu; Wauthier, Eliane; Dinh, Timothy A.; Selitsky, Sara R.; Reyna-Neyra, Andrea; Carpino, Guido; Levine, Ronald; Cardinale, Vincenzo; Klimstra, David; Gaudio, Eugenio; Alvaro, Domenico; Carrasco, Nancy; Sethupathy, Praveen; Reid, Lola M.

    2015-01-01

    The aetiology of human fibrolamellar hepatocellular carcinomas (hFL-HCCs), cancers occurring increasingly in children to young adults, is poorly understood. We present a transplantable tumour line, maintained in immune-compromised mice, and validate it as a bona fide model of hFL-HCCs by multiple methods. RNA-seq analysis confirms the presence of a fusion transcript (DNAJB1-PRKACA) characteristic of hFL-HCC tumours. The hFL-HCC tumour line is highly enriched for cancer stem cells as indicated by limited dilution tumourigenicity assays, spheroid formation and flow cytometry. Immunohistochemistry on the hFL-HCC model, with parallel studies on 27 primary hFL-HCC tumours, provides robust evidence for expression of endodermal stem cell traits. Transcriptomic analyses of the tumour line and of multiple, normal hepatic lineage stages reveal a gene signature for hFL-HCCs closely resembling that of biliary tree stem cells—newly discovered precursors for liver and pancreas. This model offers unprecedented opportunities to investigate mechanisms underlying hFL-HCCs pathogenesis and potential therapies. PMID:26437858

  15. Novel tumor suppressor candidates on chromosome 3 revealed by NotI-microarrays in cervical cancer

    PubMed Central

    Senchenko, Vera N.; Kisseljova, Natalia P.; Ivanova, Tatyana A.; Dmitriev, Alexey A.; Krasnov, George S.; Kudryavtseva, Anna V.; Panasenko, Grigory V.; Tsitrin, Evgeny B.; Lerman, Michael I.; Kisseljov, Fyodor L.; Kashuba, Vladimir I.; Zabarovsky, Eugene R.

    2013-01-01

    Genetic and epigenetic alterations in cervical carcinomas were investigated using NotI-microarrays containing 180 cloned sequences flanking all NotI-sites associated with genes on chromosome 3. In total, 48 paired normal/tumor DNA samples, specifically enriched in NotI-sites, were hybridized to NotI-microarrays. Thirty genes, including tumor suppressors or candidates (for example, VHL, RBSP3/CTDSPL, ITGA9, LRRC3B, ALDH1L1, EPHB1) and genes previously unknown as cancer-associated (ABHD5, C3orf77, PRL32, LOC285375, FGD5 and others), showed methylation/deletion in 21–44% of tumors. The genes were more frequently altered in squamous cell carcinomas (SCC) than in adenocarcinomas (ADC, p < 0.01). A set of seven potential markers (LRRN1, PRICKLE2, VHL, BHLHE40, RBSP3, CGGBP1 and SOX14) is promising for discrimination of ADC and SCC. Alterations of more than 20 genes simultaneously were revealed in 23% of SCC. Bisulfite sequencing analysis confirmed methylation as a frequent event in SCC. High down-regulation frequency was shown for RBSP3, ITGA9, VILL, APRG1/C3orf35 and RASSF1 (isoform A) genes (3p21.3 locus) in SCC. Both frequency and extent of RASSF1A and RBSP3 mRNA level decrease were more pronounced in tumors with lymph node metastases compared with non-metastatic ones (p ≤ 0.05). We confirmed by bisulfite sequencing that RASSF1 promoter methylation was a rare event in SCC and, for the first time, demonstrated RASSF1A down-regulation at both the mRNA and protein levels without promoter methylation in tumors of this histological type. Thus, our data revealed novel tumor suppressor candidates located on chromosome 3 and a frequent loss of epigenetic stability of 3p21.3 locus in combination with down-regulation of genes in cervical cancer. PMID:23478628

  16. Catabolism of Exogenous Lactate Reveals It as a Legitimate Metabolic Substrate in Breast Cancer

    PubMed Central

    Kennedy, Kelly M.; Scarbrough, Peter M.; Ribeiro, Anthony; Richardson, Rachel; Yuan, Hong; Sonveaux, Pierre; Landon, Chelsea D.; Chi, Jen-Tsan; Pizzo, Salvatore

    2013-01-01

    Lactate accumulation in tumors has been associated with metastases and poor overall survival in cancer patients. Lactate promotes angiogenesis and metastasis, providing rationale for understanding how it is processed by cells. The concentration of lactate in tumors is a balance between the amount produced, amount carried away by vasculature and if/how it is catabolized by aerobic tumor or stromal cells. We examined lactate metabolism in human normal and breast tumor cell lines and rat breast cancer: 1. at relevant concentrations, 2. under aerobic vs. hypoxic conditions, 3. under conditions of normo vs. hypoglucosis. We also compared the avidity of tumors for lactate vs. glucose and identified key lactate catabolites to reveal how breast cancer cells process it. Lactate was non-toxic at clinically relevant concentrations. It was taken up and catabolized to alanine and glutamate by all cell lines. Kinetic uptake rates of lactate in vivo surpassed that of glucose in R3230Ac mammary carcinomas. The uptake appeared specific to aerobic tumor regions, consistent with the proposed “metabolic symbiont” model; here lactate produced by hypoxic cells is used by aerobic cells. We investigated whether treatment with alpha-cyano-4-hydroxycinnamate (CHC), a MCT1 inhibitor, would kill cells in the presence of high lactate. Both 0.1 mM and 5 mM CHC prevented lactate uptake in R3230Ac cells at lactate concentrations at ≤20 mM but not at 40 mM. 0.1 mM CHC was well-tolerated by R3230Ac and MCF7 cells, but 5 mM CHC killed both cell lines ± lactate, indicating off-target effects. This study showed that breast cancer cells tolerate and use lactate at clinically relevant concentrations in vitro (± glucose) and in vivo. We provided additional support for the metabolic symbiont model and discovered that breast cells prevailingly take up and catabolize lactate, providing rationale for future studies on manipulation of lactate catabolism pathways for therapy. PMID:24069390

  17. Exome sequencing of a colorectal cancer family reveals shared mutation pattern and predisposition circuitry along tumor pathways

    PubMed Central

    Suleiman, Suleiman H.; Koko, Mahmoud E.; Nasir, Wafaa H.; Elfateh, Ommnyiah; Elgizouli, Ubai K.; Abdallah, Mohammed O. E.; Alfarouk, Khalid O.; Hussain, Ayman; Faisal, Shima; Ibrahim, Fathelrahamn M. A.; Romano, Maurizio; Sultan, Ali; Banks, Lawrence; Newport, Melanie; Baralle, Francesco; Elhassan, Ahmed M.; Mohamed, Hiba S.; Ibrahim, Muntaser E.

    2015-01-01

    The molecular basis of cancer and cancer multiple phenotypes are not yet fully understood. Next Generation Sequencing promises new insight into the role of genetic interactions in shaping the complexity of cancer. Aiming to outline the differences in mutation patterns between familial colorectal cancer cases and controls we analyzed whole exomes of cancer tissues and control samples from an extended colorectal cancer pedigree, providing one of the first data sets of exome sequencing of cancer in an African population against a background of large effective size typically with excess of variants. Tumors showed hMSH2 loss of function SNV consistent with Lynch syndrome. Sets of genes harboring insertions–deletions in tumor tissues revealed, however, significant GO enrichment, a feature that was not seen in control samples, suggesting that ordered insertions–deletions are central to tumorigenesis in this type of cancer. Network analysis identified multiple hub genes of centrality. ELAVL1/HuR showed remarkable centrality, interacting specially with genes harboring non-synonymous SNVs thus reinforcing the proposition of targeted mutagenesis in cancer pathways. A likely explanation to such mutation pattern is DNA/RNA editing, suggested here by nucleotide transition-to-transversion ratio that significantly departed from expected values (p-value 5e-6). NFKB1 also showed significant centrality along with ELAVL1, raising the suspicion of viral etiology given the known interaction between oncogenic viruses and these proteins. PMID:26442106

  18. Human T-cell leukemia virus type 1 infection leads to arrest in the G1 phase of the cell cycle.

    PubMed

    Liu, Meihong; Yang, Liangpeng; Zhang, Ling; Liu, Baoying; Merling, Randall; Xia, Zheng; Giam, Chou-Zen

    2008-09-01

    Infection by the human T-cell leukemia virus type 1 (HTLV-1) is thought to cause dysregulated T-cell proliferation, which in turn leads to adult T-cell leukemia/lymphoma. Early cellular changes after HTLV-1 infection have been difficult to study due to the poorly infectious nature of HTLV-1 and the need for cell-to-cell contact for HTLV-1 transmission. Using a series of reporter systems, we show that HeLa cells cease proliferation within one or two division cycles after infection by HTLV-1 or transduction of the HTLV-1 tax gene. HTLV-1-infected HeLa cells, like their tax-transduced counterparts, expressed high levels of p21(CIP1/WAF1) and p27(KIP1), developed mitotic abnormalities, and became arrested in G(1) in senescence. In contrast, cells of a human osteosarcoma lineage (HOS) continued to divide after HTLV-1 infection or Tax expression, albeit at a reduced growth rate and with mitotic aberrations. Unique to HOS cells is the dramatic reduction of p21(CIP1/WAF1) and p27(KIP1) expression, which is in part associated with the constitutive activation of the phosphatidylinositol-3-kinase (PI3K)-protein kinase B (Akt) pathway. The loss of p21(CIP1/WAF1) and p27(KIP1) in HOS cells apparently allows HTLV-1- and Tax-induced G(1) arrest to be bypassed. Finally, HTLV-1 infection and Tax expression also cause human SupT1 T cells to arrest in the G(1) phase of the cell cycle. These results suggest that productive HTLV-1 infection ordinarily leads to Tax-mediated G(1) arrest. However, T cells containing somatic mutations that inactivate p21(CIP1/WAF1) and p27(KIP1) may continue to proliferate after HTLV-1 infection and Tax expression. These infected cells can expand clonally, accumulate additional chromosomal abnormalities, and progress to cancer. PMID:18596104

  19. Molecular portrait of breast cancer in China reveals comprehensive transcriptomic likeness to Caucasian breast cancer and low prevalence of luminal A subtype

    PubMed Central

    Huang, Xiaoyan; Dugo, Matteo; Callari, Maurizio; Sandri, Marco; De Cecco, Loris; Valeri, Barbara; Carcangiu, Maria Luisa; Xue, Jingyan; Bi, Rui; Veneroni, Silvia; Daidone, Maria Grazia; Ménard, Sylvie; Tagliabue, Elda; Shao, Zhimin; Wu, Jiong; Orlandi, Rosaria

    2015-01-01

    The recent dramatic increase in breast cancer incidence across China with progressive urbanization and economic development has signaled the urgent need for molecular and clinical detailing of breast cancer in the Chinese population. Our analyses of a unique transethnic collection of breast cancer frozen specimens from Shanghai Fudan Cancer Center (Chinese Han) profiled simultaneously with an analogous Caucasian Italian series revealed consistent transcriptomic data lacking in batch effects. The prevalence of Luminal A subtype was significantly lower in Chinese series, impacting the overall prevalence of estrogen receptor (ER)-positive disease in a large cohort of Chinese/Caucasian patients. Unsupervised and supervised comparison of gene and microRNA (miRNA) profiles of Chinese and Caucasian samples revealed extensive similarity in the comprehensive taxonomy of transcriptional elements regulating breast cancer biology. Partition of gene expression data using gene lists relevant to breast cancer as “intrinsic” and “extracellular matrix” genes identified Chinese and Caucasian subgroups with equivalent global gene and miRNA profiles. These findings indicate that in the Chinese and Caucasian groups, breast neoplasia and the surrounding stromal characteristics undergo the same differentiation and molecular processes. Transcriptional similarity across transethnic cohorts may simplify translational medicine approaches and clinical management of breast cancer patients worldwide. PMID:25787708

  20. CDA directs metabolism of epigenetic nucleosides revealing a therapeutic window in cancer

    PubMed Central

    Zauri, Melania; Berridge, Georgina; Thézénas, Marie-Laëtitia; Pugh, Kathryn M.; Goldin, Robert; Kessler, Benedikt M.; Kriaucionis, Skirmantas

    2015-01-01

    Summary Cells require nucleotides to support DNA replication and to repair damaged DNA. In addition to de novo synthesis, cells recycle nucleotides from the DNA of dying cells or from cellular material ingested through the diet. Salvaged nucleosides come with the complication that they can contain epigenetic modifications. Since epigenetic inheritance of DNA methylation mainly relies on copying of the modification pattern from parental strands1-3, random incorporation of pre-modified bases during replication could have profound implications for epigenome fidelity and yield adverse cellular phenotypes. Although the salvage mechanism of 5-methyl-2′deoxycytidine (5mdC) has been investigated before4-6, currently it remains unknown how cells deal with the recently identified oxidised forms of 5mdC – 5-hydroxymethyl-2′deoxycytidine (5hmdC), 5-formy-2′deoxycytidine (5fdC) and 5-carboxyl-2′deoxycytidine (5cadC)7-10. Here we demonstrate that enzymes of the nucleotide salvage pathway display substrate selectivity, effectively protecting newly synthesized DNA from the incorporation of epigenetically modified forms of cytosine. Thus cell lines and animals can tolerate high doses of these modified cytidines without any deleterious effects on physiology. Interestingly, by screening cancer cell lines for growth defects following exposure to 5hmdC, we unexpectedly identify a subset of cell lines where 5hmdC or 5fdC administration leads to cell lethality. Using genomic approaches we discover that the susceptible cell lines overexpress cytidine deaminase (CDA). CDA converts 5hmdC and 5fdC into variants of uridine that are incorporated into DNA, resulting in accumulation of DNA damage and ultimately, cell death. Our observations extend current knowledge of the nucleotide salvage pathway by revealing the metabolism of oxidised epigenetic bases, and suggest a therapeutic option for cancers, such as pancreatic cancer, that have CDA overexpression and are resistant to treatment

  1. Cross-Cancer Genome-Wide Analysis of Lung, Ovary, Breast, Prostate, and Colorectal Cancer Reveals Novel Pleiotropic Associations.

    PubMed

    Fehringer, Gordon; Kraft, Peter; Pharoah, Paul D; Eeles, Rosalind A; Chatterjee, Nilanjan; Schumacher, Fredrick R; Schildkraut, Joellen M; Lindström, Sara; Brennan, Paul; Bickeböller, Heike; Houlston, Richard S; Landi, Maria Teresa; Caporaso, Neil; Risch, Angela; Amin Al Olama, Ali; Berndt, Sonja I; Giovannucci, Edward L; Grönberg, Henrik; Kote-Jarai, Zsofia; Ma, Jing; Muir, Kenneth; Stampfer, Meir J; Stevens, Victoria L; Wiklund, Fredrik; Willett, Walter C; Goode, Ellen L; Permuth, Jennifer B; Risch, Harvey A; Reid, Brett M; Bezieau, Stephane; Brenner, Hermann; Chan, Andrew T; Chang-Claude, Jenny; Hudson, Thomas J; Kocarnik, Jonathan K; Newcomb, Polly A; Schoen, Robert E; Slattery, Martha L; White, Emily; Adank, Muriel A; Ahsan, Habibul; Aittomäki, Kristiina; Baglietto, Laura; Blomquist, Carl; Canzian, Federico; Czene, Kamila; Dos-Santos-Silva, Isabel; Eliassen, A Heather; Figueroa, Jonine D; Flesch-Janys, Dieter; Fletcher, Olivia; Garcia-Closas, Montserrat; Gaudet, Mia M; Johnson, Nichola; Hall, Per; Hazra, Aditi; Hein, Rebecca; Hofman, Albert; Hopper, John L; Irwanto, Astrid; Johansson, Mattias; Kaaks, Rudolf; Kibriya, Muhammad G; Lichtner, Peter; Liu, Jianjun; Lund, Eiliv; Makalic, Enes; Meindl, Alfons; Müller-Myhsok, Bertram; Muranen, Taru A; Nevanlinna, Heli; Peeters, Petra H; Peto, Julian; Prentice, Ross L; Rahman, Nazneen; Sanchez, Maria Jose; Schmidt, Daniel F; Schmutzler, Rita K; Southey, Melissa C; Tamimi, Rulla; Travis, Ruth C; Turnbull, Clare; Uitterlinden, Andre G; Wang, Zhaoming; Whittemore, Alice S; Yang, Xiaohong R; Zheng, Wei; Buchanan, Daniel D; Casey, Graham; Conti, David V; Edlund, Christopher K; Gallinger, Steven; Haile, Robert W; Jenkins, Mark; Le Marchand, Loïc; Li, Li; Lindor, Noralene M; Schmit, Stephanie L; Thibodeau, Stephen N; Woods, Michael O; Rafnar, Thorunn; Gudmundsson, Julius; Stacey, Simon N; Stefansson, Kari; Sulem, Patrick; Chen, Y Ann; Tyrer, Jonathan P; Christiani, David C; Wei, Yongyue; Shen, Hongbing; Hu, Zhibin; Shu, Xiao-Ou; Shiraishi, Kouya; Takahashi, Atsushi; Bossé, Yohan; Obeidat, Ma'en; Nickle, David; Timens, Wim; Freedman, Matthew L; Li, Qiyuan; Seminara, Daniela; Chanock, Stephen J; Gong, Jian; Peters, Ulrike; Gruber, Stephen B; Amos, Christopher I; Sellers, Thomas A; Easton, Douglas F; Hunter, David J; Haiman, Christopher A; Henderson, Brian E; Hung, Rayjean J

    2016-09-01

    Identifying genetic variants with pleiotropic associations can uncover common pathways influencing multiple cancers. We took a two-stage approach to conduct genome-wide association studies for lung, ovary, breast, prostate, and colorectal cancer from the GAME-ON/GECCO Network (61,851 cases, 61,820 controls) to identify pleiotropic loci. Findings were replicated in independent association studies (55,789 cases, 330,490 controls). We identified a novel pleiotropic association at 1q22 involving breast and lung squamous cell carcinoma, with eQTL analysis showing an association with ADAM15/THBS3 gene expression in lung. We also identified a known breast cancer locus CASP8/ALS2CR12 associated with prostate cancer, a known cancer locus at CDKN2B-AS1 with different variants associated with lung adenocarcinoma and prostate cancer, and confirmed the associations of a breast BRCA2 locus with lung and serous ovarian cancer. This is the largest study to date examining pleiotropy across multiple cancer-associated loci, identifying common mechanisms of cancer development and progression. Cancer Res; 76(17); 5103-14. ©2016 AACR. PMID:27197191

  2. A common promoter hypomethylation signature in invasive breast, liver and prostate cancer cell lines reveals novel targets involved in cancer invasiveness

    PubMed Central

    Yi, Cao; Li, Chen Chen; Yu, Patricia; Arakelian, Ani; Tanvir, Imrana; Khan, Haseeb Ahmed; Rabbani, Shafaat

    2015-01-01

    Cancer invasion and metastasis is the most morbid aspect of cancer and is governed by different cellular mechanisms than those driving the deregulated growth of tumors. We addressed here the question of whether a common DNA methylation signature of invasion exists in cancer cells from different origins that differentiates invasive from non-invasive cells. We identified a common DNA methylation signature consisting of hyper- and hypomethylation and determined the overlap of differences in DNA methylation with differences in mRNA expression using expression array analyses. A pathway analysis reveals that the hypomethylation signature includes some of the major pathways that were previously implicated in cancer migration and invasion such as TGF beta and ERBB2 triggered pathways. The relevance of these hypomethylation events in human tumors was validated by identification of the signature in several publicly available databases of human tumor transcriptomes. We shortlisted novel invasion promoting candidates and tested the role of four genes in cellular invasiveness from the list C11orf68, G0S2, SHISA2 and TMEM156 in invasiveness using siRNA depletion. Importantly these genes are upregulated in human cancer specimens as determined by immunostaining of human normal and cancer breast, liver and prostate tissue arrays. Since these genes are activated in cancer they constitute a group of targets for specific pharmacological inhibitors of cancer invasiveness. SUMMARY Our study provides evidence that common DNA hypomethylation signature exists between cancer cells derived from different tissues, pointing to a common mechanism of cancer invasiveness in cancer cells from different origins that could serve as drug targets. PMID:26427334

  3. Aberrant gene expression in mucosa adjacent to tumor reveals a molecular crosstalk in colon cancer

    PubMed Central

    2014-01-01

    Background A colorectal tumor is not an isolated entity growing in a restricted location of the body. The patient’s gut environment constitutes the framework where the tumor evolves and this relationship promotes and includes a complex and tight correlation of the tumor with inflammation, blood vessels formation, nutrition, and gut microbiome composition. The tumor influence in the environment could both promote an anti-tumor or a pro-tumor response. Methods A set of 98 paired adjacent mucosa and tumor tissues from colorectal cancer (CRC) patients and 50 colon mucosa from healthy donors (246 samples in total) were included in this work. RNA extracted from each sample was hybridized in Affymetrix chips Human Genome U219. Functional relationships between genes were inferred by means of systems biology using both transcriptional regulation networks (ARACNe algorithm) and protein-protein interaction networks (BIANA software). Results Here we report a transcriptomic analysis revealing a number of genes activated in adjacent mucosa from CRC patients, not activated in mucosa from healthy donors. A functional analysis of these genes suggested that this active reaction of the adjacent mucosa was related to the presence of the tumor. Transcriptional and protein-interaction networks were used to further elucidate this response of normal gut in front of the tumor, revealing a crosstalk between proteins secreted by the tumor and receptors activated in the adjacent colon tissue; and vice versa. Remarkably, Slit family of proteins activated ROBO receptors in tumor whereas tumor-secreted proteins transduced a cellular signal finally activating AP-1 in adjacent tissue. Conclusions The systems-level approach provides new insights into the micro-ecology of colorectal tumorogenesis. Disrupting this intricate molecular network of cell-cell communication and pro-inflammatory microenvironment could be a therapeutic target in CRC patients. PMID:24597571

  4. In Search of a Cure for Proteostasis-Addicted Cancer: A AAA Target Revealed.

    PubMed

    Xia, Di; Ye, Yihong

    2015-11-01

    Tumorigenesis is often associated with an unbalanced protein homeostasis (proteostasis) network, which sensitizes cancer cells to drugs targeting protein quality control (PQC) regulators. In this issue of Cancer Cell, Anderson and colleagues investigated the anti-cancer activity of a new class of inhibitor against a multi-functional ATPase essential for proteostasis maintenance. PMID:26555170

  5. Differential secretome analysis reveals CST6 as a suppressor of breast cancer bone metastasis

    PubMed Central

    Jin, Lei; Zhang, Yan; Li, Hui; Yao, Ling; Fu, Da; Yao, Xuebiao; Xu, Lisa X; Hu, Xiaofang; Hu, Guohong

    2012-01-01

    Bone metastasis is a frequent complication of breast cancer and a common cause of morbidity and mortality from the disease. During metastasis secreted proteins play crucial roles in the interactions between cancer cells and host stroma. To characterize the secreted proteins that are associated with breast cancer bone metastasis, we preformed a label-free proteomic analysis to compare the secretomes of four MDA-MB-231 (MDA231) derivative cell lines with varied capacities of bone metastasis. A total of 128 proteins were found to be consistently up-/down-regulated in the conditioned medium of bone-tropic cancer cells. The enriched molecular functions of the altered proteins included receptor binding and peptidase inhibition. Through additional transcriptomic analyses of breast cancer cells, we selected cystatin E/M (CST6), a cysteine protease inhibitor down-regulated in bone-metastatic cells, for further functional studies. Our results showed that CST6 suppressed the proliferation, colony formation, migration and invasion of breast cancer cells. The suppressive function against cancer cell motility was carried out by cancer cell-derived soluble CST6. More importantly, ectopic expression of CST6 in cancer cells rescued mice from overt osteolytic metastasis and deaths in the animal study, while CST6 knockdown markedly enhanced cancer cell bone metastasis and shortened animal survival. Overall, our study provided a systemic secretome analysis of breast cancer bone tropism and established secreted CST6 as a bona fide suppressor of breast cancer osteolytic metastasis. PMID:22688893

  6. High-Resolution CRISPR Screens Reveal Fitness Genes and Genotype-Specific Cancer Liabilities.

    PubMed

    Hart, Traver; Chandrashekhar, Megha; Aregger, Michael; Steinhart, Zachary; Brown, Kevin R; MacLeod, Graham; Mis, Monika; Zimmermann, Michal; Fradet-Turcotte, Amelie; Sun, Song; Mero, Patricia; Dirks, Peter; Sidhu, Sachdev; Roth, Frederick P; Rissland, Olivia S; Durocher, Daniel; Angers, Stephane; Moffat, Jason

    2015-12-01

    The ability to perturb genes in human cells is crucial for elucidating gene function and holds great potential for finding therapeutic targets for diseases such as cancer. To extend the catalog of human core and context-dependent fitness genes, we have developed a high-complexity second-generation genome-scale CRISPR-Cas9 gRNA library and applied it to fitness screens in five human cell lines. Using an improved Bayesian analytical approach, we consistently discover 5-fold more fitness genes than were previously observed. We present a list of 1,580 human core fitness genes and describe their general properties. Moreover, we demonstrate that context-dependent fitness genes accurately recapitulate pathway-specific genetic vulnerabilities induced by known oncogenes and reveal cell-type-specific dependencies for specific receptor tyrosine kinases, even in oncogenic KRAS backgrounds. Thus, rigorous identification of human cell line fitness genes using a high-complexity CRISPR-Cas9 library affords a high-resolution view of the genetic vulnerabilities of a cell. PMID:26627737

  7. Photoactivation of Mutant Isocitrate Dehydrogenase 2 Reveals Rapid Cancer-Associated Metabolic and Epigenetic Changes

    PubMed Central

    2016-01-01

    Isocitrate dehydrogenase is mutated at a key active site arginine residue (Arg172 in IDH2) in many cancers, leading to the synthesis of the oncometabolite (R)-2-hydroxyglutarate (2HG). To investigate the early events following acquisition of this mutation in mammalian cells we created a photoactivatable version of IDH2(R172K), in which K172 is replaced with a photocaged lysine (PCK), via genetic code expansion. Illumination of cells expressing this mutant protein led to a rapid increase in the levels of 2HG, with 2HG levels reaching those measured in patient tumor samples, within 8 h. 2HG accumulation is closely followed by a global decrease in 5-hydroxymethylcytosine (5-hmC) in DNA, demonstrating that perturbations in epigenetic DNA base modifications are an early consequence of mutant IDH2 in cells. Our results provide a paradigm for rapidly and synchronously uncloaking diverse oncogenic mutations in live cells to reveal the sequence of events through which they may ultimately cause transformation. PMID:26761588

  8. Photoactivation of Mutant Isocitrate Dehydrogenase 2 Reveals Rapid Cancer-Associated Metabolic and Epigenetic Changes.

    PubMed

    Walker, Olivia S; Elsässer, Simon J; Mahesh, Mohan; Bachman, Martin; Balasubramanian, Shankar; Chin, Jason W

    2016-01-27

    Isocitrate dehydrogenase is mutated at a key active site arginine residue (Arg172 in IDH2) in many cancers, leading to the synthesis of the oncometabolite (R)-2-hydroxyglutarate (2HG). To investigate the early events following acquisition of this mutation in mammalian cells we created a photoactivatable version of IDH2(R172K), in which K172 is replaced with a photocaged lysine (PCK), via genetic code expansion. Illumination of cells expressing this mutant protein led to a rapid increase in the levels of 2HG, with 2HG levels reaching those measured in patient tumor samples, within 8 h. 2HG accumulation is closely followed by a global decrease in 5-hydroxymethylcytosine (5-hmC) in DNA, demonstrating that perturbations in epigenetic DNA base modifications are an early consequence of mutant IDH2 in cells. Our results provide a paradigm for rapidly and synchronously uncloaking diverse oncogenic mutations in live cells to reveal the sequence of events through which they may ultimately cause transformation. PMID:26761588

  9. Transcriptomic and CRISPR/Cas9 technologies reveal FOXA2 as a tumor suppressor gene in pancreatic cancer.

    PubMed

    Vorvis, Christina; Hatziapostolou, Maria; Mahurkar-Joshi, Swapna; Koutsioumpa, Marina; Williams, Jennifer; Donahue, Timothy R; Poultsides, George A; Eibl, Guido; Iliopoulos, Dimitrios

    2016-06-01

    Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with low survival rates and limited therapeutic options. Thus elucidation of signaling pathways involved in PDAC pathogenesis is essential for identifying novel potential therapeutic gene targets. Here, we used a systems approach to elucidate those pathways by integrating gene and microRNA profiling analyses together with CRISPR/Cas9 technology to identify novel transcription factors involved in PDAC pathogenesis. FOXA2 transcription factor was found to be significantly downregulated in PDAC relative to control pancreatic tissues. Functional experiments revealed that FOXA2 has a tumor suppressor function through inhibition of pancreatic cancer cell growth, migration, invasion, and colony formation. In situ hybridization analysis revealed miR-199a to be significantly upregulated in pancreatic cancer. Bioinformatics and luciferase analyses showed that miR-199a negatively but directly regulates FOXA2 expression through binding in its 3'-untranslated region (UTR). Evaluation of the functional importance of miR-199a on pancreatic cancer revealed that miR-199a acts as an inhibitor of FOXA2 expression, inducing an increase in pancreatic cancer cell proliferation, migration, and invasion. Additionally, gene ontology and network analyses in PANC-1 cells treated with a small interfering RNA (siRNA) against FOXA2 revealed an enrichment for cell invasion mechanisms through PLAUR and ERK activation. FOXA2 deletion (FOXA2Δ) by using two CRISPR/Cas9 vectors in PANC-1 cells induced tumor growth in vivo resulting in upregulation of PLAUR and ERK pathways in FOXA2Δ xenograft tumors. We have identified FOXA2 as a novel tumor suppressor in pancreatic cancer and it is regulated directly by miR-199a, thereby enhancing our understanding of how microRNAs interplay with the transcription factors to affect pancreatic oncogenesis. PMID:27151939

  10. Metabolic Profiling Reveals PAFAH1B3 as a Critical Driver of Breast Cancer Pathogenicity

    PubMed Central

    Mulvihill, Melinda M.; Benjamin, Daniel I.; Ji, Xiaodan; Le Scolan, Erwan; Louie, Sharon M.; Shieh, Alice; Green, McKenna; Narasimhalu, Tara; Morris, Patrick J.; Luo, Kunxin; Nomura, Daniel K.

    2014-01-01

    Many studies have identified metabolic pathways that underlie cellular transformation, but the metabolic drivers of cancer progression remain less well understood. The Hippo transducer pathway has been shown to confer malignant traits on breast cancer cells. In this study, we used metabolic mapping platforms to identify biochemical drivers of cellular transformation and malignant progression driven through RAS and the Hippo pathway in breast cancer, and identified platelet activating factor acetylhydrolase 1B3 (PAFAH1B3) as a key metabolic driver of breast cancer pathogenicity that is upregulated in primary human breast tumors and correlated with poor prognosis. Metabolomic profiling suggests that PAFAH1B3 inactivation attenuates cancer pathogenicity through enhancing tumor-suppressing signaling lipids. Our studies provide a map of altered metabolism that underlies breast cancer progression and put forth PAFAH1B3 as a critical metabolic node in breast cancer. PMID:24954006