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Sample records for oncofetal h19-derived mir-675

  1. The long non-coding RNA H19-derived miR-675 modulates human gastric cancer cell proliferation by targeting tumor suppressor RUNX1

    SciTech Connect

    Zhuang, Ming; Gao, Wen; Xu, Jing; Wang, Ping; Shu, Yongqian

    2014-06-06

    Graphical abstract: - Highlights: • H19 regulates gastric cancer cell proliferation phenotype via miR-675. • MiR-675 modulates cell proliferation of gastric cancer cells by targeting tumor suppressor RUNX1. • The H19/miR-675/RUNX1 axis plays an important role in the tumorigenesis of gastric cancer. - Abstract: The lncRNA H19 has been recently shown to be upregulated and play important roles in gastric cancer tumorigenesis. However, the precise molecular mechanism of H19 and its mature product miR-675 in the carcinogenesis of gastric cancer remains unclear. In this study, we found that miR-675 was positively expressed with H19 and was a pivotal mediator in H19-induced gastric cancer cell growth promotion. Subsequently, the tumor suppressor Runt Domain Transcription Factor1 (RUNX1) was confirmed to be a direct target of miR-675 using a luciferase reporter assay and Western blotting analyses. A series of rescue assays indicated that RUNX1 mediated H19/miR-67-induced gastric cancer cell phenotypic changes. Moreover, the inverse relationship between the expression of RUNX1 and H19/miR-675 was also revealed in gastric cancer tissues and gastric cancer cell lines. Taken together, our study demonstrated that the novel pathway H19/miR-675/RUNX1 regulates gastric cancer development and may serve as a potential target for gastric cancer therapy.

  2. Oncofetal H19 RNA promotes tumor metastasis.

    PubMed

    Matouk, Imad J; Raveh, Eli; Abu-lail, Rasha; Mezan, Shaul; Gilon, Michal; Gershtain, Eitan; Birman, Tatiana; Gallula, Jennifer; Schneider, Tamar; Barkali, Moshe; Richler, Carmelit; Fellig, Yakov; Sorin, Vladimir; Hubert, Ayala; Hochberg, Abraham; Czerniak, Abraham

    2014-07-01

    The oncofetal H19 gene transcribes a long non-coding RNA(lncRNA) that is essential for tumor growth. Here we found that numerous established inducers of epithelial to mesenchymal transition(EMT) also induced H19/miR-675 expression. Both TGF-β and hypoxia concomitantly induced H19 and miR-675 with the induction of EMT markers. We identified the PI3K/AKT pathway mediating the inductions of Slug, H19 RNA and miR-675 in response to TGF-β treatment, while Slug induction depended on H19 RNA. In the EMT induced multidrug resistance model, H19 level was also induced. In a mouse breast cancer model, H19 expression was tightly correlated with metastatic potential. In patients, we detected high H19 expression in all common metastatic sites tested, regardless of tumor primary origin. H19 RNA suppressed the expression of E-cadherin protein. H19 up-regulated Slug expression concomitant with the suppression of E-cadherin protein through a mechanism that involved miR-675. Slug also up-regulated H19 expression and activated its promoter. Altogether, these results may support the existence of a positive feedback loop between Slug and H19/miR-675, that regulates E-cadherin expression. H19 RNA enhanced the invasive potential of cancer cells in vitro and enhanced tumor metastasis in vivo. Additionally, H19 knockdown attenuated the scattering and tumorigenic effects of HGF/SF. Our results present novel mechanistic insights into a critical role for H19 RNA in tumor progression and indicate a previously unknown link between H19/miR-675, Slug and E-cadherin in the regulation of cancer cell EMT programs.

  3. Long noncoding RNA H19 mediates melatonin inhibition of premature senescence of c-kit(+) cardiac progenitor cells by promoting miR-675.

    PubMed

    Cai, Benzhi; Ma, Wenya; Bi, Chongwei; Yang, Fan; Zhang, Lai; Han, Zhenbo; Huang, Qi; Ding, Fengzhi; Li, Yuan; Yan, Gege; Pan, Zhenwei; Yang, Baofeng; Lu, Yanjie

    2016-08-01

    Melatonin, a hormone secreted by the pineal gland, possesses multiple biological activities such as antitumor, antioxidant, and anti-ischemia. C-kit(+) cardiac progenitor cells (CPCs) have emerged as a promising tool for the treatment of heart diseases. However, the senescence of CPCs due to pathological stimuli leads to the decline of CPCs' functions and regenerative potential. This study was conducted to demonstrate whether melatonin antagonizes the senescence of CPCs in response to oxidative stress. Here, we found that the melatonin treatment markedly inhibited the senescent characteristics of CPCs after exposed to sublethal concentration of H2 O2 , including the increase in senescence-associated β-galactosidase (SA-β-gal)-positive CPCs, senescence-associated heterochromatin loci (SAHF), secretory IL-6 level, and the upregulation of p53 and p21 proteins. Senescence-associated proliferation reduction was also attenuated by melatonin in CPCs. Luzindole, the melatonin membrane receptor blocker, may block the melatonin-mediated suppression of premature senescence in CPCs. Interestingly, we found that long noncoding RNA H19 and its derived miR-675 were downregulated by H2 O2 in CPCs, but melatonin treatment could counter this alteration. Furthermore, knockdown of H19 or miR-675 blocked antisenescence actions of melatonin on H2 O2 -treated CPCs. It was further verified that H19-derived miR-675 targeted at the 3'UTR of USP10, which resulted in the downregulation of p53 and p21 proteins. In summary, melatonin antagonized premature senescence of CPCs via H19/miR-675/USP10 pathway, which provides new insights into pharmacological actions and potential applications of melatonin on the senescence of CPCs.

  4. H19 non coding RNA-derived miR-675 enhances tumorigenesis and metastasis of breast cancer cells by downregulating c-Cbl and Cbl-b

    PubMed Central

    Vennin, Constance; Spruyt, Nathalie; Dahmani, Fatima; Julien, Sylvain; Bertucci, François; Finetti, Pascal; Chassat, Thierry; Bourette, Roland P.; Le Bourhis, Xuefen; Adriaenssens, Eric

    2015-01-01

    H19 is a long non-coding RNA precursor of miR-675microRNA. H19 is increasingly described to play key roles in the progression and metastasis of cancers from different tissue origins. We have previously shown that the H19 gene is activated by growth factors and increases breast cancer cell invasion. In this study, we established H19/miR-675 ectopic expression models of MDA-MB-231 breast cancer cells to further investigate the underlying mechanisms of H19 oncogenic action. We showed that overexpression of H19/miR-675 enhanced the aggressive phenotype of breast cancer cells including increased cell proliferation and migration in vitro, and increased tumor growth and metastasis in vivo. Moreover, we identified ubiquitin ligase E3 family (c-Cbl and Cbl-b) as direct targets of miR-675 in breast cancer cells. Using a luciferase assay, we demonstrated that H19, through its microRNA, decreased both c-Cbl and Cbl-b expression in all breast cancer cell lines tested. Thus, by directly binding c-Cbl and Cbl-b mRNA, miR-675 increased the stability and the activation of EGFR and c-Met, leading to sustained activation of Akt and Erk as well as enhanced cell proliferation and migration. Our data describe a novel mechanism of protumoral action of H19 in breast cancer. PMID:26353930

  5. H19 non coding RNA-derived miR-675 enhances tumorigenesis and metastasis of breast cancer cells by downregulating c-Cbl and Cbl-b.

    PubMed

    Vennin, Constance; Spruyt, Nathalie; Dahmani, Fatima; Julien, Sylvain; Bertucci, François; Finetti, Pascal; Chassat, Thierry; Bourette, Roland P; Le Bourhis, Xuefen; Adriaenssens, Eric

    2015-10-06

    H19 is a long non-coding RNA precursor of miR-675 microRNA. H19 is increasingly described to play key roles in the progression and metastasis of cancers from different tissue origins. We have previously shown that the H19 gene is activated by growth factors and increases breast cancer cell invasion. In this study, we established H19/miR-675 ectopic expression models of MDA-MB-231 breast cancer cells to further investigate the underlying mechanisms of H19 oncogenic action. We showed that overexpression of H19/miR-675 enhanced the aggressive phenotype of breast cancer cells including increased cell proliferation and migration in vitro, and increased tumor growth and metastasis in vivo. Moreover, we identified ubiquitin ligase E3 family (c-Cbl and Cbl-b) as direct targets of miR-675 in breast cancer cells. Using a luciferase assay, we demonstrated that H19, through its microRNA, decreased both c-Cbl and Cbl-b expression in all breast cancer cell lines tested. Thus, by directly binding c-Cbl and Cbl-b mRNA, miR-675 increased the stability and the activation of EGFR and c-Met, leading to sustained activation of Akt and Erk as well as enhanced cell proliferation and migration. Our data describe a novel mechanism of protumoral action of H19 in breast cancer.

  6. MiR675-5p Acts on HIF-1α to Sustain Hypoxic Responses: A New Therapeutic Strategy for Glioma

    PubMed Central

    Lo Dico, Alessia; Costa, Viviana; Martelli, Cristina; Diceglie, Cecilia; Rajata, Francesca; Rizzo, Aroldo; Mancone, Carmine; Tripodi, Marco; Ottobrini, Luisa; Alessandro, Riccardo; Conigliaro, Alice

    2016-01-01

    Hypoxia is a common feature in solid tumours. In glioma, it is considered the major driving force for tumour angiogenesis and correlates with enhanced resistance to conventional therapies, increased invasiveness and a poor prognosis for patients. Here we describe, for the first time, that miR675-5p, embedded in hypoxia-induced long non-coding RNA H19, plays a mandatory role in establishing a hypoxic response and in promoting hypoxia-mediated angiogenesis. We demonstrated, in vitro and in vivo, that miR675-5p over expression in normoxia is sufficient to induce a hypoxic moreover, miR675-5p depletion in low oxygen conditions, drastically abolishes hypoxic responses including angiogenesis. In addition, our data indicate an interaction of miR675-5p, HIF-1α mRNA and the RNA Binding Protein HuR in hypoxia-induced responses. We suggest the modulation of miR675-5p as a new therapeutic option to promote or abolish hypoxia induced angiogenesis. PMID:27279905

  7. Increased expression of H19/miR-675 is associated with a low fat-free mass index in patients with COPD.

    PubMed

    Lewis, Amy; Lee, Jen Y; Donaldson, Anna V; Natanek, S Amanda; Vaidyanathan, Srividya; Man, William D-C; Hopkinson, Nicholas S; Sayer, Avan A; Patel, Harnish P; Cooper, Cyrus; Syddall, Holly; Polkey, Michael I; Kemp, Paul R

    2016-06-01

    Loss of muscle mass and strength is a significant comorbidity in patients with chronic obstructive pulmonary disease (COPD) that limits their quality of life and has prognostic implications but does not affect everyone equally. To identify mechanisms that may contribute to the susceptibility to a low muscle mass, we investigated microRNA (miRNA) expression, methylation status, and regeneration in quadriceps muscle from COPD patients and the effect of miRNAs on myoblast proliferation in vitro. The relationships of miRNA expression with muscle mass and strength was also determined in a group of healthy older men. We identified miRNAs associated with a low fat-free mass (FFM) phenotype in a small group of patients with COPD using a PCR screen of 750 miRNAs. The expression of two differentially expressed miRNAs (miR-675 and miR-519a) was determined in an expanded group of COPD patients and their associations with FFM and strength identified. The association of these miRNAs with FFM and strength was also explored in a group of healthy community-dwelling older men. As the expression of the miRNAs associated with FFM could be regulated by methylation, the relative methylation of the H19 ICR was determined. Furthermore, the proportion of myofibres with centralized nuclei, as a marker of muscle regeneration, in the muscle of COPD patients was identified by immunofluorescence. Imprinted miRNAs (miR-675 and from a cluster, C19MC which includes miR-519a) were differentially expressed in the quadriceps of patients with a low fat-free mass index (FFMI) compared to those with a normal FFMI. In larger cohorts, miR-675 and its host gene (H19) were higher in patients with a low FFMI and strength. The association of miR-519a expression with FFMI was present in male patients with severe COPD. Similar associations of miR expression with lean mass and strength were not observed in healthy community dwelling older men participating in the Hertfordshire Sarcopenia Study. Relative methylation of the H19 ICR was reduced in COPD patients with muscle weakness but was not associated with FFM. In vitro, miR-675 inhibited myoblast proliferation and patients with a low FFMI had fewer centralized nuclei suggesting miR-675 represses regeneration. The data suggest that increased expression of miR-675/H19 and altered methylation of the H19 imprinting control region are associated with a low FFMI in patients with COPD but not in healthy community dwelling older men suggesting that epigenetic control of this loci may contribute to a susceptibility to a low FFMI.

  8. MiR-9-5p, miR-675-5p and miR-138-5p Damages the Strontium and LRP5-Mediated Skeletal Cell Proliferation, Differentiation, and Adhesion.

    PubMed

    Sun, Tianhao; Leung, Frankie; Lu, William W

    2016-02-15

    This study was designed to evaluate the effects of strontium on the expression levels of microRNAs (miRNAs) and to explore their effects on skeletal cell proliferation, differentiation, adhesion, and apoptosis. The targets of these miRNAs were also studied. Molecular cloning, cell proliferation assay, cell apoptosis assay, quantitative real-time PCR, and luciferase reporter assay were used. Strontium altered the expression levels of miRNAs in vitro and in vivo. miR-9-5p, miR-675-5p, and miR-138-5p impaired skeletal cell proliferation, cell differentiation and cell adhesion. miR-9-5p and miR-675-5p induced MC3T3-E1 cell apoptosis more specifically than miR-138-5p. miR-9-5p, miR-675-5p, and miR-138-5p targeted glycogen synthase kinase 3 β (GSK3β), ATPase Aminophospholipid Transporter Class I Type 8A Member 2 (ATP8A2), and Eukaryotic Translation Initiation Factor 4E Binding Protein 1 (EIF4EBP1), respectively. Low-density lipoprotein receptor-related protein 5 (LRP5) played a positive role in skeletal development. miR-9-5p, miR-675-5p, and miR-138-5p damage strontium and LRP5-mediated skeletal cell proliferation, differentiation, and adhesion, and induce cell apoptosis by targeting GSK3β, ATP8A2, and EIF4EBP1, respectively.

  9. The role of the oncofetal H19 lncRNA in tumor metastasis: orchestrating the EMT-MET decision

    PubMed Central

    Matouk, Imad J.; Halle, David; Raveh, Eli; Gilon, Michal; Sorin, Vladimir; Hochberg, Avraham

    2016-01-01

    Long non-coding RNA (lncRNA) genes are emerging as key players in the metastatic cascade. Current evidence indicate that H19 lncRNA and the microRNA(miRNA) miR-675, which is processed from it, play crucial roles in metastasis, through the regulation of critical events specifically the epithelial to mesenchymal (EMT) and the mesenchymal to epithelial transitions (MET). This review summarizes recent mechanistic pathways and tries to put together seemingly conflicting data from different reports under one proposed general scheme underlying the various roles of H19/miR-675 in the metastatic cascade. We propose several approaches to harnessing this knowledge for translational medicine. PMID:26623562

  10. Proteomics Identification of Desmin as a Potential Oncofetal Diagnostic and Prognostic Biomarker in Colorectal Cancer*

    PubMed Central

    Ma, Yanlei; Peng, Jiayuan; Liu, Weijie; Zhang, Peng; Huang, Long; Gao, Benbo; Shen, Tongyi; Zhou, Yukun; Chen, Hongqi; Chu, Zhaoxin; Zhang, Ming; Qin, Huanlong

    2009-01-01

    Colorectal cancer (CRC) is the third most common cancer worldwide and has poor prognosis. To identify the oncofetal proteins involved in CRC carcinogenesis, differentially expressed proteins among fetal colorectal tissues, CRC, and the paired tumor-adjacent normal colorectal tissues were investigated by a two-dimensional gel electrophoresis and MALDI-TOF/TOF-based proteomics approach. 42 protein spots were differentially expressed among these tissues, and 22 proteins were identified by MS analysis. Desmin and zinc finger protein 829 were found to be elevated in CRC tissue and fetal colorectal tissue compared with normal colorectal tissue. The elevated expression of desmin in CRC tissue and different developmental stages of fetus colon was confirmed by RT-PCR and Western blot analysis. Immunohistochemical analysis showed that the elevated expression of desmin was correlated with the severity and differentiation of CRC and decreased survival rate of CRC patients. Finally by developing a highly sensitive immunoassay, desmin could be detected in human serum and was significantly elevated in CRC patients compared with healthy volunteers. We propose that desmin be considered a potential oncofetal serum tumor marker for CRC that may have significance in the detection of patients with CRC. PMID:19460759

  11. Induction of epithelial-mesenchymal transition with O-glycosylated oncofetal fibronectin

    PubMed Central

    Ding, Yao; Gelfenbeyn, Kirill; Freire-de-Lima, Leonardo; Handa, Kazuko; Hakomori, Sen-itiroh

    2012-01-01

    Epithelial-mesenchymal transition (EMT) has been shown to play a key role in embryogenesis and cancer progression. We previously found that fibronectin (FN) carrying O-GalNAc at a specific site is selectively expressed in cancer and fetal cells/tissues, and termed oncofetal FN (onfFN). Here, we show that (i) a newly-established monoclonal antibody against FN lacking the O-GalNAc, termed normalFN (norFN), is useful for isolation of onfFN, (ii) onfFN, but not norFN, can induce EMT in human lung carcinoma cells, (iii) onfFN has a synergistic effect with transforming growth factor (TGF)β1 in EMT induction. PMID:22641031

  12. Oncofetal Chondroitin Sulfate Glycosaminoglycans are Key Players in Integrin Signaling and Tumor Cell Motility

    PubMed Central

    Clausen, Thomas Mandel; Pereira, Marina Ayres; Al Nakouzi, Nader; Oo, Htoo Zarni; Agerbæk, Mette Ø; Lee, Sherry; Ørum-Madsen, Maj Sofie; Christensen, Anders Riis; El-Naggar, Amal; Grandgenett, Paul M.; Grem, Jean L.; Hollingsworth, Michael A.; Holst, Peter J.; Theander, Thor; Sorensen, Poul H.; Daugaard, Mads; Salanti, Ali

    2016-01-01

    Many tumors express proteoglycans modified with oncofetal chondroitin sulfate glycosaminoglycan chains (ofCS), which are normally restricted to the placenta. However, the role of ofCS in cancer is largely unknown. The function of ofCS in cancer was analyzed using the recombinant ofCS-binding VAR2CSA protein (rVAR2) derived from the malaria parasite, Plasmodium falciparum. We demonstrate that ofCS plays a key role in tumor cell motility by affecting canonical integrin signaling pathways. Binding of rVAR2 to tumor cells inhibited the interaction of cells with extracellular matrix (ECM) components, which correlated with decreased phosphorylation of Src kinase. Moreover, rVAR2 binding decreased migration, invasion and anchorage-independent growth of tumor cells in vitro. Mass spectrometry of ofCS-modified proteoglycan complexes affinity purified from tumor cell lines on rVAR2 columns, revealed an overrepresentation of proteins involved in cell motility and integrin signaling, such as integrin β1 (ITGB1) and integrin α4 (ITGA4). Saturating concentrations of rVAR2 inhibited downstream integrin signaling, which was mimicked by knockdown of the core CS synthesis enzymes Beta-1,3-Glucuronyltransferase 1 (B3GAT1) and Chondroitin Sulfate N-Acetylgalactosaminyltransferase 1 (CSGALNACT1). The ofCS modification was highly expressed in both human and murine metastatic lesions in situ and pre-incubation or early intravenous treatment of tumor cells with rVAR2 inhibited seeding and spreading of tumor cells in mice. This was associated with a significant increase in survival of the animals. These data functionally link ofCS modifications with cancer cell motility and further highlights ofCS as a novel therapeutic cancer target. Implications The cancer specific expression of oncofetal chondroitin sulfate aids in metastatic phenotypes and is a candidate target for therapy. PMID:27655130

  13. Use of the 60 kd oncofetal protein for monitoring chemical hepatocarcinogenesis

    SciTech Connect

    Hanausek-Walaszek, M.; Walaszek, Z.; Webb, T.E.

    1986-03-01

    The 60 kd oncofetal protein (OFP-60) was earlier found in the blood plasma of tumor hosts and carcinogen-treated animals. OFP-60 was shown to be induced by all initiators of chemical carcinogenesis tested but neither by tumor promoters nor by liver regeneration. In the present study a hepatocarcinogen-mediated induction of OFP-60 and its release to circulation has been monitored by use of a sensitive biochemical assay. This assay measures, in a cell-free system, the energy-dependent release of nuclear mRNA sequences, catalyzed by OFP-60. A non-necrogenic dose by diethylnitrosamine (DENA) was administered to female Sprague Dawley rats 24 hours after partial hepatectomy. Half of the rats received phenobarbital in the diet beginning 7 weeks post-carcinogen treatment. Upon treatment with DENA, the relative activity of OFP-60 in the liver cytosol gradually increases up to 6 weeks and then remains constant. In the blood, the OFP-60 activity increases only up to 3 weeks, then decreases to basal level before a slow ascent is observed. Phenobarbital increases the activity in both liver and blood. The increase may be due to expansion of the cell population in the OFP-60 containing foci, while the decrease in the blood to immune clearance.

  14. The receptor tyrosine kinase ROR1--an oncofetal antigen for targeted cancer therapy.

    PubMed

    Hojjat-Farsangi, Mohammad; Moshfegh, Ali; Daneshmanesh, Amir Hossein; Khan, Abdul Salam; Mikaelsson, Eva; Osterborg, Anders; Mellstedt, Håkan

    2014-12-01

    Targeted cancer therapies have emerged as new treatment options for various cancer types. Among targets, receptor tyrosine kinases (RTKs) are among the most promising. ROR1 is a transmembrane RTK of importance during the normal embryogenesis for the central nervous system, heart, lung and skeletal systems, but is not expressed in normal adult tissues. However, ROR1 is overexpressed in several human malignancies and may act as a survival factor for tumor cells. Its unique expression by malignant cells may provide a target for novel therapeutics including monoclonal antibodies (mAbs) and small molecule inhibitors of tyrosine kinases (TKI) for the treatment of cancer. Promising preclinical results have been reported in e.g. chronic lymphocytic leukemia, pancreatic carcinoma, lung and breast cancer. ROR1 might also be an interesting oncofetal antigen for active immunotherapy. In this review, we provide an overview of the ROR1 structure and functions in cancer and highlight emerging therapeutic options of interest for targeting ROR1 in tumor therapy.

  15. The product of the imprinted H19 gene is an oncofetal RNA.

    PubMed Central

    Ariel, I.; Ayesh, S.; Perlman, E. J.; Pizov, G.; Tanos, V.; Schneider, T.; Erdmann, V. A.; Podeh, D.; Komitowski, D.; Quasem, A. S.; de Groot, N.; Hochberg, A.

    1997-01-01

    AIMS/BACKGROUND: The H19 gene is an imprinted, maternally expressed gene in humans. It is tightly linked and coregulated with the imprinted, paternally expressed gene of insulin-like growth factor 2. The H19 gene product is not translated into protein and functions as an RNA molecule. Although its role has been investigated for more than a decade, its biological function is still not understood fully. H19 is abundantly expressed in many tissues from early stages of embryogenesis through fetal life, and is down regulated postnatally. It is also expressed in certain childhood and adult tumours. This study was designed to screen the expression of H19 in human cancer and its relation to the expression of H19 in the fetus. METHODS: Using in situ hybridisation with a [35S] labelled probe, H19 mRNA was detected in paraffin wax sections of fetal tissues from the first and second trimesters of pregnancy and of a large array of human adult and childhood tumours arising from these tissues. RESULTS: The H19 gene is expressed in tumours arising from tissues which express this gene in fetal life. Its expression in the fetus and in cancer is closely linked with tissue differentiation. CONCLUSIONS: Based on these and previous data, H19 is neither a tumour suppressor gene nor an oncogene. Its product is an oncofetal RNA. The potential use of this RNA as a tumour marker should be evaluated. Images PMID:9208812

  16. Development and use of monoclonal antibodies against an oncofetal protein associated with carcinogenesis and tumorigenesis.

    PubMed

    Runge, S W; Larroya-Runge, S N; Schumm, D E; Webb, T E

    1991-06-01

    An oncofetal protein (OFP) studied in our laboratory associated with embryogenesis, carcinogenesis and tumorigenesis has as its known biological function the modification of RNA release from isolated nuclei. In the present study, we have developed and investigated the use of monoclonal antibodies against OFP. Six hybridoma cell lines (A-F) were isolated by screening the hybridoma culture media for anti-OFP antibodies (MOFP) with an indirect ELISA and by testing the ability of these antibodies complexed with anti-mouse IgG-agarose to bind to rat OFP and remove its associated RNA transport activity from solution (Immunobioassay). An inhibition ELISA developed to measure OFP gave a linear response up to 20 ng of plasma protein from a tumor-bearing rat. Western blot analysis using these monoclonals showed that OFP from a rat tumor (H7777) cytosol that shed to the blood consisted of two species exhibiting molecular weights of 50 and 55 kD respectively. In order to show the usefulness of our assays, a preliminary study showing the ability of the immunobioassay to detect the expression of OFP in the plasma of carcinogen treated rats in a dosage dependent manner has been presented. Since OFP is produced in the target organ of rats shortly after treatment with carcinogens and persists in the preneoplastic foci and subsequent tumors, these monoclonal antibodies will be valuable in studying its involvement in chemical carcinogenesis and tumorigenesis.

  17. Involvement of O-glycosylation defining oncofetal fibronectin in epithelial-mesenchymal transition process

    PubMed Central

    Freire-de-Lima, Leonardo; Gelfenbeyn, Kirill; Ding, Yao; Mandel, Ulla; Clausen, Henrik; Handa, Kazuko; Hakomori, Sen-itiroh

    2011-01-01

    The process termed “epithelial–mesenchymal transition” (EMT) was originally discovered in ontogenic development, and has been shown to be one of the key steps in tumor cell progression and metastasis. Recently, we showed that the expression of some glycosphingolipids (GSLs) is down-regulated during EMT in human and mouse cell lines. Here, we demonstrate the involvement of GalNAc-type (or mucin-type) O-glycosylation in EMT process, induced with transforming growth factor β (TGF-β) in human prostate epithelial cell lines. We found that: (i) TGF-β treatment caused up-regulation of oncofetal fibronectin (onfFN), which is defined by mAb FDC6, and expressed in cancer or fetal cells/tissues, but not in normal adult cells/tissues. The reactivity of mAb FDC6 requires the addition of an O-glycan at a specific threonine, inside the type III homology connective segment (IIICS) domain of FN. (ii) This change is associated with typical EMT characteristics; i.e., change from epithelial to fibroblastic morphology, enhanced cell motility, decreased expression of a typical epithelial cell marker, E-cadherin, and enhanced expression of mesenchymal markers. (iii) TGF-β treatment up-regulated mRNA level of FN containing the IIICS domain and GalNAc-T activity for the IIICS domain peptide substrate containing the FDC6 onfFN epitope. (iv) Knockdown of GalNAc-T6 and T3 inhibited TGF-β–induced up-regulation of onfFN and EMT process. (v) Involvement of GSLs was not detectable with the EMT process in these cell lines. These findings indicate the important functional role of expression of onfFN, defined by site-specific O-glycosylation at IIICS domain, in the EMT process. PMID:22006308

  18. Delineation of a cellular hierarchy in lung cancer reveals an oncofetal antigen expressed on tumor-initiating cells.

    PubMed

    Damelin, Marc; Geles, Kenneth G; Follettie, Maximillian T; Yuan, Ping; Baxter, Michelle; Golas, Jonathon; DiJoseph, John F; Karnoub, Maha; Huang, Shuguang; Diesl, Veronica; Behrens, Carmen; Choe, Sung E; Rios, Carol; Gruzas, Janet; Sridharan, Latha; Dougher, Maureen; Kunz, Arthur; Hamann, Philip R; Evans, Deborah; Armellino, Douglas; Khandke, Kiran; Marquette, Kimberly; Tchistiakova, Lioudmila; Boghaert, Erwin R; Abraham, Robert T; Wistuba, Ignacio I; Zhou, Bin-Bing S

    2011-06-15

    Poorly differentiated tumors in non-small cell lung cancer (NSCLC) have been associated with shorter patient survival and shorter time to recurrence following treatment. Here, we integrate multiple experimental models with clinicopathologic analysis of patient tumors to delineate a cellular hierarchy in NSCLC. We show that the oncofetal protein 5T4 is expressed on tumor-initiating cells and associated with worse clinical outcome in NSCLC. Coexpression of 5T4 and factors involved in the epithelial-to-mesenchymal transition were observed in undifferentiated but not in differentiated tumor cells. Despite heterogeneous expression of 5T4 in NSCLC patient-derived xenografts, treatment with an anti-5T4 antibody-drug conjugate resulted in complete and sustained tumor regression. Thus, the aggressive growth of heterogeneous solid tumors can be blocked by therapeutic agents that target a subpopulation of cells near the top of the cellular hierarchy.

  19. The oncofetal protein IMP3: a novel biomarker for endometrial serous carcinoma.

    PubMed

    Zheng, Wenxin; Yi, Xiaofang; Fadare, Oluwole; Liang, Sharon X; Martel, Maritza; Schwartz, Peter E; Jiang, Zhong

    2008-02-01

    Insulin-like growth factor II mRNA-binding protein 3 (IMP3) is an oncofetal protein highly expressed in fetal tissue and malignant tumors but rarely found in adult benign tissues. The aim of this study is to determine the expression of IMP3 in benign endometrium, endometrial cancer, and its precursor lesions, trying to see whether IMP3 has any diagnostic usage. Two hundred ninety-eight endometrial samples were examined for IMP3 expression by immunohistochemistry. These included benign endometrium (n=68), atypical hyperplasia or endometrial intraepithelial neoplasia (n=35), endometrial glandular dysplasia (n=21), endometrial intraepithelial carcinoma (n=18), endometrioid carcinoma (n=70), mucinous carcinoma (n=8), serous carcinoma (n=51), clear cell carcinoma (n=12), and other malignancies (n=15). Maturational patterns in the 68 benign endometrial samples included atrophic (n=12), proliferative (n=18), secretory (n=14), menstrual (n=8), and gestational (n=16). Most of the carcinomas were histologically pure; where mixed, the second component constituted <10% of the total tumor volume. The extent and intensity of IMP3 expression was semiquantitatively determined and scored for all samples. A renal cell carcinoma with known IMP3 expression was used as positive control for each immunohistochemistry run. Among the malignant cases, IMP3 expression was predominantly found in endometrial serous carcinoma and its putative precursor lesions, with 3 (14%) of 21 endometrial glandular dysplasia, 16 (89%) of 18 serous endometrial intraepithelial carcinoma, and 48 (94%) of 51 serous carcinomas (P<0.001). In contrast, the frequency of IMP3 expression was significantly lesser in nonserous malignancies with 0 (0%) of 35, 5 (7%) of 70, 0 (0%) of 8, 3 (25%) of 12, and 5 (33%) of 15 positive expression rates in atypical hyperplasia or endometrial intraepithelial neoplasia, endometrioid, mucinous, clear cell carcinomas, and other malignancies, respectively. The IMP3 staining was

  20. An oncofetal antigen, IMP-3-derived long peptides induce immune responses of both helper T cells and CTLs

    PubMed Central

    Hirayama, Masatoshi; Tomita, Yusuke; Yuno, Akira; Tsukamoto, Hirotake; Senju, Satoru; Imamura, Yuya; Sayem, Mohammad Abu; Irie, Atsushi; Yoshitake, Yoshihiro; Fukuma, Daiki; Shinohara, Masanori; Hamada, Akinobu; Jono, Hirofumi; Yuba, Eiji; Kono, Kenji; Yoshida, Koji; Tsunoda, Takuya; Nakayama, Hideki; Nishimura, Yasuharu

    2016-01-01

    ABSTRACT Insulin-like growth factor II mRNA-binding protein 3 (IMP-3), an oncofetal antigen identified using genome-wide cDNA microarray analyses, is overexpressed in several malignancies. IMP-3-derived cytotoxic T lymphocyte (CTL) epitopes have been used for peptide-based immunotherapies against various cancers. In addition to CTLs, induction of tumor-associated antigen (TAA)-specific helper T (Th) cells is crucial for establishment of effective antitumor immunity. In this study, we aimed to identify IMP-3-derived long peptides (IMP-3-LPs) carrying CTL and promiscuous Th-cell epitopes for use in cancer immunotherapy. IMP-3-derived Th-cell epitopes that bind to multiple HLA-class II molecules were predicted by in silico analysis, and their immunogenicity was determined by utilizing human T cells. We identified two highly immunogenic IMP-3-LPs presented by multiple HLA-class II molecules. One of the IMP-3-LPs encompassed two CTL epitopes that have been used for peptide-vaccine immunotherapy in ongoing clinical trials. IMP-3-LPs-specific Th cells responded to autologous dendritic cells (DCs) loaded with the recombinant IMP-3 proteins, suggesting that these s (LPs) can be naturally processed and presented. The IMP-3-LPs and specific Th cells augmented the expansion of IMP-3-specific CTLs, which was further enhanced by programmed cell death-1 (PD-1) blockade. In addition, IMP-3-LP encapsulated in liposomes was efficiently cross-presented in vitro, and this LP successfully cross-primed CTLs in HLA-A2 transgenic mice (Tgm) in vivo. Furthermore, one of the IMP-3-LPs induced IMP-3-specific Th cells from peripheral blood mononuclear cells (PBMCs) of head-and-neck malignant tumor (HNMT) patients. These findings suggest the potential usefulness of IMP-3-LPs in propagating both Th cells and CTLs and may have implications for IMP-3-LPs-based cancer immunotherapy. PMID:27471607

  1. An oncofetal antigen, IMP-3-derived long peptides induce immune responses of both helper T cells and CTLs.

    PubMed

    Hirayama, Masatoshi; Tomita, Yusuke; Yuno, Akira; Tsukamoto, Hirotake; Senju, Satoru; Imamura, Yuya; Sayem, Mohammad Abu; Irie, Atsushi; Yoshitake, Yoshihiro; Fukuma, Daiki; Shinohara, Masanori; Hamada, Akinobu; Jono, Hirofumi; Yuba, Eiji; Kono, Kenji; Yoshida, Koji; Tsunoda, Takuya; Nakayama, Hideki; Nishimura, Yasuharu

    2016-06-01

    Insulin-like growth factor II mRNA-binding protein 3 (IMP-3), an oncofetal antigen identified using genome-wide cDNA microarray analyses, is overexpressed in several malignancies. IMP-3-derived cytotoxic T lymphocyte (CTL) epitopes have been used for peptide-based immunotherapies against various cancers. In addition to CTLs, induction of tumor-associated antigen (TAA)-specific helper T (Th) cells is crucial for establishment of effective antitumor immunity. In this study, we aimed to identify IMP-3-derived long peptides (IMP-3-LPs) carrying CTL and promiscuous Th-cell epitopes for use in cancer immunotherapy. IMP-3-derived Th-cell epitopes that bind to multiple HLA-class II molecules were predicted by in silico analysis, and their immunogenicity was determined by utilizing human T cells. We identified two highly immunogenic IMP-3-LPs presented by multiple HLA-class II molecules. One of the IMP-3-LPs encompassed two CTL epitopes that have been used for peptide-vaccine immunotherapy in ongoing clinical trials. IMP-3-LPs-specific Th cells responded to autologous dendritic cells (DCs) loaded with the recombinant IMP-3 proteins, suggesting that these s (LPs) can be naturally processed and presented. The IMP-3-LPs and specific Th cells augmented the expansion of IMP-3-specific CTLs, which was further enhanced by programmed cell death-1 (PD-1) blockade. In addition, IMP-3-LP encapsulated in liposomes was efficiently cross-presented in vitro, and this LP successfully cross-primed CTLs in HLA-A2 transgenic mice (Tgm) in vivo. Furthermore, one of the IMP-3-LPs induced IMP-3-specific Th cells from peripheral blood mononuclear cells (PBMCs) of head-and-neck malignant tumor (HNMT) patients. These findings suggest the potential usefulness of IMP-3-LPs in propagating both Th cells and CTLs and may have implications for IMP-3-LPs-based cancer immunotherapy.

  2. Developmental transcription factor NFIB is a putative target of oncofetal miRNAs and is associated with tumour aggressiveness in lung adenocarcinoma.

    PubMed

    Becker-Santos, Daiana D; Thu, Kelsie L; English, John C; Pikor, Larissa A; Martinez, Victor D; Zhang, May; Vucic, Emily A; Luk, Margaret Ty; Carraro, Anita; Korbelik, Jagoda; Piga, Daniela; Lhomme, Nicolas M; Tsay, Mike J; Yee, John; MacAulay, Calum E; Lam, Stephen; Lockwood, William W; Robinson, Wendy P; Jurisica, Igor; Lam, Wan L

    2016-10-01

    Genes involved in fetal lung development are thought to play crucial roles in the malignant transformation of adult lung cells. Consequently, the study of lung tumour biology in the context of lung development has the potential to reveal key developmentally relevant genes that play critical roles in lung cancer initiation/progression. Here, we describe for the first time a comprehensive characterization of miRNA expression in human fetal lung tissue, with subsequent identification of 37 miRNAs in non-small cell lung cancer (NSCLC) that recapitulate their fetal expression patterns. Nuclear factor I/B (NFIB), a transcription factor essential for lung development, was identified as a potential frequent target for these 'oncofetal' miRNAs. Concordantly, analysis of NFIB expression in multiple NSCLC independent cohorts revealed its recurrent underexpression (in ∼40-70% of tumours). Interrogation of NFIB copy number, methylation, and mutation status revealed that DNA level disruption of this gene is rare, and further supports the notion that oncofetal miRNAs are likely the primary mechanism responsible for NFIB underexpression in NSCLC. Reflecting its functional role in regulating lung differentiation, low expression of NFIB was significantly associated with biologically more aggressive subtypes and, ultimately, poorer survival in lung adenocarcinoma patients. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  3. Repression by sustained-release. beta. -glucuronidase inhibitors of chemical carcinogen-mediated induction of a marker oncofetal protein in rodents

    SciTech Connect

    Walaszek, Z.; Hanausek-Walaszek, M.; Webb, T.E.

    1988-01-01

    The degree of induction of an oncofetal protein marker in rodents by selected chemical carcinogens has been correlated with changes in carcinogenicity induced by dietary D-glucaro-1,4-lactone (GL) based anticarcinogens. These potent anticarcinogens may act to increase the clearance of carcinogens as glucuronides through the inhibition of ..beta..-glucuronidase. The sustained-release forms are particularly effective, 1.5 mmol/kg of GL maintaining serum ..beta..-glucuronidase activity at or below 50% for only 1 h, while an equivalent amount of calcium glucarate (CGT) maintained this level of inhibition for over 5 h. CGT or other sustained-release inhibitors, when fed to rodents during administration of carcinogens that undergo glucuronidation, caused a marked reduction in the induction of the marker protein. For those systems where other markers of carcinogenesis were also assessed, it was determined the inhibition of marker-protein induction was quantitatively similar to both the inhibition of binding of the carcinogen to DNA and the subsequent induction of tumors in target organs. The following carcinogens were administered intraperitoneally: benzo(a)pryene; 7,12-demethylbenz(a)anthracene; 3-methylcholanthrene; 2-acetylaminofluorene; 2-naphthylamine; N-nitroso-N,N-dibutylamine; aflatoxin B1; 1-nitropyrene.

  4. Long-term tumor regression induced by an antibody-drug conjugate that targets 5T4, an oncofetal antigen expressed on tumor-initiating cells.

    PubMed

    Sapra, Puja; Damelin, Marc; Dijoseph, John; Marquette, Kimberly; Geles, Kenneth G; Golas, Jonathon; Dougher, Maureen; Narayanan, Bitha; Giannakou, Andreas; Khandke, Kiran; Dushin, Russell; Ernstoff, Elana; Lucas, Judy; Leal, Mauricio; Hu, George; O'Donnell, Christopher J; Tchistiakova, Lioudmila; Abraham, Robert T; Gerber, Hans-Peter

    2013-01-01

    Antibody-drug conjugates (ADC) represent a promising therapeutic modality for the clinical management of cancer. We sought to develop a novel ADC that targets 5T4, an oncofetal antigen expressed on tumor-initiating cells (TIC), which comprise the most aggressive cell population in the tumor. We optimized an anti-5T4 ADC (A1mcMMAF) by sulfydryl-based conjugation of the humanized A1 antibody to the tubulin inhibitor monomethylauristatin F (MMAF) via a maleimidocaproyl linker. A1mcMMAF exhibited potent in vivo antitumor activity in a variety of tumor models and induced long-term regressions for up to 100 days after the last dose. Strikingly, animals showed pathologic complete response in each model with doses as low as 3 mg antibody/kg dosed every 4 days. In a non-small cell lung cancer patient-derived xenograft model, in which 5T4 is preferentially expressed on the less differentiated tumor cells, A1mcMMAF treatment resulted in sustained tumor regressions and reduced TIC frequency. These results highlight the potential of ADCs that target the most aggressive cell populations within tumors, such as TICs. In exploratory safety studies, A1mcMMAF exhibited no overt toxicities when administered to cynomolgus monkeys at doses up to 10 mg antibody/kg/cycle × 2 and displayed a half-life of 5 days. The preclinical efficacy and safety data established a promising therapeutic index that supports clinical testing of A1mcMMAF.

  5. Comparing prothrombin induced by vitamin K absence-II (PIVKA-II) with the oncofetal proteins glypican-3, Alpha feto protein and carcinoembryonic antigen in diagnosing hepatocellular carcinoma among Egyptian patients.

    PubMed

    Abd El Gawad, Iman A; Mossallam, Ghada I; Radwan, Noha H; Elzawahry, Heba M; Elhifnawy, Niveen M

    2014-06-01

    Hepatocellular carcinoma (HCC) is usually asymptomatic in the early stage and does not show elevated alpha-feto protein (AFP). AFP shows 60-80% sensitivity in diagnosing HCC. Glypican3 (GPC-3) is an oncofetal protein that is only detected in HCC cells but not in benign liver tissues, while Carcinoembryonic antigen (CEA) is expressed in various neoplasms including HCC. Although, it is not specific for HCC. Prothrombin induced by vitamin K absence-II (PIVKA-II) is an abnormal prothrombin protein that is increased in the serum of HCC patients. It has higher sensitivity and specificity compared to AFP. The aim of this study is to compare the clinical utility of PIVKA-II with GPC-3, AFP and CEA in diagnosing HCC. This study included 40 patients with HCC, 10 patients with cirrhosis as a benign control group, and 10 apparently healthy volunteers as normal controls. Serum samples were subjected to routine laboratory investigations, measurement of CEA, AFP using MEIA technique (Axsym), glypican3, and PIVKA-II using ELISA technique in the sera of all patients and controls. All markers showed the highest results in the HCC group. Higher concentrations of PIVKA-II were detected in patients with splenomegaly, and in tumors with size (>3cm). Combination of Glypican-3 and PIVKA-II showed the highest sensitivity, while GPC-3 alone and combination of GPC-3 and AFP showed the highest specificity to differentiate HCC from liver cirrhosis and normal controls. GPC-3, PIVKAII, and combination of both showed the highest sensitivity, while GPC-3 alone showed the highest specificity to differentiate HCC from liver cirrhosis. Glypican-3 is the only oncofetal antigen that showed comparable high diagnostic accuracy as PIVKA-II in diagnosing HCC among Egyptian patients. Copyright © 2014. Production and hosting by Elsevier B.V.

  6. Blood presence of circulating oncofetal fibronectin mRNA, by RT-PCR, does not represent a useful specific marker for the management and follow-up of thyroid cancer patients.

    PubMed

    Vendittelli, Francesca; Raffaelli, Marco; Fadda, Guido; Carelli-Alinovi, Cristiana; Paolillo, Carmela; Bellantone, Rocco; Zuppi, Cecilia; Capoluongo, Ettore

    2012-01-06

    Recent studies strongly suggest the use of oncofetal fibronectin (onfFN) mRNA in diagnostic follow-up and staging due to its very high specificity for thyroid cancers. Since the use of this marker has not been well established yet, particularly in the monitoring of minimal residual disease, we have tried to verify the diagnostic power of onfFN and its usefulness as a prognostic molecular marker. For this reason, we evaluated (by RT-PCR) the presence of onfFN mRNAs, not only in blood samples and thyroid tissues (both normal and neoplastic), but also in different biological fluids (such as K3-EDTA blood samples, saliva and urine) belonging to healthy individuals. Molecular investigations, such as RT-PCR protocol, and sequencing of onfFN cDNAs evaluation of the above-mentioned samples were performed. The onfFN transcript was largely expressed in all benign and malignant thyroid tissues [differentiated thyroid carcinomas (DTCs)] tested as well as in a large number of biological fluids; in particular, 100% urine samples were positive for onfFN transcript as compared to the thyroglobulin (Tg) mRNA (75%), while saliva was always positive for onfFN and never for Tg. These findings indicate that onfFN cannot be considered a marker specific for thyroid cancer presence. Finally, Tg results were positive in a large part of the samples, but not always in concomitance with onfFN. We underline how the complexity of onfFN transcripts could affect the RT-PCR procedure. In addition, the presence of onfFN transcripts in several normal and cancer tissues, along with non-thyroid biological fluids or cells, does not allow the use of this marker for cancer monitoring.

  7. Cross-Presentation of the Oncofetal Tumor Antigen 5T4 from Irradiated Prostate Cancer Cells--A Key Role for Heat-Shock Protein 70 and Receptor CD91.

    PubMed

    Salimu, Josephine; Spary, Lisa K; Al-Taei, Saly; Clayton, Aled; Mason, Malcolm D; Staffurth, John; Tabi, Zsuzsanna

    2015-06-01

    Immune responses contribute to the success of radiotherapy of solid tumors; however, the mechanism of triggering CD8(+) T-cell responses is poorly understood. Antigen cross-presentation from tumor cells by dendritic cells (DC) is a likely dominant mechanism to achieve CD8(+) T-cell stimulation. We established a cross-presentation model in which DCs present a naturally expressed oncofetal tumor antigen (5T4) from irradiated DU145 prostate cancer cells to 5T4-specific T cells. The aim was to establish which immunogenic signals are important in radiation-induced cross-presentation. Radiation (12 Gy) caused G2-M cell-cycle arrest and cell death, increased cellular 5T4 levels, high-mobility protein group-B1 (HMGB1) release, and surface calreticulin and heat-shock protein-70 (Hsp70) expression in DU145 cells. DCs phagocytosed irradiated tumor cells efficiently, followed by upregulation of CD86 on phagocytic DCs. CD8(+) 5T4-specific T cells, stimulated with these DCs, proliferated and produced IFNγ. Inhibition of HMGB1 or the TRIF/MyD88 pathway only had a partial effect on T-cell stimulation. Unlike previous investigators, we found no evidence that DCs carrying Asp299Gly Toll-like receptor-4 (TLR4) single-nucleotide polymorphism had impaired ability to cross-present tumor antigen. However, pretreatment of tumor cells with Hsp70 inhibitors resulted in a highly statistically significant and robust prevention of antigen cross-presentation and CD86 upregulation on DCs cocultured with irradiated tumor cells. Blocking the Hsp70 receptor CD91 also abolished cross-presentation. Together, the results from our study demonstrate that irradiation induces immunologically relevant changes in tumor cells, which can trigger CD8(+) T-cell responses via a predominantly Hsp70-dependent antigen cross-presentation process.

  8. UG311, An Oncofetal Marker Lost with Prostate Cancer Progression

    DTIC Science & Technology

    2001-04-01

    replacement therapy where no increased risk of prostate cancer was found [57]. That said, in acromegaly , a condition characterized by chronically...Carter, H.B. and Coffey, D.S. 1988, J. Urol., 140, 173. 134. Rajah, R., Nachajon, R.V., Collins, M.H., Hakonarson, H., et al. 1999, Am. J. Respir . Cell

  9. Urinary oncofetal ED-A fibronectin correlates with poor prognosis in patients with bladder cancer.

    PubMed

    Arnold, Shanna A; Loomans, Holli A; Ketova, Tatiana; Andl, Claudia D; Clark, Peter E; Zijlstra, Andries

    2016-01-01

    The extracellular matrix protein fibronectin (FN) contributes to the structural integrity of tissues as well as the adhesive and migratory functions of cells. While FN is abundantly expressed in adult tissues, the expression of several alternatively spliced FN isoforms is restricted to embryonic development, tissue remodeling and cancer. These FN isoforms, designated ED-A and ED-B, are frequently expressed by cancer cells, tumor-associated fibroblasts and newly forming blood vessels. Using a highly sensitive collagen-based indirect ELISA, we evaluated the correlation of urinary ED-A and ED-B at time of cystectomy with overall survival in patients with high-grade bladder cancer (BCa). Detectable levels of total FN as well as ED-A and ED-B were found in urine from 85, 73 and 51 % of BCa patients, respectively. The presence of urinary ED-A was a significant independent predictor of 2-year overall survival (OS) after adjusting for age, tumor stage, lymph node stage, and urinary creatinine by multivariable Logistic Regression (p = 0.029, OR = 4.26, 95 % CI 1.16-15.71) and improved accuracy by 3.6 %. Furthermore, detection of ED-A in the urine was a significant discriminator of survival specifically in BCa patients with negative lymph node status (Log-Rank, p = 0.006; HR = 5.78, 95 % CI 1.39-24.13). Lastly, multivariable Cox proportional hazards analysis revealed that urinary ED-A was an independent prognostic indicator of 5-year OS rate for patients with BCa (p = 0.04, HR = 2.20, 95 % CI 1.04-4.69). Together, these data suggest that cancer-derived, alternatively spliced FN isoforms can act as prognostic indicators and that additional studies are warranted to assess the clinical utility of ED-A in BCa.

  10. The Increasing Complexity of the Oncofetal H19 Gene Locus: Functional Dissection and Therapeutic Intervention

    PubMed Central

    Matouk, Imad; Raveh, Eli; Ohana, Patricia; Lail, Rasha Abu; Gershtain, Eitan; Gilon, Michal; De Groot, Nathan; Czerniak, Abraham; Hochberg, Abraham

    2013-01-01

    The field of the long non-coding RNA (lncRNA) is advancing rapidly. Currently, it is one of the most popular fields in the biological and medical sciences. It is becoming increasingly obvious that the majority of the human transcriptome has little or no-protein coding capacity. Historically, H19 was the first imprinted non-coding RNA (ncRNA) transcript identified, and the H19/IGF2 locus has served as a paradigm for the study of genomic imprinting since its discovery. In recent years, we have extensively investigated the expression of the H19 gene in a number of human cancers and explored the role of H19 RNA in tumor development. Here, we discuss recently published data from our group and others that provide further support for a central role of H19 RNA in the process of tumorigenesis. Furthermore, we focus on major transcriptional modulators of the H19 gene and discuss them in the context of the tumor-promoting activity of the H19 RNA. Based on the pivotal role of the H19 gene in human cancers, we have developed a DNA-based therapeutic approach for the treatment of cancers that have upregulated levels of H19 expression. This approach uses a diphtheria toxin A (DTA) protein expressed under the regulation of the H19 promoter to treat tumors with significant expression of H19 RNA. In this review, we discuss the treatment of four cancer indications in human subjects using this approach, which is currently under development. This represents perhaps one of the very few examples of an existing DNA-based therapy centered on an lncRNA system. Apart from cancer, H19 expression has been reported also in other conditions, syndromes and diseases, where deregulated imprinting at the H19 locus was obvious in some cases and will be summarized below. Moreover, the H19 locus proved to be much more complicated than initially thought. It houses a genomic sequence that can transcribe, yielding various transcriptional outputs, both in sense and antisense directions. The major transcriptional outputs of the H19 locus are presented here. PMID:23429271

  11. HCC-DETECT: a combination of nuclear, cytoplasmic, and oncofetal proteins as biomarkers for hepatocellular carcinoma.

    PubMed

    Attallah, Abdelfattah M; El-Far, Mohamed; Malak, Camelia A Abdel; Omran, Mohamed M; Shiha, Gamal E; Farid, Khaled; Barakat, Lamiaa A; Albannan, Mohamed S; Attallah, Ahmed A; Abdelrazek, Mohamed A; Elbendary, Mohamed S; Sabry, Refaat; Hamoda, Gehan A; Elshemy, Mohamed M; Ragab, Abdallah A; Foda, Basma M; Abdallah, Sanaa O

    2015-09-01

    Currently, the search for suitable hepatocellular carcinoma (HCC) biomarkers is very intensive. Besides, efficacy and cost/effectiveness of screening and surveillance of cirrhotics for the diagnosis of HCC is still debated. So, the present study is concerned with the evaluation of cytokeratin-1 (CK-1) and nuclear matrix protein-52 (NMP-52) for identifying HCC. Two-hundred and eighty individuals categorized into three groups [liver fibrosis (F1-F3), cirrhosis (F4), and HCC] constituted this study. Western blot was used for identifying CK-1 and NMP-52 in serum samples. As a result, a single immunoreactive band was shown at 67 and 52 kDa corresponding to CK-1 and NMP-52, respectively. Both CK-1 and NMP-52 bands were cut and electroeluted separately. These markers were quantified in sera using ELISA. Patients with HCC were associated with higher concentrations of CK-1 and NMP-52 than those without HCC with a significant difference (P < 0.0001). CK-1 showed an area under receiver-operating characteristic curve (AUC) of 0.83 with 75 % sensitivity and 82 % specificity while NMP-52 yielded 0.72 AUC with 62 % sensitivity and 70 % specificity for identifying HCC. HCC-DETECT comprising CK-1 and NMP-52 together with AFP was then constructed yielding 0.90 AUC for identifying HCC with 80 % sensitivity and 92 % specificity. HCC-DETECT was then tested for separating HCC from F1-F3 showing 0.94 AUC with 80 % sensitivity and 93 % specificity. In conclusion, CK-1 in conjunction with NMP-52 and AFP could have a potential role for improving the detection of HCC with a high degree of accuracy.

  12. Oncofetal protein, IMP-3, a potential marker for prediction of postoperative peritoneal dissemination in gastric adenocarcinoma.

    PubMed

    Okada, Kaoru; Fujiwara, Yoshiyuki; Nakamura, Yurika; Takiguchi, Shuji; Nakajima, Kiyokazu; Miyata, Hiroshi; Yamasaki, Makoto; Kurokawa, Yukinori; Takahashi, Tsuyoshi; Mori, Masaki; Doki, Yuichiro

    2012-06-15

    The aim of this study was to determine the expression of insulin-like growth factor-II messenger RNA (mRNA)-binding protein-3 (IMP-3) and its clinical significance in gastric cancers, as well the prognostic value of its expression in the peritoneal lavage fluid after surgery. IMP-3 expression was examined by immunohistochemistry in 96 primary gastric tumors. IMP-3 mRNA expression in peritoneal lavage fluid obtained at laparotomy was determine by real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). Positive staining for IMP-3 was observed in 74% (71/96) of the tumors. IMP-3 expression in gastric tumors correlated significantly with worst overall survival (OS) and recurrence-free survival. Multivariate analyses identified pathological N stage and IMP-3 expression as significant independent prognostic factors for disease-free survival. Eight (28%) of 36 peritoneal lavage samples were cytologically negative but positive for IMP-3 mRNA expression by RT-PCR. The OS of patients with IMP-3-positive peritoneal lavage was significantly worse than of those with negative expression. IMP-3 expression in primary gastric tumors was an independent poor prognostic factor. IMP-3 mRNA expression in peritoneal lavage fluid was a predictor of recurrence after surgery in gastric cancer and a marker of poor prognosis. Copyright © 2011 Wiley Periodicals, Inc.

  13. Extra-domain B in Oncofetal Fibronectin Structurally Promotes Fibrillar Head-to-tail Dimerization of Extracellular Matrix Protein*

    PubMed Central

    Schiefner, André; Gebauer, Michaela; Skerra, Arne

    2012-01-01

    The type III extra-domain B (ED-B) is specifically spliced into fibronectin (Fn) during embryogenesis and neoangiogenesis, including many cancers. The x-ray structure of the recombinant four-domain fragment FnIII7B89 reveals a tightly associated, extended head-to-tail dimer, which is stabilized via pair-wise shape and charge complementarity. A tendency toward ED-B-dependent dimer formation in solution was supported by size exclusion chromatography and analytical ultracentrifugation. When amending the model with the known three-dimensional structure of the FnIII10 domain, its RGD loop as well as the adhesion synergy region in FnIII9–10 become displayed on the same face of the dimer; this should allow simultaneous binding of at least two integrins and, thus, receptor clustering on the cell surface and intracellular signaling. Insertion of ED-B appears to stabilize overall head-to-tail dimerization of two separate Fn chains, which, together with alternating homodimer formation via disulfide bridges at the C-terminal Fn tail, should lead to the known macromolecular fibril formation. PMID:22442152

  14. H19 Long Noncoding RNA Regulates Intestinal Epithelial Barrier Function via MicroRNA 675 by Interacting with RNA-Binding Protein HuR

    PubMed Central

    Zou, Tongtong; Jaladanki, Suraj K.; Liu, Lan; Xiao, Lan; Chung, Hee Kyoung; Wang, Jun-Yao; Xu, Yan; Gorospe, Myriam

    2016-01-01

    The disruption of the intestinal epithelial barrier function occurs commonly in various pathologies, but the exact mechanisms responsible are unclear. The H19 long noncoding RNA (lncRNA) regulates the expression of different genes and has been implicated in human genetic disorders and cancer. Here, we report that H19 plays an important role in controlling the intestinal epithelial barrier function by serving as a precursor for microRNA 675 (miR-675). H19 overexpression increased the cellular abundance of miR-675, which in turn destabilized and repressed the translation of mRNAs encoding tight junction protein ZO-1 and adherens junction E-cadherin, resulting in the dysfunction of the epithelial barrier. Increasing the level of the RNA-binding protein HuR in cells overexpressing H19 prevented the stimulation of miR-675 processing from H19, promoted ZO-1 and E-cadherin expression, and restored the epithelial barrier function to a nearly normal level. In contrast, the targeted deletion of HuR in intestinal epithelial cells enhanced miR-675 production in the mucosa and delayed the recovery of the gut barrier function after exposure to mesenteric ischemia/reperfusion. These results indicate that H19 interacts with HuR and regulates the intestinal epithelial barrier function via the H19-encoded miR-675 by altering ZO-1 and E-cadherin expression posttranscriptionally. PMID:26884465

  15. Serum oncofetal fibronectin (onfFN) mRNA in differentiated thyroid carcinoma (DTC): large overlap between disease-free and metastatic patients.

    PubMed

    Sritara, Chanika; Charoenphun, Putthiporn; Ponglikitmongkol, Mathurose; Musikarat, Suchawadee; Utamakul, Chirawat; Chokesuwattanasakul, Payap; Thakkinstian, Ammarin

    2012-01-01

    This study assessed if onfFN mRNA in the peripheral blood of patients with DTC can identify individuals with metastatic disease. Comparison of onfFN mRNA was made among 3 groups: disease-free, lymph node metastasis, and distant metastasis using real-time RT-PCR on 5 ml blood samples from each DTC patient. Fifty-one patients were included: 30 (59%) were disease-free; 7 (13.7%) had lymph node metastasis; and 14 (27.5%) had distant metastasis. OnfFN mRNA levels in the 3 groups were significantly different (P=0.001) but with a large overlap and the expression being highest in the disease-free group. Subgroup analysis of the metastatic groups did not show any effect of age, cell type, and serum TSH, Tg, and antiTg on onfFN mRNA. The within-run and between-run root mean square coefficients of variations were <2%. OnfFN mRNA in patients with DTC cannot identify those with metastatic disease.

  16. Parental imprinting regulates insulin-like growth factor signaling: a Rosetta Stone for understanding the biology of pluripotent stem cells, aging and cancerogenesis.

    PubMed

    Ratajczak, M Z; Shin, D-M; Schneider, G; Ratajczak, J; Kucia, M

    2013-04-01

    In recent years, solid evidence has accumulated that insulin-like growth factor-1 (IGF-1) and 2 (IGF-2) regulate many biological processes in normal and malignant cells. Recently, more light has been shed on the epigenetic mechanisms regulating expression of genes involved in IGF signaling (IFS) and it has become evident that these mechanisms are crucial for initiation of embryogenesis, maintaining the quiescence of pluripotent stem cells deposited in adult tissues (for example, very-small embryonic-like stem cells), the aging process, and the malignant transformation of cells. The expression of several genes involved in IFS is regulated at the epigenetic level by imprinting/methylation within differentially methylated regions (DMRs), which regulate their expression from paternal or maternal chromosomes. The most important role in the regulation of IFS gene expression is played by the Igf-2-H19 locus, which encodes the autocrine/paracrine mitogen IGF-2 and the H19 gene, which gives rise to a non-coding RNA precursor of several microRNAs that negatively affect cell proliferation. Among these, miR-675 has recently been demonstrated to downregulate expression of the IGF-1 receptor. The proper imprinting of DMRs at the Igf-2-H19 locus, with methylation of the paternal chromosome and a lack of methylation on the maternal chromosome, regulates expression of these genes so that Igf-2 is transcribed only from the paternal chromosome and H19 (including miR-675) only from the maternal chromosome. In this review, we will discuss the relevance of (i) proper somatic imprinting, (ii) erasure of imprinting and (iii) loss of imprinting within the DMRs at the Igf-2-H19 locus to the expression of genes involved in IFS, and the consequences of these alternative patterns of imprinting for stem cell biology.

  17. SIX1 Oncoprotein as a Biomarker in a Model of Hormonal Carcinogenesis and in Human Endometrial Cancer.

    EPA Science Inventory

    The oncofetal protein sine oculis-related homeobox 1 (SIX1) is a developmental transcription factor associated with carcinogenesis in animal models and humans. In a model of hormonal carcinogenesis, mice neonatally exposed to the soy phytoestrogen, genistein (GEN), or the synthet...

  18. Application of monoclonal antibodies in tumor pathology

    SciTech Connect

    Ruiter, D.J. ); Fleuren, G.J.; Warnaar, S.O. )

    1987-01-01

    This book contains papers under the following three section headings: Basic and technical aspects; Tumor associated antigens; and Practical application and case presentations. Some of the paper titles are: Monoclonal antibodies to oncofetal antigens; Monoclonal antibodies against ovarian cancer; and Tumor associated antigens and oncogene products defined by monoclonal antibodies.

  19. Investigative approaches to the problem of pancreatic cancer.

    PubMed Central

    Moossa, A. R.

    1979-01-01

    A prospective study of 134 patients suspected of having pancreatic cancer is reported. Ultrasonography and duodenal drainage studies are the best initial investigations. Endoscopic retrograde cholangiopancreatography with cytological examination is the test most likely to provide a definitive diagnosis. Arteriography is essential before laparotomy to delineate anomalies in the foregut vasculature. Pancreatic oncofetal antigen is the only tumour marker that is useful in diagnosis and in monitoring therapy. Images FIG. 5 FIG. 7 PMID:434747

  20. Regulation of human chondrocyte function through direct inhibition of cartilage master regulator SOX9 by microRNA-145 (miRNA-145).

    PubMed

    Martinez-Sanchez, Aida; Dudek, Katarzyna A; Murphy, Chris L

    2012-01-06

    Articular cartilage enables weight bearing and near friction-free movement in the joints. Critical to its function is the production of a specialized, mechanocompetent extracellular matrix controlled by master regulator transcription factor SOX9. Mutations in SOX9 cause campomelic dysplasia, a haploinsufficiency disorder resulting in severe skeletal defects and dwarfism. Although much is understood about how SOX9 regulates cartilage matrix synthesis and hence joint function, how this master regulator is itself regulated remains largely unknown. Here we identify a specific microRNA, miR-145, as a direct regulator of SOX9 in normal healthy human articular chondrocytes. We show that miR-145 directly represses SOX9 expression in human cells through a unique binding site in its 3'-UTR not conserved in mice. Modulation of miR-145 induced profound changes in the human chondrocyte phenotype. Specifically, increased miR-145 levels cause greatly reduced expression of critical cartilage extracellular matrix genes (COL2A1 and aggrecan) and tissue-specific microRNAs (miR-675 and miR-140) and increased levels of the hypertrophic markers RUNX2 and MMP13, characteristic of changes occurring in osteoarthritis. We propose miR-145 as an important regulator of human chondrocyte function and a new target for cartilage repair.

  1. O-GlcNAc inhibits interaction between Sp1 and Elf-1 transcription factors

    SciTech Connect

    Lim, Kihong; Chang, Hyo-Ihl

    2009-03-13

    The novel protein modification, O-linked N-acetylglucosamine (O-GlcNAc), plays an important role in various aspects of cell regulation. Although most of nuclear transcription regulatory factors are modified by O-GlcNAc, O-GlcNAc effects on transcription remain largely undefined yet. In this study, we show that O-GlcNAc inhibits a physical interaction between Sp1 and Elf-1 transcription factors, and negatively regulates transcription of placenta and embryonic expression oncofetal protein gene (Pem). These findings suggest that O-GlcNAc inhibits Sp1-mediated gene transcription possibly by interrupting Sp1 interaction with its cooperative factor.

  2. NEUROPLASTICITY, AXONAL GUIDANCE, AND MICRORNA GENES ARE ASSOCIATED WITH MORPHINE SELF-ADMINISTRATION BEHAVIOR

    PubMed Central

    Tapocik, Jenica D.; Luu, Truong V.; Mayo, Cheryl L.; Wang, Bi-Dar; Doyle, Erin; Lee, Alec D.; Lee, Norman H.; Elmer, Greg I.

    2012-01-01

    Neuroadaptations in the ventral striatum (VS) and ventral midbrain (VMB) following chronic opioid administration are thought to contribute to the pathogenesis and persistence of opiate addiction. In order to identify candidate genes involved in these neuroadaptations we utilized a behavior genetics strategy designed to associate contingent intravenous drug self-administration with specific patterns of gene expression in inbred mice differentially predisposed to the rewarding effects of morphine. In a yoked-control paradigm, C57BL/6J mice showed clear morphine-reinforced behavior whereas DBA/2J mice did not. Moreover, the yoked-control paradigm revealed the powerful consequences of self-administration versus passive administration at the level of gene expression. Morphine self-administration in the C57BL/6J mice uniquely up- or down-regulated 237 genes in the VS and 131 genes in the VMB. Interestingly, only a handful of the C57BL/6J self-administration genes (<3%) exhibited a similar expression pattern in the DBA/2J mice. Hence, specific sets of genes could be confidently assigned to regional effects of morphine in a contingent- and genotype-dependent manner. Bioinformatics analysis revealed that neuroplasticity, axonal guidance and microRNAs (miRNAs) were among the key themes associated with drug self-administration. Noteworthy were the primary miRNA genes H19 and microRNA containing gene (Mirg), processed respectively to mature miRNAs miR-675 and miR-154, since they are prime candidates to mediate network-like changes in responses to chronic drug administration. These miRNAs have postulated roles in dopaminergic neuron differentiation and mu-opioid receptor regulation. The strategic approach designed to focus on reinforcement-associated genes provides new insight into the role of neuroplasticity pathways and miRNAs in drug addiction. PMID:22804800

  3. Expression of the 11p13 Wilms' tumor gene, WT1, correlates with histologic category of Wilms' tumor.

    PubMed Central

    Gerald, W. L.; Gramling, T. S.; Sens, D. A.; Garvin, A. J.

    1992-01-01

    Homozygous inactivation of WT1, a Wilms' tumor gene located on chromosome 11 at p13, is believed to predispose to Wilms' tumor and therefore may be a common occurrence in this cancer. The expression of this gene in primary Wilms' tumors was examined by northern and quantitative RNA slot blot analysis and compared with clinical, histologic, and molecular features of each case. The characteristic 3.2 kb RNA was readily detected in most primary tumors although there was marked variation in the level of WT1-specific transcripts. No abnormal-sized RNA products were detected and expression of WT1 was not coordinated with that of several other oncofetal genes: N-myc, insulinlike growth factor 2 (IGF-2), and c-myc. The relative abundance of WT1-specific RNA did correlate with the histologic category of Wilms' tumor such that those tumors with heterologous differentiation have, in general, lower relative levels of WT1 transcripts than those tumors without heterologous differentiation. These results further establish the heterogeneity of Wilms' tumor with respect to the expression of tumor-associated oncofetal genes and suggest a relationship between WT1 expression and cellular differentiation. Images Figure 1 PMID:1316081

  4. Targeting SALL4 by entinostat in lung cancer

    PubMed Central

    Hong, Clarice Kit Yee; Zhao, Wenxiu; Wang, Fei; Tatetsu, Hiro; Yan, Benedict; Qi, Lihua; Fletcher, Jonathan A.; Yang, Henry; Soo, Ross

    2016-01-01

    The overall survival of lung cancer patients remains dismal despite the availability of targeted therapies. Oncofetal protein SALL4 is a novel cancer target. We herein report that SALL4 was aberrantly expressed in a subset of lung cancer patients with poor survival. SALL4 silencing by RNA interference or SALL4 peptide inhibitor treatment led to impaired lung cancer cell growth. Expression profiling of SALL4-knockdown cells demonstrated that both the EGFR and IGF1R signaling pathways were affected. Connectivity Map analysis revealed the HDAC inhibitor entinostat as a potential drug in treating SALL4-expressing cancers, and this was confirmed in 17 lung cancer cell lines. In summary, we report for the first time that entinostat can target SALL4-positive lung cancer. This lays the foundation for future clinical studies evaluating the therapeutic efficacy of entinostat in SALL4-positive lung cancer patients. PMID:27705911

  5. Genomic analysis of hepatoblastoma identifies distinct molecular and prognostic subgroups.

    PubMed

    Sumazin, Pavel; Chen, Yidong; Treviño, Lisa R; Sarabia, Stephen F; Hampton, Oliver A; Patel, Kayuri; Mistretta, Toni-Ann; Zorman, Barry; Thompson, Patrick; Heczey, Andras; Comerford, Sarah; Wheeler, David A; Chintagumpala, Murali; Meyers, Rebecka; Rakheja, Dinesh; Finegold, Milton J; Tomlinson, Gail; Parsons, D Williams; López-Terrada, Dolores

    2017-01-01

    Despite being the most common liver cancer in children, hepatoblastoma (HB) is a rare neoplasm. Consequently, few pretreatment tumors have been molecularly profiled, and there are no validated prognostic or therapeutic biomarkers for HB patients. We report on the first large-scale effort to profile pretreatment HBs at diagnosis. Our analysis of 88 clinically annotated HBs revealed three risk-stratifying molecular subtypes that are characterized by differential activation of hepatic progenitor cell markers and metabolic pathways: high-risk tumors were characterized by up-regulated nuclear factor, erythroid 2-like 2 activity; high lin-28 homolog B, high mobility group AT-hook 2, spalt-like transcription factor 4, and alpha-fetoprotein expression; and high coordinated expression of oncofetal proteins and stem-cell markers, while low-risk tumors had low lin-28 homolog B and lethal-7 expression and high hepatic nuclear factor 1 alpha activity.

  6. Molecular Mechanisms Regulating the Dendritic Development of Newborn Olfactory Bulb Interneurons in a Sensory Experience-Dependent Manner

    PubMed Central

    Yoshihara, Sei-ichi; Takahashi, Hiroo; Tsuboi, Akio

    2016-01-01

    Inhibitory interneurons in the olfactory bulb are generated continuously throughout life in the subventricular zone and differentiate into periglomerular and granule cells. Neural circuits that undergo reorganization by newborn olfactory bulb interneurons are necessary for odor detection, odor discrimination, olfactory memory, and innate olfactory responses. Although sensory experience has been shown to regulate development in a variety of species and in various structures, including the retina, cortex, and hippocampus, little is known about how sensory experience regulates the dendritic development of newborn olfactory bulb interneurons. Recent studies revealed that the 5T4 oncofetal trophoblast glycoprotein and the neuronal Per/Arnt/Sim domain protein 4 (Npas4) transcription factor regulate dendritic branching and dendritic spine formation, respectively, in olfactory bulb interneurons. Here, we summarize the molecular mechanisms that underlie the sensory input-dependent development of newborn interneurons and the formation of functional neural circuitry in the olfactory bulb. PMID:26793053

  7. Autoimmune damage to spermatogenesis in rodents immunized with mouse F9 embryonic carcinoma cells.

    PubMed Central

    Vojtiskova, M; Pokorna, Z; Draber, P

    1983-01-01

    Significant inhibition of spermatogenesis and appearance of antibodies against spermatogenic cells identified by cytotoxicity and immunofluorescence reactions were observed in mice of inbred strains 129/Sv and BALB/c and in albino guinea pigs after syngeneic, allogeneic, and xenogeneic immunization with mouse F9 embryonic carcinoma cells and Freund's complete adjuvant. A similar syngeneic immunization with PYS-2 cells was ineffective. Appropriate absorption experiments confirmed the similarity between the antigens of F9 and spermatogenic cells and the absence of such a similarity with antigens of PYS-2 cells. These results support the hypothesis that the oncofetal F9 antigens represent spermatogenic differentiation antigens and thus play an essential role in spermatogenic cell differentiation. PMID:6340100

  8. Suppression of hamster lymphocyte reactivity to simian virus 40 tumor surface antigens by spleen cells from pregnant hamsters

    SciTech Connect

    Weppner, W.A.; Adkinson, L.R.; Coggin, J.H.Jr

    1980-09-01

    SV40-transformed tumor cells in hamsters have been found to have cell surface antigens cross-reactive with antigens temporally expressed on fetal tissues. Using a lymphocyte transformation assay, spleen cells from pregnant hamsters were found to be incapable of responding to preparations of either hamster fetal tissue or SV40-transformed cells. However, a suppressor component can be demonstrated in spleen cell populations of both primi-and multiparous hamsters during pregnancy that is capable of reducing the response of lymphocytes sensitized against SV40 tumor-associated antigens. The degree of suppression is proportional to the ratio of responder cells to spleen cells from pregnant animals. These results suggest there is a subpopulation of spleen cells involved in immunoregulation during pregnancy that has the ability to suppress the reactivity of lymphocytes sensitized against SV40-associated oncofetal antigens.

  9. Alpha fetoprotein DNA prime and adenovirus boost immunization of two hepatocellular cancer patients

    PubMed Central

    2014-01-01

    Background Alpha fetoprotein (AFP) is an oncofetal antigen over-expressed by many hepatocellular cancers (HCC). We previously demonstrated that HLA-A2-restricted epitopes derived from AFP are immunogenic in vitro and in vivo despite high circulating levels of this oncofetal antigen. In order to test a more broadly applicable, HLA-unrestricted, inexpensive, cell-free vaccine platform capable of activating tumor antigen-specific CD8+ and CD4+ T cells, we tested full length AFP in a plasmid DNA construct in combination with an AFP-expressing replication-deficient adenovirus (AdV) in a prime-boost vaccine strategy. Methods HCC patients who had an AFP+ tumor and previous treatment for HCC were screened and two patients received vaccination with three plasmid DNA injections followed by a single AdV injection, all delivered intramuscularly (i.m.). Results The vaccine was well tolerated and safe. Both patients showed immunologic evidence of immunization. The first patient had a weak AFP-specific T cell response, a strong AdV-specific cellular response and recurred with an AFP-expressing HCC at nine months. The second patient developed a strong AFP-specific CD8+ and CD4+ cellular response and an AdV neutralizing antibody response, and recurred at 18 months without an increase in serum AFP. Conclusions The AFP DNA prime-AdV boost vaccine was safe and immunogenic. Circulating anti-AdV neutralizing antibodies at baseline did not prohibit the development of AFP-specific cellular immunity. The patient who developed CD8+ and CD4+ AFP-specific T cell immunity had more favorable progression-free survival. The observations with these two patients support development of this vaccine strategy in a larger clinical trial. Trial registration ClinicalTrials.gov: NCT00093548 PMID:24708667

  10. miRNA and mRNA expression profiling identifies members of the miR-200 family as potential regulators of epithelial-mesenchymal transition in pterygium

    PubMed Central

    Engelsvold, David H.; Utheim, Tor P.; Olstad, Ole K.; Gonzalez, Pedro; Eidet, Jon R.; Lyberg, Torstein; Trøseid, Anne-Marie S.; Dartt, Darlene A.; Raeder, Sten

    2014-01-01

    The current study investigates whether microRNA (miRNA) regulators of epithelial-mesenchymal transition (EMT), tissue fibrosis, and angiogenesis are differentially expressed in human primary pterygium. Genome-wide miRNA and mRNA expression profiling of paired pterygium and normal conjunctiva was performed in the context of conventional excision of pterygium with autotransplantation of conjunctiva (n=8). Quantitative real time polymerase chain reaction (qRT-PCR) was used to validate the expression of key molecules previously detected by microarray. In pterygium, 25 miRNAs and 31 mRNAs were significantly differentially expressed by more than two-fold compared to normal conjunctiva. 14 miRNAs were up-regulated (miR-1246, −486, −451, −3172, −3175, −1308, −1972, −143, −211, −665, −1973, −18a, 143, and −663b), whereas 11 were down-regulated (miR-675, −200b-star, −200a-star, −29b, −200b, −210, −141, −31, −200a, −934, and −375). Unsupervised hierarchical cluster analysis demonstrated that members of the miR-200 family were coexpressed and down-regulated in pterygium. The molecular and cellular functions that were most significant to the miRNA data sets were cellular development, cellular growth and proliferation, and cellular movement. qRT-PCR confirmed the expression of 15 of the 16 genes tested and revealed that miR-429 was down-regulated by more than two-fold in pterygium. The concerted down-regulation of four members from both clusters of the miR-200 family (miR-200a/−200b/−429 and miR-200c/−141), which are known to regulate EMT, and up-regulation of the predicted target and mesenchymal marker fibronectin (FN1), suggest that EMT could potentially play a role in the pathogenesis of pterygium and might constitute promising new targets for therapeutic intervention in pterygium. PMID:23872359

  11. SALL4 represents fetal gut differentiation of gastric cancer, and is diagnostically useful in distinguishing hepatoid gastric carcinoma from hepatocellular carcinoma.

    PubMed

    Ushiku, Tetsuo; Shinozaki, Aya; Shibahara, Junji; Iwasaki, Yoshiaki; Tateishi, Yoko; Funata, Nobuaki; Fukayama, Masashi

    2010-04-01

    The novel stem cell marker SALL4 has been identified as a diagnostic marker of germ cell tumors, especially yolk sac tumors, in gonadal organs. To clarify the significance of SALL4 as an oncofetal protein, we investigated SALL4 expression by immunohistochemistry in non-neoplastic stomach and gastric carcinoma with particular emphasis on á-fetoprotein (AFP)-producing gastric carcinoma, as AFP-producing gastric carcinoma shares expression of AFP and glypican 3 (GPC3) with yolk sac tumors and hepatic neoplasms. A total of 338 gastric carcinomas, 60 hepatocellular carcinomas, and 48 cholangiocellular carcinomas were studied by immunohistochemistry on tissue microarrays. In addition, more detailed whole tissue section immunohistochemistry was performed on non-neoplastic gastric tissue from 5 adult and 8 fetal specimens, 6 hepatoblastomas, and 31 cases of AFP-producing gastric carcinomas. SALL4 expression was observed in the neofetal stomach in gestational week 9 and disappeared thereafter. It was also identified by tissue microarray study in a fraction of gastric carcinomas (51 of 338, 15%), associated with older age (P=0.0001), male sex (P=0.0033), intestinal-type histology (P=0.0001), and synchronous liver metastasis (P=0.0047). AFP and GPC3 were closely associated with SALL4 expression in gastric carcinoma (both, P<0.0001), and a full-section study indicated that SALL4 was positive in all 31 cases of AFP-producing gastric carcinoma with diffuse staining in 24 cases (78%). Diffuse SALL4 expression was observed in the histologic patterns of hepatoid (89%), glandular (57%), and clear cell (39%) AFP-producing gastric carcinoma. In addition, SALL4 expression was completely negative in hepatoblastoma (n=6) and hepatocellular carcinoma (n=60). SALL4 is an oncofetal protein similar to AFP and GPC3, but it represents fetal gut differentiation in gastric carcinoma. SALL4 is a sensitive marker for AFP-producing gastric carcinoma and is especially useful to distinguish hepatoid

  12. H19/let-7/LIN28 reciprocal negative regulatory circuit promotes breast cancer stem cell maintenance

    PubMed Central

    Peng, Fei; Li, Ting-Ting; Wang, Kai-Li; Xiao, Guo-Qing; Wang, Ju-Hong; Zhao, Hai-Dong; Kang, Zhi-Jie; Fan, Wen-Jun; Zhu, Li-Li; Li, Mei; Cui, Bai; Zheng, Fei-Meng; Wang, Hong-Jiang; Lam, Eric W-F; Wang, Bo; Xu, Jie; Liu, Quentin

    2017-01-01

    Long noncoding RNA-H19 (H19), an imprinted oncofetal gene, has a central role in carcinogenesis. Hitherto, the mechanism by which H19 regulates cancer stem cells, remains elusive. Here we show that breast cancer stem cells (BCSCs) express high levels of H19, and ectopic overexpression of H19 significantly promotes breast cancer cell clonogenicity, migration and mammosphere-forming ability. Conversely, silencing of H19 represses these BCSC properties. In concordance, knockdown of H19 markedly inhibits tumor growth and suppresses tumorigenesis in nude mice. Mechanistically, we found that H19 functions as a competing endogenous RNA to sponge miRNA let-7, leading to an increase in expression of a let-7 target, the core pluripotency factor LIN28, which is enriched in BCSC populations and breast patient samples. Intriguingly, this gain of LIN28 expression can also feedback to reverse the H19 loss-mediated suppression of BCSC properties. Our data also reveal that LIN28 blocks mature let-7 production and, thereby, de-represses H19 expression in breast cancer cells. Appropriately, H19 and LIN28 expression exhibits strong correlations in primary breast carcinomas. Collectively, these findings reveal that lncRNA H19, miRNA let-7 and transcriptional factor LIN28 form a double-negative feedback loop, which has a critical role in the maintenance of BCSCs. Consequently, disrupting this pathway provides a novel therapeutic strategy for breast cancer. PMID:28102845

  13. H19/let-7/LIN28 reciprocal negative regulatory circuit promotes breast cancer stem cell maintenance.

    PubMed

    Peng, Fei; Li, Ting-Ting; Wang, Kai-Li; Xiao, Guo-Qing; Wang, Ju-Hong; Zhao, Hai-Dong; Kang, Zhi-Jie; Fan, Wen-Jun; Zhu, Li-Li; Li, Mei; Cui, Bai; Zheng, Fei-Meng; Wang, Hong-Jiang; Lam, Eric W-F; Wang, Bo; Xu, Jie; Liu, Quentin

    2017-01-19

    Long noncoding RNA-H19 (H19), an imprinted oncofetal gene, has a central role in carcinogenesis. Hitherto, the mechanism by which H19 regulates cancer stem cells, remains elusive. Here we show that breast cancer stem cells (BCSCs) express high levels of H19, and ectopic overexpression of H19 significantly promotes breast cancer cell clonogenicity, migration and mammosphere-forming ability. Conversely, silencing of H19 represses these BCSC properties. In concordance, knockdown of H19 markedly inhibits tumor growth and suppresses tumorigenesis in nude mice. Mechanistically, we found that H19 functions as a competing endogenous RNA to sponge miRNA let-7, leading to an increase in expression of a let-7 target, the core pluripotency factor LIN28, which is enriched in BCSC populations and breast patient samples. Intriguingly, this gain of LIN28 expression can also feedback to reverse the H19 loss-mediated suppression of BCSC properties. Our data also reveal that LIN28 blocks mature let-7 production and, thereby, de-represses H19 expression in breast cancer cells. Appropriately, H19 and LIN28 expression exhibits strong correlations in primary breast carcinomas. Collectively, these findings reveal that lncRNA H19, miRNA let-7 and transcriptional factor LIN28 form a double-negative feedback loop, which has a critical role in the maintenance of BCSCs. Consequently, disrupting this pathway provides a novel therapeutic strategy for breast cancer.

  14. Reciprocal interplay between thyroid hormone and microRNA-21 regulates hedgehog pathway–driven skin tumorigenesis

    PubMed Central

    Di Girolamo, Daniela; Ambrosio, Raffaele; De Stefano, Maria A.; Mancino, Giuseppina; Porcelli, Tommaso; Luongo, Cristina; Di Cicco, Emery; Scalia, Giulia; Vecchio, Luigi Del; Colao, Annamaria; Dlugosz, Andrzej A.; Missero, Caterina; Salvatore, Domenico

    2016-01-01

    The thyroid hormone–inactivating (TH-inactivating) enzyme type 3 iodothyronine deiodinase (D3) is an oncofetal protein that is rarely expressed in adult life but has been shown to be reactivated in the context of proliferation and neoplasms. D3 terminates TH action within the tumor microenvironment, thereby enhancing cancer cell proliferation. However, the pathological role of D3 and the contribution of TH metabolism in cancer have yet to be fully explored. Here, we describe a reciprocal regulation between TH action and the cancer-associated microRNA-21 (miR21) in basal cell carcinoma (BCC) skin tumors. We found that, besides being negatively regulated by TH at the transcriptional level, miR21 attenuates the TH signal by increasing D3 levels. The ability of miR21 to positively regulate D3 was mediated by the tumor suppressor gene GRHL3, a hitherto unrecognized D3 transcriptional inhibitor. Finally, in a BCC mouse model, keratinocyte-specific D3 depletion markedly reduced tumor growth. Together, our results establish TH action as a critical hub of multiple oncogenic pathways and provide functional and mechanistic evidence of the involvement of TH metabolism in BCC tumorigenesis. Moreover, our results identify a miR21/GRHL3/D3 axis that reduces TH in the tumor microenvironment and has potential to be targeted as a therapeutic approach to BCC. PMID:27159391

  15. Upregulation of H19 indicates a poor prognosis in gallbladder carcinoma and promotes epithelial-mesenchymal transition

    PubMed Central

    Wang, Shou-Hua; Wu, Xiao-Cai; Zhang, Ming-Di; Weng, Ming-Zhe; Zhou, Di; Quan, Zhi-Wei

    2016-01-01

    The imprinted oncofetal long non-coding RNA H19 has been reported to be involved in many kinds of human cancers. However, whether lncRNA H19 implicate in oncogenesis and cancer progression in gallbladder cancer remain largely unknown. In the present study, compared with adjacent normal tissues, the level of H19 was significantly upregulated in gallbladder cancer tissues and was positively associated with lymphatic metastasis and tumor size. The overall survival is shorter in those who had higher H19 expression among GBC patients. In vitro, both TGF-β1 and IL-6 treatment induced upregulation of H19, downregulated the protein level of E-cadherin while increased Vimentin, indicating an epithelial-mesenchymal transition (EMT) phenotype in GBC. The overexpression of H19 in GBC cells enhanced tumor invasion and promoted EMT by upregulated transcription factor Twist1. On the contrary, Loss of function studies indicated that H19 interference in GBC suppressed tumor cell invasion and promoted mesenchymal-epithelial transition (MET) via suppressing Twist expression. In vivo, the volume of the tumors in H19-inteference group was significantly decreased compared to those in the control group of nude mice. Both western-blot and immunohistochemistry confirmed that a MET phenotype existed in the H19 interference group when compared to control group. These results defined H19 as a novel prognostic factor for GBC, and indicated that it might play important regulatory roles in the EMT process. PMID:27073719

  16. WT1 expression is increased in primary fibroblasts derived from Dupuytren's disease tissues.

    PubMed

    Crawford, Justin; Raykha, Christina; Charles, Daevina; Gan, Bing Siang; O'Gorman, David B

    2015-12-01

    Dupuytren's disease (DD) is a fibroproliferative and contractile fibrosis of the palmar fascia that, like all other heritable fibroses, is currently incurable. While DD is invariably benign, it exhibits some molecular similarities to malignant tumours, including increased levels of ß-catenin, onco-fetal fibronectin, periostin and insulin-like growth factor (IGF)-II. To gain additional insights into the pathogenesis of DD, we have assessed the expression of WT1, encoding Wilm's tumour 1, an established tumour biomarker that is syntenic with IGF2, the gene encoding IGF-II in humans. We found that WT1 expression is robustly and consistently up regulated in primary fibroblasts derived from the fibrotic palmar fascia of patients with DD (DD cells), whereas syngeneic fibroblasts derived from the macroscopically unaffected palmar fascia in these patients and allogeneic fibroblasts derived from normal palmar fascia exhibited very low or undetectable WT1 transcript levels. WT1 immunoreactivity was evident in a subset of cells in the fibrotic palmar fascia of patients with DD, but not in macroscopically unaffected palmar fascia. These findings identify WT1 expression as a novel biomarker of fibrotic palmar fascia and are consistent with the hypothesis that the pathogeneses of DD and malignant tumours have molecular similarities.

  17. The Interplay of LncRNA-H19 and Its Binding Partners in Physiological Process and Gastric Carcinogenesis

    PubMed Central

    Zhang, Li; Zhou, Yuhang; Huang, Tingting; Cheng, Alfred S. L.; Yu, Jun; Kang, Wei; To, Ka Fai

    2017-01-01

    Long non-coding RNA (lncRNA), a novel and effective modulator in carcinogenesis, has become a study hotspot in recent years. The imprinted oncofetal lncRNA H19 is one of the first identified imprinted lncRNAs with a high expression level in embryogenesis but is barely detectable in most tissues after birth. Aberrant alterations of H19 expression have been demonstrated in various tumors, including gastric cancer (GC), implicating a crucial role of H19 in cancer progression. As one of the top malignancies in the world, GC has already become a serious concern to public health with poor prognosis. The regulatory roles of H19 in gastric carcinogenesis have been explored by various research groups, which leads to the development of GC therapy. This review comprehensively summarizes the current knowledge of H19 in tumorigenesis, especially in GC pathogenesis, with emphasis on the underneath molecular mechanisms depicted from its functional partners. Furthermore, the accumulated knowledge of H19 will provide better understanding on targeted therapy of GC. PMID:28230721

  18. Type II congenital pulmonary airway malformation associated with intralobar pulmonary sequestration: report of a case and review of classification criteria.

    PubMed

    Mastrogiulio, M G; Barone, A; Disanto, M G; Ginori, A; Ambrosio, M R; Carbone, S F; Spina, D

    2016-03-01

    Pulmonary congenital abnormalities are rare disorders including congenital pulmonary airway malformations (CPAM) and pulmonary sequestration (PS). CPAM is a lesion characterized by the presence of anomalous bronchiolar or acinar structures, variable in size, either cystic or not cystic. PS is generally defined as nonfunctioning lung tissue that is not in normal continuity with the tracheobronchial tree and that derives its blood supply from systemic vessels. We describe a case of a baby girl with a very rare association between CPAM type 2 and intralobar pulmonary sequestration (IPS) focusing on the cystic lesions typical of CPAM and on the lymphatic and blood vessels. The cells lining the cysts often were positive for D2-40 (oncofetal protein M2A). Lymphatic endothelial cells, positive for D2-40, were widely present in the lung parenchyma and dilated lymphatic vessels were present also in the inter-alveolar septa. Moreover, we discuss the pathogenesis of CPAM and its classification criteria. © Copyright Società Italiana di Anatomia Patologica e Citopatologia Diagnostica, Divisione Italiana della International Academy of Pathology.

  19. The B-cell tumor–associated antigen ROR1 can be targeted with T cells modified to express a ROR1-specific chimeric antigen receptor

    PubMed Central

    Schmitt, Thomas M.; Baskar, Sivasubramanian; Lupo-Stanghellini, Maria Teresa; Nishida, Tetsuya; Yamamoto, Tori N.; Bleakley, Marie; Turtle, Cameron J.; Chang, Wen-Chung; Greisman, Harvey A.; Wood, Brent; Maloney, David G.; Jensen, Michael C.; Rader, Christoph; Riddell, Stanley R.

    2010-01-01

    Monoclonal antibodies and T cells modified to express chimeric antigen receptors specific for B-cell lineage surface molecules such as CD20 exert antitumor activity in B-cell malignancies, but deplete normal B cells. The receptor tyrosine kinase-like orphan receptor 1 (ROR1) was identified as a highly expressed gene in B-cell chronic lymphocytic leukemia (B-CLL), but not normal B cells, suggesting it may serve as a tumor-specific target for therapy. We analyzed ROR1-expression in normal nonhematopoietic and hematopoietic cells including B-cell precursors, and in hematopoietic malignancies. ROR1 has characteristics of an oncofetal gene and is expressed in undifferentiated embryonic stem cells, B-CLL and mantle cell lymphoma, but not in major adult tissues apart from low levels in adipose tissue and at an early stage of B-cell development. We constructed a ROR1-specific chimeric antigen receptor that when expressed in T cells from healthy donors or CLL patients conferred specific recognition of primary B-CLL and mantle cell lymphoma, including rare drug effluxing chemotherapy resistant tumor cells that have been implicated in maintaining the malignancy, but not mature normal B cells. T-cell therapies targeting ROR1 may be effective in B-CLL and other ROR1-positive tumors. However, the expression of ROR1 on some normal tissues suggests the potential for toxi-city to subsets of normal cells. PMID:20702778

  20. The B-cell tumor-associated antigen ROR1 can be targeted with T cells modified to express a ROR1-specific chimeric antigen receptor.

    PubMed

    Hudecek, Michael; Schmitt, Thomas M; Baskar, Sivasubramanian; Lupo-Stanghellini, Maria Teresa; Nishida, Tetsuya; Yamamoto, Tori N; Bleakley, Marie; Turtle, Cameron J; Chang, Wen-Chung; Greisman, Harvey A; Wood, Brent; Maloney, David G; Jensen, Michael C; Rader, Christoph; Riddell, Stanley R

    2010-11-25

    Monoclonal antibodies and T cells modified to express chimeric antigen receptors specific for B-cell lineage surface molecules such as CD20 exert antitumor activity in B-cell malignancies, but deplete normal B cells. The receptor tyrosine kinase-like orphan receptor 1 (ROR1) was identified as a highly expressed gene in B-cell chronic lymphocytic leukemia (B-CLL), but not normal B cells, suggesting it may serve as a tumor-specific target for therapy. We analyzed ROR1-expression in normal nonhematopoietic and hematopoietic cells including B-cell precursors, and in hematopoietic malignancies. ROR1 has characteristics of an oncofetal gene and is expressed in undifferentiated embryonic stem cells, B-CLL and mantle cell lymphoma, but not in major adult tissues apart from low levels in adipose tissue and at an early stage of B-cell development. We constructed a ROR1-specific chimeric antigen receptor that when expressed in T cells from healthy donors or CLL patients conferred specific recognition of primary B-CLL and mantle cell lymphoma, including rare drug effluxing chemotherapy resistant tumor cells that have been implicated in maintaining the malignancy, but not mature normal B cells. T-cell therapies targeting ROR1 may be effective in B-CLL and other ROR1-positive tumors. However, the expression of ROR1 on some normal tissues suggests the potential for toxi-city to subsets of normal cells.

  1. Two Isoforms of the RNA Binding Protein, Coding Region Determinant-binding Protein (CRD-BP/IGF2BP1), Are Expressed in Breast Epithelium and Support Clonogenic Growth of Breast Tumor Cells.

    PubMed

    Fakhraldeen, Saja A; Clark, Rod J; Roopra, Avtar; Chin, Emily N; Huang, Wei; Castorino, John; Wisinski, Kari B; Kim, TaeWon; Spiegelman, Vladimir S; Alexander, Caroline M

    2015-05-22

    CRD-BP/IGF2BP1 has been characterized as an "oncofetal" RNA binding protein typically highly expressed in embryonic tissues, suppressed in normal adult tissues, but induced in many tumor types. In this study, we show that adult breast tissues express ubiquitous but low levels of CRD-BP protein and mRNA. Although CRD-BP mRNA expression is induced in breast tumor cells, levels remain ∼1000-fold lower than in embryonic tissues. Despite low expression levels, CRD-BP is required for clonogenic growth of breast cancer cells. We reveal that because the most common protein isoform in normal adult breast and breast tumors has an N-terminal deletion (lacking two RNA recognition motif (RRM) domains) and is therefore missing antibody epitopes, CRD-BP expression has been under-reported by previous studies. We show that a CRD-BP mutant mouse strain retains expression of the shorter transcript (ΔN-CRD-BP), which originates in intron 2, suggesting that the impact of complete ablation of this gene in mice is not yet known. Either the full-length CRD-BP or the N-terminally truncated version can rescue the clonogenicity of CRD-BP knockdown breast cancer cells, suggesting that clonogenic function is served by either CRD-BP isoform. In summary, although CRD-BP expression levels are low in breast cancer cells, this protein is necessary for clonogenic activity.

  2. Nature of immobilization surface affects antibody specificity to placental alkaline phosphatase.

    PubMed

    Kumar, Mukesh; Khan, Imran; Sinha, Subrata

    2015-01-01

    Retention of native conformation of immobilized protein is essential for various applications including selection and detection of specific recombinant antibodies (scFvs). Placental alkaline phosphatase (PAP), an onco-fetal antigen expressed on the surface of several tumors, was immobilized on supermagnetic particles for selection of recombinant antibodies from a human phage display antibody library. The isolated antibodies were found to be cross-reactive to either of the isozymes of alkaline phosphatase, i.e., bone alkaline phosphatase (BAP) or intestinal alkaline phosphatase (IAP) and could not be used for tumor targeting. A specific anti-PAP monoclonal antibody H17E2 was tested for retention of specificity under these conditions. Binding of the antibody to magnetic beads conjugated IAP and BAP along with PAP and the ability of the two isozymes to inhibit its binding to PAP depicted the loss of isozyme specificity of the antibody. However, the antibody retained its specificity to PAP immobilized on polyvinyl chloride (PVC) surface. Enzyme activity was observed on both surfaces. This demonstrates that nature of immobilization may affect antigen-antibody binding in subtle ways, resulting in alteration of conformation of the epitopes. This may have consequences for determining the specificity of antibody binding for proteins that share a high degree of homology.

  3. Use of H19 Gene Regulatory Sequences in DNA-Based Therapy for Pancreatic Cancer

    PubMed Central

    Scaiewicz, V.; Sorin, V.; Fellig, Y.; Birman, T.; Mizrahi, A.; Galula, J.; Abu-lail, R.; Shneider, T.; Ohana, P.; Buscail, L.; Hochberg, A.; Czerniak, A.

    2010-01-01

    Pancreatic cancer is the eighth most common cause of death from cancer in the world, for which palliative treatments are not effective and frequently accompanied by severe side effects. We propose a DNA-based therapy for pancreatic cancer using a nonviral vector, expressing the diphtheria toxin A chain under the control of the H19 gene regulatory sequences. The H19 gene is an oncofetal RNA expressed during embryo development and in several types of cancer. We tested the expression of H19 gene in patients, and found that 65% of human pancreatic tumors analyzed showed moderated to strong expression of the gene. In vitro experiments showed that the vector was effective in reducing Luciferase protein activity on pancreatic carcinoma cell lines. In vivo experiment results revealed tumor growth arrest in different animal models for pancreatic cancer. Differences in tumor size between control and treated groups reached a 75% in the heterotopic model (P = .037) and 50% in the orthotopic model (P = .007). In addition, no visible metastases were found in the treated group of the orthotopic model. These results indicate that the treatment with the vector DTA-H19 might be a viable new therapeutic option for patients with unresectable pancreatic cancer. PMID:21052499

  4. Acetylation directs survivin nuclear localization to repress STAT3 oncogenic activity.

    PubMed

    Wang, Haijuan; Holloway, Michael P; Ma, Li; Cooper, Zachary A; Riolo, Matthew; Samkari, Ayman; Elenitoba-Johnson, Kojo S J; Chin, Y Eugene; Altura, Rachel A

    2010-11-12

    The multiple functions of the oncofetal protein survivin are dependent on its selective expression patterns within immunochemically distinct subcellular pools. The mechanism by which survivin localizes to these compartments, however, is only partly understood. Here we show that nuclear accumulation of survivin is promoted by CREB-binding protein (CBP)-dependent acetylation on lysine 129 (129K, Lys-129). We demonstrate a mechanism by which survivin acetylation at this position results in its homodimerization, while deacetylation promotes the formation of survivin monomers that heterodimerize with CRM1 and facilitate its nuclear export. Using proteomic analysis, we identified the oncogenic transcription factor STAT3 as a binding partner of nuclear survivin. We show that acetylated survivin binds to the N-terminal transcriptional activation domain of the STAT3 dimer and represses STAT3 transactivation of target gene promoters. Using multiplex PCR and DNA sequencing, we identified a single-nucleotide polymorphism (A → G) at Lys-129 that exists as a homozygous mutation in a neuroblastoma cell line and corresponds with a defect in survivin nuclear localization. Our results demonstrate that the dynamic equilibrium between survivin acetylation and deacetylation at amino acid 129 determines its interaction with CRM1, its subsequent subcellular localization, and its ability to inhibit STAT3 transactivation, providing a potential route for therapeutic intervention in STAT3-dependent tumors.

  5. Spontaneous development of hepatocellular carcinoma with cancer stem cell properties in PR-SET7-deficient livers

    PubMed Central

    Nikolaou, Kostas C; Moulos, Panagiotis; Chalepakis, George; Hatzis, Pantelis; Oda, Hisanobu; Reinberg, Danny; Talianidis, Iannis

    2015-01-01

    PR-SET7-mediated histone 4 lysine 20 methylation has been implicated in mitotic condensation, DNA damage response and replication licensing. Here, we show that PR-SET7 function in the liver is pivotal for maintaining genome integrity. Hepatocyte-specific deletion of PR-SET7 in mouse embryos resulted in G2 phase arrest followed by massive cell death and defect in liver organogenesis. Inactivation at postnatal stages caused cell duplication-dependent hepatocyte necrosis, accompanied by inflammation, fibrosis and compensatory growth induction of neighboring hepatocytes and resident ductal progenitor cells. Prolonged necrotic regenerative cycles coupled with oncogenic STAT3 activation led to the spontaneous development of hepatic tumors composed of cells with cancer stem cell characteristics. These include a capacity to self-renew in culture or in xenografts and the ability to differentiate to phenotypically distinct hepatic cells. Hepatocellular carcinoma in PR-SET7-deficient mice displays a cancer stem cell gene signature specified by the co-expression of ductal progenitor markers and oncofetal genes. PMID:25515659

  6. [Alphafetoprotein in hepatic tumours and benign liver diseases].

    PubMed

    Forones, N M; Queiroz, L A; Ferraz, M L; Parise, E R

    1995-01-01

    AFP is an oncofetal protein found in increased levels in hepatocellular carcinoma, liver metastasis and other benign liver diseases. PURPOSE--To know the behaviour of this protein in each of these clinical situations would undoubtedly help us to discriminate between hepatocellular carcinoma and benign diseases. PATIENTS--A hundred forty nine patients were divided into 4 groups: 1. acute hepatitis (AH) n = 24, 2. chronic liver disease, viral or alcoholic (CLD) n = 81, 3. hepatic metastasis (HM) n = 29, 4. hepatocellular carcinoma (HCC) n = 15. AFP assays were done by ELISA (Abbott Diagnostica, ref. value: 15ng/mL). RESULTS--The results observed were as follows: AFP < 15ng/mL: AH 75%, CLD 86.4%, HM 79.3%, HCC 6.6%, AFP > 15 e < 100ng/mL: AH 25%, CLD 8.6%, HM 20.6%, HCC 20%, AFP > 100ng/mL: AH zero, CLD 4.9%, HM zero, HCC 49%. It is clear that depending on the cut off level, there is a decrease of sensibility which is paralleled by an increase in specificity. CONCLUSIONS--AFP levels are increased in benign liver diseases (AH, CLD) and HM, how ever levels above 100ng/mL occur much more frequently in HCC. In our sample, 93.3% of the HCC showed high levels of AFP, probably because most of the patients had advanced clinical stages of the disease.

  7. Genetic Differences Between Great Apes and Humans: Implications for Human Evolution

    SciTech Connect

    Varki, Ajit

    2004-03-17

    When considering protein sequences, humans are 99-100% identical to chimpanzees and bonobos, our closest evolutionary relatives. The evolution of humans (and the unique features of our species) from a common ancestor with these great apes involved many steps, influenced by interactions amongst factors of genetic, developmental, ecological, microbial, climatic, behavioral, cultural and social origin. The genetic factors can be approached by direct comparisons of human and great ape genomes, genes and gene products, and by elucidating biochemical and biological consequences of the differences. We have discovered multiple genetic and biochemical differences between humans and great apes, particularly in relationship to a family of cell surface molecules called sialic acids. These differences have implications for the human condition, ranging from susceptibility or resistance to microbial pathogens; effects on endogenous receptors in the immune system; potential effects on placental signaling; the expression of oncofetal antigens in cancers; consequences of dietary intake of animal foods; and the development of the mammalian brain. This talk will provide an overview of these and other genetic differences between humans and great apes, with attention to differences potentially relevant to the evolution of humans.

  8. Genetic Differences Between Humans and Great Apes -- Implications for the Evolution of Humans

    NASA Astrophysics Data System (ADS)

    Varki, Ajit

    2004-06-01

    At the level of individual protein sequences, humans are 97-100% identical to the great apes, our closest evolutionary relatives. The evolution of humans (and of human intelligence) from a common ancestor with the chimpanzee and bonobo involved many steps, influenced by interactions amongst factors of genetic, developmental, ecological, microbial, climatic, behavioral, cultural and social origin. The genetic factors can be approached by direct comparisons of human and great ape genomes, genes and gene products, and by elucidating biochemical and biological consequences of any differences found. We have discovered multiple genetic and biochemical differences between humans and great apes, particularly with respect to a family of cell surface molecules called sialic acids, as well as in the metabolism of thyroid hormones. The hormone differences have potential consequences for human brain development. The differences in sialic acid biology have multiple implications for the human condition, ranging from susceptibility or resistance to microbial pathogens, effects on endogenous receptors in the immune system, and potential effects on placental signaling, expression of oncofetal antigens in cancers, consequences of dietary intake of animal foods, and development of the mammalian brain.

  9. Immunohistochemical study of pancreatoblastoma.

    PubMed

    Ohaki, Y; Misugi, K; Fukuda, J; Okudaira, M; Hirose, M

    1987-10-01

    Three cases of pancreatoblastoma in children were examined immunohistochemically and the results were compared with those of pancreatic duct carcinoma in adults. The pancreatoblastoma demonstrated positive reactions to alpha-fetoprotein (AFP) (67%: 2/3), alpha-1-antitrypsin (AAT) (100%: 3/3), carcinoembryonic antigen (CEA) (67%: 2/3) and keratin (33%: 1/3), although CEA was only weakly positive in both cases. On the other hand, adult pancreatic duct carcinoma showed positive reactions as follows; AFP: 3% (1/29), AAT: 21% (6/29), CEA: 97% (28/29) and keratin: 93% (27/29). Also, endocrine substances including insulin, glucagon and somatostatin were all negative in the pancreatoblastomas. Two cases of pancreatoblastoma which were immunohistochemically positive for AFP also showed elevation of the serum AFP level clinically. The different expressive pattern of oncofetal antigens in pancreatoblastoma as compared with pancreatic duct carcinoma in adults may provide further supporting evidence for the embryonic nature of pancreatoblastoma, and suggests that such a pattern might be used as a tumor marker for pancreatoblastoma.

  10. Enzyme-linked PNA lectin binding assay compared with CA19-9 and CEA radioimmunoassay as a diagnostic blood test for pancreatic cancer.

    PubMed Central

    Ching, C. K.; Rhodes, J. M.

    1989-01-01

    Previous studies have shown that sera from patients with pancreatic cancer often contain a mucus glycoprotein that expresses the oncofetal antigen galactose 1-3, N-acetyl galactosamine, which is the T blood group antigen and the binding site for the lectin peanut agglutinin (PNA). An enzyme-linked lectin assay has been developed to quantify PNA-binding glycoproteins in serum and has been evaluated as a serological test for pancreatic cancer. Sera were studied from 53 patients with pancreatic cancer and 154 controls, including benign obstructive jaundice, acute and chronic pancreatitis, chronic liver disease and inflammatory bowel disease. The enzyme-linked peanut lectin assay proved highly reproducible and has 77% sensitivity and 83% specificity for pancreatic cancer, results that are very similar to those achieved in the same sera by CA19-9 radioimmunoassay (75% sensitivity, 82% specificity with the upper limit of normal set at 37 u ml-1). CEA assay proved less useful (60% sensitivity, 47% specificity). In this study better results were obtained if an upper limit of normal of 50 u ml-1 was used for CA19-9 (75% sensitivity, 92% specificity). Combination of CA19-9 assay with the upper limit set at 50 u ml-1 and the peanut lectin assay improved the sensitivity to 85% with only a slight fall in specificity (85%). These results compare well with published results for ultrasound and CT scanning. PMID:2736232

  11. Evaluation of MVA-5T4 as a novel immunotherapeutic vaccine in colorectal, renal and prostate cancer.

    PubMed

    Amato, Robert J; Stepankiw, Mika

    2012-03-01

    This paper reviews the development of the combination of modified vaccinia Ankara (MVA) to deliver the tumor-associated antigen 5T4 as a novel immunotherapeutic vaccine. The oncofetal antigen 5T4 is highly expressed in 80% of breast, kidney, colorectal, prostate and ovarian carcinomas, making it an ideal antigen for vaccine therapy. To date, more than 3000 doses of MVA-5T4 have been administered to colorectal, renal and prostate cancer patients, with rare occurrences of grade 3 or 4 vaccination-related adverse events being observed. Studies have demonstrated that MVA-5T4 is safe and highly immunogenic, both as monotherapy and in combination with other standard of care therapies. Although an immune response has been observed, antitumor activity has been modest or absent in clinical trials. A Phase III trial resulted in the development of an immune response surrogate that is to be applied to all future MVA-5T4 clinical trials. With minimal side effects and the ability to produce a strong immunogenic response, MVA-5T4 is a viable addition to the cancer therapy arsenal.

  12. Carcinoembryonic antigen (CEA)-based cancer vaccines: recent patents and antitumor effects from experimental models to clinical trials.

    PubMed

    Turriziani, Mario; Fantini, Massimo; Benvenuto, Monica; Izzi, Valerio; Masuelli, Laura; Sacchetti, Pamela; Modesti, Andrea; Bei, Roberto

    2012-09-01

    Carcinoembryonic antigen (CEA), a glycosylated protein of MW 180 kDa, is overexpressed in a wide range of human carcinomas, including colorectal, gastric, pancreatic, non-small cell lung and breast carcinomas. Accordingly, CEA is one of several oncofetal antigens that may serve as a target for active anti-cancer specific immunotherapy. Experimental results obtained by employing animal models have supported the design of clinical trials using a CEA-based vaccine for the treatment of different types of human cancers. This review reports findings from experimental models and clinical evidence on the use of a CEA-based vaccine for the treatment of cancer patients. Among the diverse CEA-based cancer vaccines, DCs- and recombinant viruses-based vaccines seem the most valid. However, although vaccination was shown to induce a strong immune response to CEA, resulting in a delay in tumor progression and prolonged survival in some cancer patients, it failed to eradicate the tumor in most cases, owing partly to the negative effect exerted by the tumor microenvironment on immune response. Thus, in order to develop more efficient and effective cancer vaccines, it is necessary to design new clinical trials combining cancer vaccines with chemotherapy, radiotherapy and drugs which target those factors responsible for immunosuppression of immune cells. This review also discusses relevant patents relating to the use of CEA as a cancer vaccine.

  13. An unusual primary malignant tumor of the stomach: Fetal gut-like Gastric adenocarcinoma with "blastoma"-like component.

    PubMed

    Taher, Altaf; Denic, Nebojsa; Kalimuthu, Sangeetha N; Chetty, Runjan

    2017-03-15

    An unusual case of a polypoid, malignant gastric tumor in a 62-year man is presented. Endoscopy and subsequent polypectomy revealed an 8.5 x 6.5 x 4.5cm lesion in the body of the stomach. Microscopy showed surface dysplasia with an invasive adenocarcinoma displaying prominent tubulopapillary areas composed of large vacuolated cells, pleomorphic nuclei and occasional cytoplasmic hyaline globules. This component then blended with tubular structures lined by more primitive appearing vacuolated cells embedded within a stroma made up of cellular primitive, high-grade blastema-like areas and, less cellular more pleomorphic foci with spindle and several bizarre, large cells. Immunohistochemistry showed the adenocarcinoma and primitive tubules to be strongly SALL4 and epithelial marker positive, but only focal expression of α-fetoprotein and glypican-3. The stromal component made up of blastema-like areas displayed strong immunoreactivity for glypican-3. The pleomorphic stromal areas were negative for all markers, including epithelial and muscle markers. The overall morphology and expression of primitive oncofetal proteins, especially SALL4 and glypican-3, are in keeping with this being a primitive adenocarcinoma showing fetal gut-like differentiation with an accompanying blastoma-like component, a combination not previously described in a primary gastric cancer.

  14. Dysregulated serum response factor triggers formation of hepatocellular carcinoma

    PubMed Central

    Ohrnberger, Stefan; Thavamani, Abhishek; Braeuning, Albert; Lipka, Daniel B; Kirilov, Milen; Geffers, Robert; Authenrieth, Stella E; Römer, Michael; Zell, Andreas; Bonin, Michael; Schwarz, Michael; Schütz, Günther; Schirmacher, Peter; Plass, Christoph; Longerich, Thomas; Nordheim, Alfred

    2015-01-01

    The ubiquitously expressed transcriptional regulator serum response factor (SRF) is controlled by both Ras/MAPK (mitogen-activated protein kinase) and Rho/actin signaling pathways, which are frequently activated in hepatocellular carcinoma (HCC). We generated SRF-VP16iHep mice, which conditionally express constitutively active SRF-VP16 in hepatocytes, thereby controlling subsets of both Ras/MAPK- and Rho/actin-stimulated target genes. All SRF-VP16iHep mice develop hyperproliferative liver nodules that progresses to lethal HCC. Some murine (m)HCCs acquire Ctnnb1 mutations equivalent to those in human (h)HCC. The resulting transcript signatures mirror those of a distinct subgroup of hHCCs, with shared activation of oncofetal genes including Igf2, correlating with CpG hypomethylation at the imprinted Igf2/H19 locus. Conclusion: SRF-VP16iHep mHCC reveal convergent Ras/MAPK and Rho/actin signaling as a highly oncogenic driver mechanism for hepatocarcinogenesis. This suggests simultaneous inhibition of Ras/MAPK and Rho/actin signaling as a treatment strategy in hHCC therapy. (Hepatology 2015;61:979–989) PMID:25266280

  15. Differential expression of IMP3 between male and female mature teratomas--immunohistochemical evidence of malignant nature.

    PubMed

    Goodman, Steven; Zhang, Liping; Cheng, Liang; Jiang, Zhong

    2014-10-01

    Ovarian mature teratoma is a benign tumour, whereas mature teratoma in adult testicular germ cell tumours (TGCTs) is considered to be a malignant tumour. IMP3, an oncofetal protein, plays an important role in embryogenesis and carcinogenesis. IMP3 has been demonstrated to be a malignant biomarker that is mainly expressed in malignant neoplasms rather than benign tissues. The aim of this study was to analyse IMP3 expression in germ cell tumours, and compare its expression between male and female teratomas. One hundred and seventy-eight cases (62 TGCTs, 52 ovarian teratomas, 27 metastatic testicular teratomas, and 37 cases of normal testicular tissue) obtained from the archives of two large academic medical centres were examined for IMP3 expression. Of the 62 TGCTs, 30 had mature teratoma components. IMP3 expression was present in 100% (30/30) of testicular mature teratoma components, and in 96% (26/27) of metastatic testicular teratomas. Other TGCT components also expressed IMP3 in 99% of cases (78/79). IMP3 expression was negative in all female mature teratomas. We describe for the first time an immunostaining marker that has differential expression in male and female mature teratomas, indicating that their pathogenesis differs. High expression of IMP3 in adult mature testicular teratomas supports their malignant nature. © 2014 John Wiley & Sons Ltd.

  16. Aberrant glycosylation promotes lung cancer metastasis through adhesion to galectins in the metastatic niche

    PubMed Central

    Reticker-Flynn, Nathan E.; Bhatia, Sangeeta N.

    2015-01-01

    Metastasis is the leading cause of cancer-associated deaths. While dissemination of tumor cells likely occurs early in tumorigenesis, the constituents of the microenvironment play essential rate-limiting roles in determining whether these cells will form clinically-relevant tumors. Recent studies have uncovered many molecular factors that contribute to establishment of a pro-tumorigenic metastatic niche. Here, we demonstrate that galectin-3, whose expression has clinical associations with advanced malignancy and poor outcome, contributes to metastatic niche formation by binding to carbohydrates on metastatic cells. We show that galectin-3 is expressed early during tumorigenesis by both CD11b+Gr-1+ and CD11b+Ly-6Chi leukocytes. Tumors mobilize these myeloid populations through secretion of soluble factors including IL-6. We find that metastatic cancer cells exhibit elevated presentation of the oncofetal galectin-3 carbohydrate ligand, the Thomsen-Friedenreich Antigen, on their surfaces as a result of altered C2GnT2 and St6GalNAcIV glycosyltransferase activity that inhibits further glycosylation of this carbohydrate motif and promotes metastasis. PMID:25421439

  17. IMP3 expression in serous tumors of the ovary.

    PubMed

    Chisté, Marcela; Alexis, John; Recine, Monica

    2014-10-01

    Insulin-like growth factor II mRNA-binding protein (IMP3) is an oncofetal protein involved in embryogenesis, which is expressed in a variety of malignant neoplasms. It is rarely expressed in normal adult tissue and benign tumors. The aim of this study was to evaluate the expression of IMP3 in benign and malignant serous tumors of the ovary. Seventy-nine ovarian tumors were examined for IMP3 expression by immunohistochemical analysis, comprising 16 benign serous tumors, 19 borderline serous tumors, and 44 serous carcinomas. Positive staining was defined as brown staining in the cytoplasm. Negative staining was defined as absent staining or staining of <5% of tumor cells. The intensity of staining (weak, moderate, and strong) and percentage (0% to 100%) of neoplastic cells staining positive for cytoplasmic IMP3 staining were recorded in each case. Moderate to strong cytoplasmic staining for IMP3 was observed in 30 of 44 (68%) serous carcinomas of the ovary; in contrast, <5% of the borderline and benign serous tumors expressed IMP3 ranging from weak to strong cytoplasmic staining. Statistically, the difference in IMP3 expression between these groups of tumors was highly significant (P<0.0001). Our findings demonstrate moderate to strong expression of IMP3 in the majority of ovarian serous carcinomas as compared with benign/borderline serous tumors, which demonstrated weak to strong expression in a small minority (<5%) of the tumors. Thus, IMP3 may be a useful adjunctive tool in the pathologic evaluation of ovarian serous tumors.

  18. Anti-Cadherin-17 Antibody Modulates Beta-Catenin Signaling and Tumorigenicity of Hepatocellular Carcinoma

    PubMed Central

    Liu, Ling Xiao; Zhang, Xiao Qian; Yuan, Yi; Khin, Ester; Hu, Mei-yu; Wang, Jian Hua; Poon, Ronnie T. P.; Hong, Wanjin; Lee, Nikki P.; Luk, John M.

    2013-01-01

    Cadherin-17 (CDH17) is an oncofetal molecule associated with poor prognostic outcomes of hepatocellular carcinoma (HCC), for which the treatment options are very limited. The present study investigates the therapeutic potential of a monoclonal antibody (Lic5) that targets the CDH17 antigen in HCC. In vitro experiments showed Lic5 could markedly reduce CDH17 expression in a dose-dependent manner, suppress β-catenin signaling, and induce cleavages of apoptotic enzymes caspase-8 and -9 in HCC cells. Treatment of animals in subcutaneous HCC xenograft model similarly demonstrated significant tumor growth inhibition (TGI) using Lic5 antibody alone (5 mg/kg, i.p., t.i.w.; ca.60–65% TGI vs. vehicle at day 28), or in combination with conventional chemotherapy regimen (cisplatin 1 mg/kg; ca. 85–90% TGI). Strikingly, lung metastasis was markedly suppressed by Lic5 treatments. Immunohistochemical and western blot analyses of xenograft explants revealed inactivation of the Wnt pathway and suppression of Wnt signaling components in HCC tissues. Collectively, anti-CDH17 antibody promises as an effective biologic agent for treating malignant HCC. PMID:24039755

  19. High-throughput fluorescence anisotropy screen for inhibitors of the oncogenic mRNA binding protein, IMP-1.

    PubMed

    Mahapatra, Lily; Mao, Chengjian; Andruska, Neal; Zhang, Chen; Shapiro, David J

    2014-03-01

    Cancer cell proliferation is regulated by oncogenes, such as c-Myc. An alternative approach to directly targeting individual oncogenes is to target IMP-1, an oncofetal protein that binds to and stabilizes messenger RNAs (mRNAs), leading to elevated expression of c-Myc and other oncogenes. Expression of IMP-1 is tightly correlated with a poor prognosis and reduced survival in ovarian, lung, and colon cancer. Small-molecule inhibitors of IMP-1 have not been reported. We established a fluorescence anisotropy/polarization microplate assay (FAMA) for analyzing binding of IMP-1 to a fluorescein-labeled 93 nucleotide c-Myc mRNA target (flMyc), developed the assay as a highly robust (Z' factor = 0.60) FAMA-based high-throughput screen for inhibitors of binding of IMP-1 to flMyc, and carried out a successful pilot screen of 17,600 small molecules. Our studies support rapidly filtering out toxic nonspecific inhibitors using an early cell-based assay in control cells lacking the target protein. The physiologic importance of verified hits from the in vitro high-throughput screen was demonstrated by identification of the first small-molecule IMP-1 inhibitor, a lead compound that selectively inhibits proliferation of IMP-1-positive cancer cells with very little or no effect on proliferation of IMP-1-negative cells.

  20. High-Throughput Fluorescence Anisotropy Screen for Inhibitors of the Oncogenic mRNA-binding Protein, IMP-1

    PubMed Central

    Mahapatra, Lily; Mao, Chengjian; Andruska, Neal; Zhang, Chen; Shapiro, David J.

    2014-01-01

    Cancer cell proliferation is regulated by oncogenes, such as c-Myc. An alternative approach to directly targeting individual oncogenes is to target IMP-1, an oncofetal protein that binds to and stabilizes mRNAs, leading to elevated expression of c-Myc and other oncogenes. Expression of IMP-1 is tightly correlated with a poor prognosis and reduced survival in ovarian, lung and colon cancer. Small molecule inhibitors of IMP-1 have not been reported. We established a fluorescence anisotropy/polarization microplate assay (FAMA) for analyzing binding of IMP-1 to a fluorescein-labeled 93 nucleotide c-Myc mRNA target (flMyc), developed the assay as a highly robust (Z’ factor = 0.60) FAMA-based high throughput screen for inhibitors of binding of IMP-1 to flMyc, and carried out a successful pilot screen of 17,600 small molecules. Our studies support rapidly filtering out toxic non-specific inhibitors using an early cell-based assay in control cells lacking the target protein. The physiologic importance of verified hits from the in vitro high throughput screen was demonstrated by identification of the first small molecule IMP-1 inhibitor; a lead compound that selectively inhibits proliferation of IMP-1 positive cancer cells with very little or no effect on proliferation of IMP-1 negative cells. PMID:24108120

  1. Effect of increasing maximal aerobic exercise on serum gonadal hormones and alpha-fetoprotein in the luteal phase of professional female soccer players.

    PubMed

    Otağ, Aynur; Hazar, Muhsin; Otağ, İlhan; Beyleroğlu, Malik

    2016-03-01

    [Purpose] The performance of female athletes during their menstrual period has attracted the attention of researchers for many years. It is known that the menstrual period changes with exercise. Alpha-fetoprotein (AFP) is an oncofetal protein. In this study, the effect of maximal aerobic exercise in the luteal phase on some hormones and AFP in female athletes was researched. [Subjects and Methods] Twelve volunteers and healthy female footballers with normal menstrual cycles volunteered for this study as subjects. All the participants performed a shuttle run test. Blood samples were taken before, after, and one hour after exercise. Serum AFP, estrogen, progesterone, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) values were measured using an auto analyzer and original kits. Heart rate measurements were performed before and after the exercise. [Results] AFP activity had significantly decreased after 1 h of recovery from the exercise in the female soccer players, and estrogen and LH activity had significantly increased immediately after the exercise. Progesterone activity had significantly decreased immediately after the exercise. FSH values had significantly increased immediately after the exercise. [Conclusion] The results of the present study show there were significant decreases in the values of AFP, which is a cancer parameter, 1 hour after the exercise. This result may be valuable in future physiotherapy studies on the relationship between exercise and cancer.

  2. Design, characterization and anti-tumour cytotoxicity of a panel of recombinant, mammalian ribonuclease-based immunotoxins.

    PubMed Central

    Deonarain, M. P.; Epenetos, A. A.

    1998-01-01

    Bovine seminal ribonuclease (BSRNase) is an unusual member of the ribonuclease superfamily, because of its remarkable anti-tumour and immunosuppressive properties. We describe here the construction, expression, purification and characterization of a panel of six immunotoxins based upon this enzyme and show that we can increase its anti-tumour activity by over 2 x 10(4)-fold. This is achieved by improving tumour cell targeting using a single-chain Fv (scFv) directed against the oncofetal antigen placental alkaline phosphatase. As well as the simple scFv-BSRNase fusion protein, we have constructed five other derivatives with additional peptides designed to improve folding and intracellular trafficking and delivery. We find that the molecule most cytotoxic to antigen (PLAP)-positive cells in vitro is one that contains a C-terminal 'KDEL' endoplasmic reticulum retention signal and a peptide sequence derived from diphtheria toxin. All these molecules are produced in Escherichia coli (E. coli) as insoluble inclusion bodies and require extensive in vitro processing to recover antigen binding and ribonuclease activity. Despite incomplete ribonuclease activity and quaternary assembly, these molecules are promising reagents for specific chemotherapy of cancer and are potentially less harmful and immunogenic than current immunotoxins. Images Figure 2 PMID:9484808

  3. The Interplay of LncRNA-H19 and Its Binding Partners in Physiological Process and Gastric Carcinogenesis.

    PubMed

    Zhang, Li; Zhou, Yuhang; Huang, Tingting; Cheng, Alfred S L; Yu, Jun; Kang, Wei; To, Ka Fai

    2017-02-20

    Long non-coding RNA (lncRNA), a novel and effective modulator in carcinogenesis, has become a study hotspot in recent years. The imprinted oncofetal lncRNA H19 is one of the first identified imprinted lncRNAs with a high expression level in embryogenesis but is barely detectable in most tissues after birth. Aberrant alterations of H19 expression have been demonstrated in various tumors, including gastric cancer (GC), implicating a crucial role of H19 in cancer progression. As one of the top malignancies in the world, GC has already become a serious concern to public health with poor prognosis. The regulatory roles of H19 in gastric carcinogenesis have been explored by various research groups, which leads to the development of GC therapy. This review comprehensively summarizes the current knowledge of H19 in tumorigenesis, especially in GC pathogenesis, with emphasis on the underneath molecular mechanisms depicted from its functional partners. Furthermore, the accumulated knowledge of H19 will provide better understanding on targeted therapy of GC.

  4. Fibronectin-guided migration of carcinoma collectives

    PubMed Central

    Gopal, Sandeep; Veracini, Laurence; Grall, Dominique; Butori, Catherine; Schaub, Sébastien; Audebert, Stéphane; Camoin, Luc; Baudelet, Emilie; Radwanska, Agata; Beghelli-de la Forest Divonne, Stéphanie; Violette, Shelia M.; Weinreb, Paul H.; Rekima, Samah; Ilie, Marius; Sudaka, Anne; Hofman, Paul; Van Obberghen-Schilling, Ellen

    2017-01-01

    Functional interplay between tumour cells and their neoplastic extracellular matrix plays a decisive role in malignant progression of carcinomas. Here we provide a comprehensive data set of the human HNSCC-associated fibroblast matrisome. Although much attention has been paid to the deposit of collagen, we identify oncofetal fibronectin (FN) as a major and obligate component of the matrix assembled by stromal fibroblasts from head and neck squamous cell carcinomas (HNSCC). FN overexpression in tumours from 435 patients corresponds to an independent unfavourable prognostic indicator. We show that migration of carcinoma collectives on fibrillar FN-rich matrices is achieved through αvβ6 and α9β1 engagement, rather than α5β1. Moreover, αvβ6-driven migration occurs independently of latent TGF-β activation and Smad-dependent signalling in tumour epithelial cells. These results provide insights into the adhesion-dependent events at the tumour–stroma interface that govern the collective mode of migration adopted by carcinoma cells to invade surrounding stroma in HNSCC. PMID:28102238

  5. Expression and structural characterization of anti-T-antigen single-chain antibodies (scFvs) and analysis of their binding to T-antigen by surface plasmon resonance and NMR spectroscopy.

    PubMed

    Yuasa, Noriyuki; Koyama, Tsubasa; Subedi, Ganesh P; Yamaguchi, Yoshiki; Matsushita, Misao; Fujita-Yamaguchi, Yoko

    2013-12-01

    T-antigen (Galβ1-3GalNAcα-1-Ser/Thr), also known as Thomsen-Friedenreich antigen (TF antigen), is an oncofetal antigen commonly found in cancerous tissues. Availability of anti-T-antigen human antibodies could lead to the development of cancer diagnostics and therapeutics. Four groups of single-chain variable fragment (scFv) genes were previously isolated from a phage library (Matsumoto-Takasaki et al. (2009) Isolation and characterization of anti-T-antigen single chain antibodies from a phage library. BioSci Trends 3:87-95.). Here, four anti-T-antigen scFv genes belonging to Group 1-4 were expressed and produced in a Drosophila S2 cell expression system. ELISA and surface plasmon resonance (SPR) analyses confirmed the binding activity of 1E8 scFv protein to various T-antigen presenting conjugates. NMR experiments provided evidence of the folded nature of the 1E8 scFv protein. ScFv-ligand contact was identified by STD NMR, indicating that the galactose unit of T-antigen at the non-reducing end was primarily recognized by 1E8 scFv. This thus provides direct evidence of T-antigen specificity.

  6. Spatial-temporal protein expression of inhibitor of differentiation-1 (Id1) during fetal embryogenesis and in different mouse and human cancer types.

    PubMed

    Redrado, Miriam; Bodegas, Elena; Villaro, Ana Cristina; Nguewa, Paul A; Lopez, Ines; Gil-Bazo, Ignacio; Calvo, Alfonso

    2013-08-01

    Inhibitor of differentiation-1 (Id1) plays a role in cell proliferation, acquisition of epithelial to mesenchymal transition (EMT) features and angiogenesis. Id1 was shown to be expressed in some tumor types, mainly in advanced dedifferentiated stages. However, recent studies using a validated and highly specific monoclonal antibody against Id1 have challenged many of the results obtained by immunohistochemistry. The goal of our work was to perform a thorough analysis of Id1 expression in mouse embryos and adult tissues, as well as healthy and malignant mouse and human samples using this validated antibody (Perk et al., 2006). Our results show that Id1 was highly expressed in the oropharyngeal cavity, lung, cartilage and skin of E14 and E15 mouse embryos, but expression was progressively reduced in more developed embryos. Immunostaining only remained in epithelial cells of the gut and uterus of adult mice. Mammary MMTV-Myc and MMTV-Myc/VEGF transgenic mouse tumors, and squamous cell carcinomas of the lung induced by N-nitroso-tris-chloroethylurea (NTCU) were highly positive for Id1, unlike their respective healthy counterparts. Id1 immunostaining in a human tissue microarray (TMA) revealed strong expression in cancers of the oral cavity, bladder and cervix. Some tumor specimens of esophagus, thyroid and breast were also strongly positive. Our results suggest that Id1 is an oncofetal protein highly expressed in particular tumor types that should be reanalyzed in future studies using large cohorts of patients to reassess its diagnostic/prognostic value. Moreover, MMTV-Myc- and NTCU-induced tumors could serve as appropriate mouse models to study Id1 functions in breast and lung cancer, respectively.

  7. Proper exercise decreases plasma carcinoembryonic antigen levels with the improvement of body condition in elderly women.

    PubMed

    Ko, Il-Gyu; Park, Eung-Mi; Choi, Hye-Jung; Yoo, Jaehyun; Lee, Jong-Kyun; Jee, Yong-Seok

    2014-05-01

    Aging increases the risk of chronic diseases including cancers. Physical exercise has the beneficial effects for the elderly susceptible to the development of cancers, through maintaining a healthy body condition and improving the immune system. However, excessive or insufficient exercise might increase the risk for cancer. In the present study, we investigated what exercise frequency improves cancer-related biomarkers, such as carcinoembryonic antigen (CEA), alpha fetoprotein (AFP), red blood cell (RBC), and white blood cell (WBC), and the body composition of elderly women. Fifty-four females, aged 70 to 77 years, were divided into 4 groups: control, 1-day exercise (1E), 2-3-day exercise (2-3E), and 5-day exercise (5E) groups. The control group did not participate in any physical activity, while the subjects in the exercise groups underwent the exercise program for 12 weeks. As results, CEA was significantly decreased in the exercise groups, with the lowest values in 2-3E group. In contrast, AFP, RBC and WBC were not significantly changed. CEA is an oncofetal glycoprotein that is overexpressed in adenocarcinomas. Although the function of CEA has not been fully understood, CEA has been suggested to be involved in the release of pro-inflammatory cytokines via stimulating monocytes and macrophages. Moreover, body weight and body mass index were improved in the exercise groups, with the lowest levels in 5E group. Thus, we suggest that exercise for 2-3 days per week decreases the expression of CEA and improves body condition, without loading fatigue or stress, which may contribute to preventing cancer in the elderly women.

  8. Distribution of [sup 65]Zn labeled alpha-fetoprotein during proliferation of the BW7756 murine hepatoma

    SciTech Connect

    Keenan, J.F.

    1990-01-01

    The radiolabeling of alpha-fetoprotein (AFP) with [sup 65]Zn for the determination of its biodistribution was studied in mice bearing the BW7756 murine hepatoma as compared to that found with normal mice. AFP is an oncofetal protein of about 70,000 daltons associated with pregnancy and certain cancers (e.g., hepatoma). The AFP was purified from mouse amniotic fluid (MAF) using polyacrylamide gel electrophoresis (PAGE) and higher performance liquid chromatography (HPLC). The biological activity of AFP was maintained through the separation procedures and the purity was determined using double immunodiffusion (DID), immunoelectrophoresis (IEP) and sodium dodecyl sulfate electrophoresis (SDS). The labeling procedures included removal of intrinsic metal with EDTA, incubation with radiotracer ([sup 65]Zn) and buffer, followed by removal of unbound [sup 65]Zn using gel filtration chromatography. The results correlated well with Zn fluctuations recorded by other techniques (RIXRF, radiotracer [sup 65]Zn). Large amounts of [sup 65]Zn-AFP were localized in the liver, spleen and tumor with significant elevations above normal in the log growth phase (day 14-18). [sup 65]Zn-AFP levels in the skin, pancreas, brain and thyroid decreased as the tumor mass increased. Tumor [sup 65]Zn-AFP uptake increased with time but leveled off in the late log phase (day 21) due to tumor necrosis. In light of the results of this investigation, and previous work stating that AFP binds Zn with a higher affinity than does albumin, it is suggestive that the Zn fluctuations observed in the earlier hepatoma studies were due to the in vivo binding of Zn to AFP. These results confirm the thesis that intrinsic labeling (replacement of naturally bound ligands with radioactive analogs) does not alter the biochemical integrity as non-intrinsic labeling (e.g., Iodine) may.

  9. Breast Cancer Cells in Three-dimensional Culture Display an Enhanced Radioresponse after Coordinate Targeting of Integrin ?5?1 and Fibronectin

    SciTech Connect

    Nam, Jin-Min; Onodera, Yasuhito; Bissell, Mina J; Park, Catherine C

    2010-04-07

    Tactics to selectively enhance cancer radioresponse are of great interest. Cancer cells actively elaborate and remodel their extracellular matrix (ECM) to aid in survival and progression. Previous work has shown that {beta}1-integrin inhibitory antibodies can enhance the growth-inhibitory and apoptotic responses of human breast cancer cell lines to ionizing radiation, either when cells are cultured in three-dimensional laminin-rich ECM (3D lrECM) or grown as xenografts in mice. Here, we show that a specific {alpha} heterodimer of {beta}1-integrin preferentially mediates a prosurvival signal in human breast cancer cells that can be specifically targeted for therapy. 3D lrECM culture conditions were used to compare {alpha}-integrin heterodimer expression in malignant and nonmalignant cell lines. Under these conditions, we found that expression of {alpha}5{beta}1-integrin was upregulated in malignant cells compared with nonmalignant breast cells. Similarly, we found that normal and oncofetal splice variants of fibronectin, the primary ECM ligand of {alpha}5{beta}1-integrin, were also strikingly upregulated in malignant cell lines compared with nonmalignant acini. Cell treatment with a peptide that disrupts the interactions of {alpha}5{beta}1-integrin with fibronectin promoted apoptosis in malignant cells and further heightened the apoptotic effects of radiation. In support of these results, an analysis of gene expression array data from breast cancer patients revealed an association of high levels of {alpha}5-integrin expression with decreased survival. Our findings offer preclinical validation of fibronectin and {alpha}5{beta}1-integrin as targets for breast cancer therapy.

  10. Chemopreventive effect of Korean Angelica root extract on TRAMP carcinogenesis and integrative "omic" profiling of affected neuroendocrine carcinomas.

    PubMed

    Zhang, Jinhui; Wang, Lei; Zhang, Yong; Li, Li; Tang, Suni; Xing, Chengguo; Kim, Sung-Hoon; Jiang, Cheng; Lü, Junxuan

    2015-12-01

    Angelica gigas Nakai (AGN) root ethanol extract exerts anti-cancer activity in several allograft and xenograft models. Here we examined its chemopreventive efficacy through gavage administration against primary carcinogenesis in the transgenic adenocarcinoma of mouse prostate (TRAMP) model. Male C57BL/6 TRAMP mice and wild type littermates were given a daily gavage (5 mg/mouse, Monday-Friday) of AGN or vehicle, beginning at 8 wk of age (WOA). All mice were terminated at 24 WOA, unless earlier euthanasia was necessitated by large tumors. Whereas AGN-treated TRAMP mice decreased dorsolateral prostate lesion growth by 30% (P = 0.009), they developed fewer and smaller neuroendocrine-carcinomas (NE-Ca) (0.12 g/mouse) than vehicle-treated counterparts (0.81 g/mouse, P = 0.037). We analyzed the proteome and transcriptome of banked NE-Ca to gain molecular insights. Angiogenesis-antibody array detected a substantial reduction in AGN-treated NE-Ca of basic fibroblast growth factor (FGF2), an angiogenesis stimulator. iTRAQ proteomics plus data mining suggested changes of genes upstream and downstream of FGF2 functionally consistent with AGN inhibiting FGF2/FGFR1 signaling at different levels of the transduction cascade. Moreover, AGN upregulated mRNA of genes related to immune responses, restored expression of many tumor suppressor genes, and prostate function and muscle differentiation genes. On the other hand, AGN down-regulated mRNA of genes related to neuron signaling, oncofetal antigens, inflammation, and mast cells, Wnt signaling, embryonic morphogenesis, biosynthesis, cell adhesion, motility, invasion, and angiogenesis. These changes suggest not only multiple cancer cell targeting actions of AGN but also impact on the tumor microenvironments such as angiogenesis, inflammation, and immune surveillance.

  11. Pharmacokinetics, biodistribution and antitumour effects of Sclerotium rolfsii lectin in mice.

    PubMed

    Anupama, S; Laha, Preeti; Sharma, Mamta; Pathak, Kamal; Bane, Sanjay; Ingle, Arvind D; Gota, Vikram; Kalraiya, Rajiv D; Yu, Lu-Gang; Rhodes, Jonathan M; Swamy, Bale M; Inamdar, Shashikala R

    2017-04-03

    Sclerotium rolfsii lectin (SRL) is a lectin isolated from the fungus Sclerotium rolfsii and has exquisite binding specificity towards the oncofetal Thomsen-Friedenreich antigen (TF-Ag; Galβ1-3GalNAcα-O-Ser/Thr) and its derivatives. Previous studies have shown that SRL inhibits the proliferation of human colon, breast and ovarian cancer cells in vitro and suppresses tumour growth in mice when introduced intratumourally. The present study assessed the effect of SRL on tumour growth when introduced intraperitoneally in BALB/c nude mice and investigated the pharmacokinetics and biodistribution of SRL in Swiss albino mice. When 9 doses of SRL (30 mg/kg body weight/mice) was administered to BALB/c nude mice bearing human colon cancer HT-29 xenografts, a substantial reduction in tumour size was observed. A 35.8% reduction in tumour size was noted in the treated animals after 17 days. SRL treatment also inhibited angiogenesis, and the tumours from the treated animals were observed to carry fewer blood vessels and express less angiogenesis marker protein CD31, than that from the control animals. Pharmacokinetics and biodistribution analysis revealed that SRL was detected in the serum after 1 h and its level peaked after 24 h. SRL was not detected in any of the organs apart from the kidney where a trace amount was detected after 24 h of SRL injection. No significant changes were observed in any of the biochemical parameters tested including SGOT, SGPT, LDH, CREAT and BUN in the SRL-treated mice compared to these levels in the controls. This suggests that SRL has good potential to be developed as a therapeutic agent for cancer treatment and warrant further investigations in vivo and subsequent clinical trials.

  12. Chemopreventive effect of Korean Angelica root extract on TRAMP carcinogenesis and integrative “omic” profiling of affected neuroendocrine carcinomas

    PubMed Central

    Zhang, Jinhui; Wang, Lei; Zhang, Yong; Li, Li; Tang, Suni; Xing, Chengguo; Kim, Sung-Hoon; Jiang, Cheng; Lü, Junxuan

    2016-01-01

    Angelica gigas Nakai (AGN) root ethanol extract exerts anti-cancer activity in several allograft and xenograft models. Here we examined its chemopreventive efficacy through gavage administration against primary carcinogenesis in the transgenic adenocarcinoma of mouse prostate (TRAMP) model. Male C57BL/6 TRAMP mice and wild type littermates were given a daily gavage (5 mg/mouse, Monday-Friday) of AGN or vehicle, beginning at 8 weeks of age (WOA). All mice were terminated at 24 WOA, unless earlier euthanasia was necessitated by large tumors. Whereas AGN-treated TRAMP mice decreased dorsolateral prostate lesion growth by 30% (P = 0.009), they developed fewer and smaller neuroendocrine-carcinomas (NE-Ca) (0.12 g/mouse) than vehicle-treated counterparts (0.81g/mouse, P = 0.037). We analyzed the proteome and transcriptome of banked NE-Ca to gain molecular insights. Angiogenesis-antibody array detected a substantial reduction in AGN-treated NE-Ca of basic fibroblast growth factor (FGF2), an angiogenesis stimulator. iTRAQ proteomics plus data mining suggested changes of genes upstream and downstream of FGF2 functionally consistent with AGN inhibiting FGF2/FGFR1 signaling at different levels of the transduction cascade. Moreover, AGN upregulated mRNA of genes related to immune responses, restored expression of many tumor suppressor genes, and prostate function and muscle differentiation genes. On the other hand, AGN down-regulated mRNA of genes related to neuron signaling, oncofetal antigens, inflammation and mast cells, Wnt signaling, embryonic morphogenesis, biosynthesis, cell adhesion, motility, invasion and angiogenesis. These changes suggest not only multiple cancer cell targeting actions of AGN but also impact on the tumor microenvironments such as angiogenesis, inflammation and immune surveillance. PMID:25307620

  13. Molecular mechanism of anticancer effect of Sclerotium rolfsii lectin in HT29 cells involves differential expression of genes associated with multiple signaling pathways: A microarray analysis.

    PubMed

    Barkeer, Srikanth; Guha, Nilanjan; Hothpet, Vishwanathreddy; Saligrama Adavigowda, Deepak; Hegde, Prajna; Padmanaban, Arunkumar; Yu, Lu-Gang; Swamy, Bale M; Inamdar, Shashikala R

    2015-12-01

    Sclerotium rolfsii lectin (SRL) is a lectin isolated from fungus S. rolfsii and has high binding specificity toward the oncofetal Thomsen-Friedenreich carbohydrate antigen (Galβ1-3GalNAc-α-O-Ser/Thr, T or TF), which is expressed in more than 90% of human cancers. Our previous studies have shown that binding of SRL to human colon, breast and ovarian cancer cells induces cell apoptosis in vitro and suppresses tumor growth in vivo. This study investigated the SRL-mediated cell signaling in human colon cancer HT29 cells by mRNA and miRNA microarrays. It was found that SRL treatment results in altered expression of several hundred molecules including mitogen-activated protein kinase (MAPK) and c-JUN-associated, apoptosis-associated and cell cycle and DNA replication-associated signaling molecules. Pathway analysis using GeneSpring 12.6.1 revealed that SRL treatment induces changes of MAPK and c-JUN-associated signaling pathways as early as 2 h while changes of cell cycle, DNA replication and apoptosis pathways were significantly affected only after 24 h. A significant change of cell miRNA expression was also observed after 12 h treatment of the cells with SRL. These changes were further validated by quantitative real time polymerase chain reaction and immunoblotting. This study thus suggests that the presence of SRL affects multiple signaling pathways in cancer cells with early effects on cell proliferation pathways associated with MAPK and c-JUN, followed by miRNA-associated cell activity and apoptosis. This provides insight information into the molecular mechanism of the anticancer activity of this fungal lectin.

  14. Diagnostic value of immunohistochemical IMP3 expression in core needle biopsies of pancreatic ductal adenocarcinoma.

    PubMed

    Wachter, David Lukas; Schlabrakowski, Anne; Hoegel, Josef; Kristiansen, Glen; Hartmann, Arndt; Riener, Marc-Oliver

    2011-06-01

    The oncofetal protein, insulin-like growth factor II messenger ribonucleic acid-binding protein 3 (IMP3), has been analyzed in many different tumors. Various studies have found that IMP3 is a marker for malignancy and is correlated with increased tumor aggressiveness and reduced overall survival. The diagnosis of pancreatic ductal adenocarcinoma (PDAC) in core needle biopsies can be challenging, and immunohistochemical markers are needed. We studied IMP3 expression in 177 core needle biopsies of the pancreas, including 112 PDACs, 55 cases with chronic sclerosing pancreatitis, and 10 biopsies with tumor-free pancreatic tissue without inflammation. An additional 18 biopsies of PDAC metastases (16 liver biopsies and 2 lymph node biopsies) were analyzed. To study IMP3 expression in large tissue sections, 45 pancreatic resection specimens (26 with PDAC and 19 with chronic sclerosing pancreatitis) were investigated. In contrast to normal or inflamed pancreatic tissue, which was negative in 47 of 65 (72.3%) cases and weakly positive in 15 of 65 (23.1%) cases, strong IMP3 expression was found in 99 of 112 (88.4%) PDACs. Therefore, sensitivity and specificity of IMP3 expression in the differential diagnosis of PDAC and chronic sclerosing pancreatitis using core needle biopsies were found to be 88.4% and 94.6%, respectively. These results were confirmed in the pancreas resection specimens. Furthermore, strong IMP3 expression was found in 17 of 18 (94.4%) of the PDAC metastases that were analyzed. Our study shows that IMP3 is an easy to use and potentially new immunohistochemical marker for the diagnosis of PDAC in core needle biopsies.

  15. Evidence for a human-specific mechanism for diet and antibody-mediated inflammation in carcinoma progression

    PubMed Central

    Hedlund, Maria; Padler-Karavani, Vered; Varki, Nissi M.; Varki, Ajit

    2008-01-01

    Patients with cancer have circulating heterophile antibodies that agglutinate animal red cells via recognition of the mammalian cell surface sialic acid N-glycolylneuraminic acid (Neu5Gc), which was long considered an oncofetal antigen in humans. However, humans are genetically deficient in Neu5Gc production and instead metabolically accumulate Neu5Gc from dietary sources, particularly red meats and milk products. Moreover, mice with a human-like defect showed no alternate pathway for Neu5Gc synthesis and even normal humans express anti-Neu5Gc antibodies. We show here that human tumors accumulate Neu5Gc that is covalently attached to multiple classes of glycans. The paradox of human tumor Neu5Gc accumulation in the face of circulating anti-Neu5Gc antibodies was hypothesized to be due to facilitation of tumor progression by the resulting low-grade chronic inflammation. Indeed, murine tumors expressing human-like levels of Neu5Gc show accelerated growth in syngeneic mice with a human-like Neu5Gc deficiency, coincident with the induction of anti-Neu5Gc antibodies and increased infiltration of inflammatory cells. Transfer of polyclonal monospecific syngeneic mouse anti-Neu5Gc serum also enhanced growth of transplanted syngeneic tumors bearing human-like levels of Neu5Gc, with tumors showing evidence for antibody deposition, enhanced angiogenesis and chronic inflammation. These effects were suppressed by a cyclooxygenase-2 inhibitor, a drug type known to reduce human carcinoma risk. Finally, affinity-purified human anti-Neu5Gc antibodies also accelerate growth of Neu5Gc-containing tumors in Neu5Gc-deficient mice. Taken together, the data suggest that the human propensity to develop diet-related carcinomas is contributed to by local chronic inflammation, resulting from interaction of metabolically-accumulated dietary Neu5Gc with circulating anti-Neu5Gc antibodies. PMID:19017806

  16. Efficient Preparation of Site-Specific Antibody-Drug Conjugates Using Cysteine Insertion.

    PubMed

    Dimasi, Nazzareno; Fleming, Ryan; Zhong, Haihong; Bezabeh, Binyam; Kinneer, Krista; Christie, Ronald J; Fazenbaker, Christine; Wu, Herren; Gao, Changshou

    2017-05-01

    Antibody-drug conjugates (ADCs) are a class of biopharmaceuticals that combine the specificity of antibodies with the high-potency of cytotoxic drugs. Engineering cysteine residues in the antibodies using mutagenesis is a common method to prepare site-specific ADCs. With this approach, solvent accessible amino acids in the antibody have been selected for substitution with cysteine for conjugating maleimide-bearing cytotoxic drugs, resulting in homogeneous and stable site-specific ADCs. Here we describe a cysteine engineering approach based on the insertion of cysteines before and after selected sites in the antibody, which can be used for site-specific preparation of ADCs. Cysteine-inserted antibodies have expression level and monomeric content similar to the native antibodies. Conjugation to a pyrrolobenzodiazepine dimer (SG3249) resulted in comparable efficiency of site-specific conjugation between cysteine-inserted and cysteine-substituted antibodies. Cysteine-inserted ADCs were shown to have biophysical properties, FcRn, and antigen binding affinity similar to the cysteine-substituted ADCs. These ADCs were comparable for serum stability to the ADCs prepared using cysteine-mutagenesis and had selective and potent cytotoxicity against human prostate cancer cells. Two of the cysteine-inserted variants abolish binding of the resulting ADCs to FcγRs in vitro, thereby potentially preventing non-target mediated uptake of the ADCs by cells of the innate immune system that express FcγRs, which may result in mitigating off-target toxicities. A selected cysteine-inserted ADC demonstrated potent dose-dependent anti-tumor activity in a xenograph tumor mouse model of human breast adenocarcinoma expressing the oncofetal antigen 5T4.

  17. Expression of different carbohydrate tumour markers and galectins 1 and 3 in normal squamous and malignant epithelia of the upper aaerodigestive tract.

    PubMed

    Wiest, Irmi; Alexiou, Christoph; Kuhn, Christina; Schulze, Sandra; Kunze, Susanne; Mayr, Doris; Betz, Peter; Jeschke, Udo; Dian, Darius

    2012-05-01

    Tumour markers hold a great relevance in the diagnosis and the follow-up treatment of different kinds of human carcinoma. Although head and neck cancer occurs frequently, there is still lack of appropriate tumour markers. Our investigation on the expression of sialyl Lewis A (CA19-9) in laryngeal carcinomas, consists of systematical analysis of oncofetal carbohydrates and of galectins 1 and 3 in different normal and malignant tissues of the aerodigestive tract. Paraffin-embedded sections of normal tongue, vocal cord, larynx, pharynx and epiglottis, representing normal control tissue and laryngeal cancer tissue were incubated with monoclonal antibodies against sialyl Lewis A and X (sLeA and X), Lewis Y (LeY), the Thomsen-Friedenreich (TF) antigen and galectin 1 and 3 (Gal-1 and -3). A staining reaction was carried out with ABC-peroxidase and diaminobenzidine (DAB). Tissue of breast cancer was used as a positive control. Mouse IgM, as isotype control antibody, was used as a negative control. Semi quantitative evaluation was carried out double-blinded, by two independent investigators, including a pathologist. Squamous epithelia of all investigated normal tissues of the aerodigestive tract show nearly the same pattern. Most impressive findings are the very weak expression of Gal-1 and the total absence of the TF antigen. Laryngeal cancer reveals high amounts of sLeA, Gal-1 and the TF antigen. On the basis of our findings in normal tissue of the aeradigestive tract, these three markers qualified as potential tumour markers for carcinoma of the aerodigestive tract. In particular, the high expression of TF in cancer tissue and its absence from the normal tissue is promising for its establishment as a new tumour marker in this field.

  18. Hepatic miR-29ab1 expression modulates chronic hepatic injury

    PubMed Central

    Kogure, Takayuki; Costinean, Stefan; Yan, Irene; Braconi, Chiara; Croce, Carlo; Patel, Tushar

    2012-01-01

    MicroRNAs (miRNAs) are small, regulatory non-coding RNAs that have potent effects on gene expression. Several miRNA are deregulated in cellular processes involved in human liver diseases and regulation of cellular processes. Recent studies have identified the involvement of miR-29 in hepatic fibrosis and carcinogenesis. Although several targets of miR-29 have been identified, there is limited information regarding the cell-type specific roles of miR-29 in the liver, and we sought to evaluate the role of this miRNA in hepatic pathobiology. We report the generation of a tissue–specific knockout mouse to evaluate the role of miR-29 in hepatic fibrosis and carcinogenesis in response to injury. We hypothesized that miR-29 contributes to the hepatocyte driven response to chronic cellular injury that results in fibrosis. In support of this hypothesis, fibrosis and mortality were enhanced in miR29 knockout mice in response to carbon tetrachloride. Genome-wide gene expression analysis identified an over-representation of genes associated with fibrosis. The oncofetal RNA H19 was modulated in a miR-29 dependent manner following exposure to carbon tetrachloride in vivo. The impact of a hepatocyte specific miR-29 knockout on survival following chronic hepatic injury in vivo implicates this miRNA as a potential target for intervention. These results provide evidence of the involvement of miR-29 in chronic hepatic injury, and suggest a role for deregulated hepatocyte expression of miR-29 in the response to hepatic injury, fibrosis and carcinogenesis. PMID:22469499

  19. Mouse models expressing human carcinoembryonic antigen (CEA) as a transgene: Evaluation of CEA-based cancer vaccines

    PubMed Central

    Hance, Kenneth W.; Zeytin, Hasan E.; Greiner, John W.

    2010-01-01

    In recent years, investigators have carried out several studies designed to evaluate whether human tumor-associated antigens might be exploited as targets for active specific immunotherapy, specifically human cancer vaccines. Not too long ago such an approach would have been met with considerable skepticism because the immune system was believed to be a rigid discriminator between self and non-self which, in turn, protected the host from a variety of pathogens. That viewpoint has been challenged in recent years by a series of studies indicating that antigenic determinants of self have not induced absolute host immune tolerance. Moreover, under specific conditions that evoke danger signals, peptides from self-antigen can be processed by the antigen-presenting cellular machinery, loaded onto the major histocompatibility antigen groove to serve as targets for immune intervention. Those findings provide the rationale to investigate a wide range of tumor-associated antigens, including differentiation antigens, oncogenes, and tumor suppressor genes as possible immune-based targets. One of those tumor-associated antigens is the carcinoembryonic antigen (CEA). Described almost 40 years ago, CEA is a Mr 180–200,000 oncofetal antigen that is one of the more widely studied human tumor-associated antigens. This review will provide: (i) a brief overview of the CEA gene family, (ii) a summary of early preclinical findings on overcoming immune tolerance to CEA, and (iii) the rationale to develop mouse models which spontaneously develop gastrointestinal tumors and express the CEA transgene. Those models have been used extensively in the study of overcoming host immune tolerance to CEA, a self, tumor-associated antigen, and the experimental findings have served as the rationale for the design of early clinical trials to evaluate CEA-based cancer vaccines. PMID:15888344

  20. IMP3 Predicts Invasion and Prognosis in Human Lung Adenocarcinoma.

    PubMed

    Yan, Jinhai; Wei, Qingzhu; Jian, Wenjing; Qiu, Bo; Wen, Jing; Liu, Jianghuan; Fu, Bo; Zhou, Xinhua; Zhao, Tong

    2016-02-01

    Insulin-like growth factor II mRNA-binding protein 3 (IMP3) is an oncofetal protein associated with several aggressive and advanced cancers. Whether IMP3 can predict invasion, and prognosis in patients with human lung adenocarcinoma (LAC) remains unclear. Ninety-five LAC and 75 non-tumor lung tissue samples were included in a tissue microarray. IMP3 expression was assessed by immunohistochemical examination. Correlation between IMP3 expression levels, clinicopathological characteristics, and overall prognosis was evaluated. In a separate in vitro study, RNA interference method was applied for knockdown of IMP3 gene in human LAC cell lines. Invasive potential of LAC cells was then evaluated by transwell migration assay. IMP3 immunoreactivity was observed in 39 out of 95 (41.1 %) LAC patients, but not in non-tumor lung tissues. IMP3 expression levels were closely associated with histological grade (P = 0.037), TNM stage (P = 0.034), and lymph node metastasis (P = 0.011). Patients presenting with positive IMP3 expression (P = 0.000), an advanced TNM stage (P = 0.000), and lymph node metastasis (P = 0.001) had a worse overall survival, compared to those lacking these characteristics. Both IMP3 expression (hazard ratio [HR], 2.310; 95 % confidence interval [CI] 1.192-4.476; P = 0.013) and TNM stage (HR 2.338; 95 % CI 1.393-3.925; P = 0.001) were independent predictors of poor prognosis. The invasive potential of LAC cells was significantly inhibited by IMP3 knockdown. IMP3 appears to play an important role in tumor invasion in patients with LAC and may serve as a useful prognostic biomarker in these patients.

  1. Purification and refolding of anti-T-antigen single chain antibodies (scFvs) expressed in Escherichia coli as inclusion bodies.

    PubMed

    Yuasa, Noriyuki; Koyama, Tsubasa; Fujita-Yamaguchi, Yoko

    2014-02-01

    T-antigen (Galβ1-3GalNAcα-1-Ser/Thr) is an oncofetal antigen that is commonly expressed as a carbohydrate determinant in many adenocarcinomas. Since it is associated with tumor progression and metastasis, production of recombinant antibodies specific for T-antigen could lead to the development of cancer diagnostics and therapeutics. Previously, we isolated and characterized 11 anti-T-antigen phage clones from a phage library displaying human single-chain antibodies (scFvs) and purified one scFv protein, 1G11. More recently, we purified and characterized 1E8 scFv protein using a Drosophila S2 expression system. In the current study, four anti-T-antigen scFv genes belonging to Groups 1-4 were purified from inclusion bodies expressed in Escherichia coli cells. Inclusion bodies isolated from E. coli cells were denatured in 3.5 M Gdn-HCl. Solubilized His-tagged scFv proteins were purified using Ni(2+)-Sepharose column chromatography in the presence of 3.5 M Gdn-HCl. Purified scFv proteins were refolded according to a previously published method of step-wise dialysis. Two anti-T-antigen scFv proteins, 1E6 and 1E8 that belong to Groups 1 and 2, respectively, were produced in sufficient amounts, thus allowing further characterization of their binding activity with T-antigen. Specificity and affinity constants determined using enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR), respectively, provided evidence that both 1E8 and 1E6 scFv proteins are T-antigen specific and suggested that 1E8 scFv protein has a higher affinity for T-antigen than 1E6 scFv protein.

  2. Biosynthesis and turnover of O-acetyl and N-acetyl groups in the gangliosides of human melanoma cells

    SciTech Connect

    Manzi, A.E.; Sjoberg, E.R.; Diaz, S.; Varki, A.

    1990-08-05

    We and others previously described the melanoma-associated oncofetal glycosphingolipid antigen 9-O-acetyl-GD3, a disialoganglioside O-acetylated at the 9-position of the outer sialic acid residue. We have now developed methods to examine the biosynthesis and turnover of disialogangliosides in cultured melanoma cells and in Golgi-enriched vesicles from these cells. O-Acetylation was selectively expressed on di- and trisialogangliosides, but not on monosialogangliosides, nor on glycoprotein-bound sialic acids. Double-labeling of cells with (3H)acetate and (14C)glucosamine introduced easily detectable labels into each of the components of the ganglioside molecules. Pulse-chase studies of such doubly labeled molecules indicated that the O-acetyl groups turn over faster than the parent molecule. When Golgi-enriched vesicles from these cells were incubated with (acetyl-3H)acetyl-coenzyme A, the major labeled products were disialogangliosides. (Acetyl-3H)O-acetyl groups were found at both the 7- and the 9-positions, indicating that both 7-O-acetyl GD3 and 9-O-acetyl GD3 were synthesized by the action of O-acetyltransferase(s) on endogenous GD3. Analysis of the metabolically labeled molecules confirmed the existence of both 7- and 9-O-acetylated GD3 in the intact cells. Surprisingly, the major 3H-labeled product of the in vitro labeling reaction was not O-acetyl-GD3, but GD3, with the label exclusively in the sialic acid residues. Fragmentation of the labeled sialic acids by enzymatic and chemical methods showed that the 3H-label was exclusively in (3H)N-acetyl groups. Analyses of the double-labeled sialic acids from intact cells also showed that the 3H-label from (3H)acetate was exclusively in the form of (3H)N-acetyl groups, whereas the 14C-label was at the 4-position.

  3. mRNA expression and protein distribution of fibronectin splice variants and high-molecular weight tenascin-C in different phases of human fracture healing.

    PubMed

    Kilian, O; Dahse, R; Alt, V; Zardi, L; Hentschel, J; Schnettler, R; Kosmehl, H

    2008-08-01

    Fracture healing is a reparative physiological process, which proceeds in stages, each characterized by the predominant tissue in the fracture gap. The tissue matrix is continuously reorganized by cell migration, proliferation, and differentiation. Adhesive proteins such as fibronectin and tenascin transmit information between matrix and cells. As a result of alternative splicing of pre-RNA, EDA + fibronectin, EDB + fibronectin, and high-molecular weight (hm) tenascin-C are generated. By definition, EDB + fibronectin is an oncofetal protein because it is extremely rare in normal adult tissue and plasma, whereas it is expressed in fetal and tumor tissues and during wound healing. In this study, we for the first time describe EDA + fibronectin, EDB + fibronectin, and hm tenascin-C expression in human fracture gap tissue during various stages of differentiation. We demonstrate mRNA expression of all three splice variants in the initial fibrin matrix with upregulation in the enchondral ossification/osteoid and woven bone stages. Of all variants, EDA + fibronectin mRNA has the highest concentration in all stages. For the analysis, we used LightCycler-based relative mRNA quantification and immunohistochemistry. Our data demonstrate that EDA + fibronectin and hm tenascin-C show a diffuse distribution pattern in fracture gap connective tissue, while EDB + fibronectin is focally concentrated in osteoblastic cells at the margins of woven bone. EDA + fibronectin and hm tenascin represent markers for active granulation processes, whereas EDB + fibronectin is specific for cells forming the enchondral and osteoid matrix. The possibility of stimulating fracture healing by EDB + fibronectin-cytokine complexes should be tested in further investigations.

  4. Enrichment of Human Stem-Like Prostate Cells with s-SHIP Promoter Activity Uncovers a Role in Stemness for the Long Noncoding RNA H19.

    PubMed

    Bauderlique-Le Roy, Hélène; Vennin, Constance; Brocqueville, Guillaume; Spruyt, Nathalie; Adriaenssens, Eric; Bourette, Roland P

    2015-05-15

    Understanding normal and cancer stem cells should provide insights into the origin of prostate cancer and their mechanisms of resistance to current treatment strategies. In this study, we isolated and characterized stem-like cells present in the immortalized human prostate cell line, RWPE-1. We used a reporter system with green fluorescent protein (GFP) driven by the promoter of s-SHIP (for stem-SH2-domain-containing 5'-inositol phosphatase) whose stem cell-specific expression has been previously shown. We observed that s-SHIP-GFP-expressing RWPE-1 cells showed stem cell characteristics such as increased expression of stem cell surface markers (CD44, CD166, TROP2) and pluripotency transcription factors (Oct4, Sox2), and enhanced sphere-forming capacity and resistance to arsenite-induced cell death. Concomitant increased expression of the long noncoding RNA H19 was observed, which prompted us to investigate a putative role in stemness for this oncofetal gene. Targeted suppression of H19 with siRNA decreased Oct4 and Sox2 gene expression and colony-forming potential in RWPE-1 cells. Conversely, overexpression of H19 significantly increased gene expression of these two transcription factors and the sphere-forming capacity of RWPE-1 cells. Analysis of H19 expression in various prostate and mammary human cell lines revealed similarities with Sox2 expression, suggesting that a functional relationship may exist between H19 and Sox2. Collectively, we provide the first evidence that s-SHIP-GFP promoter reporter offers a unique marker for the enrichment of human stem-like cell populations and highlight a role in stemness for the long noncoding RNA H19.

  5. IMP-1 displays cross-talk with K-Ras and modulates colon cancer cell survival through the novel proapoptotic protein CYFIP2.

    PubMed

    Mongroo, Perry S; Noubissi, Felicite K; Cuatrecasas, Miriam; Kalabis, Jiri; King, Catrina E; Johnstone, Cameron N; Bowser, Mark J; Castells, Antoni; Spiegelman, Vladimir S; Rustgi, Anil K

    2011-03-15

    Insulin-like growth factor 2 mRNA-binding protein-1 (IMP-1) is an oncofetal protein that binds directly to and stabilizes oncogenic c-Myc and regulates, in turn, its posttranscriptional expression and translation. In contrast to normal adult tissue, IMP-1 is reexpressed and/or overexpressed in human cancers. We show that knockdown of c-Myc in human colon cancer cell lines increases the expression of mature let-7 miRNA family members and downregulates several of its mRNA targets: IMP-1, Cdc34, and K-Ras. We further show that loss of IMP-1 inhibits Cdc34, Lin-28B, and K-Ras, suppresses SW-480 cell proliferation and anchorage-independent growth, and promotes caspase- and lamin-mediated cell death. We also found that IMP-1 binds to the coding region and 3'UTR of K-Ras mRNA. RNA microarray profiling and validation by reverse transcription PCR reveals that the p53-inducible proapoptotic protein CYFIP2 is upregulated in IMP-1 knockdown SW480 cells, a novel finding. We also show that overexpression of IMP-1 increases c-Myc and K-Ras expression and LIM2405 cell proliferation. Furthermore, we show that loss of IMP-1 induces Caspase-3- and PARP-mediated apoptosis, and inhibits K-Ras expression in SW480 cells, which is rescued by CYFIP2 knockdown. Importantly, analysis of 228 patients with colon cancers reveals that IMP-1 is significantly upregulated in differentiated colon tumors (P ≤ 0.0001) and correlates with K-Ras expression (r = 0.35, P ≤ 0.0001) relative to adjacent normal mucosa. These findings indicate that IMP-1, interrelated with c-Myc, acts upstream of K-Ras to promote survival through a novel mechanism that may be important in colon cancer pathogenesis. ©2011 AACR.

  6. RNA-binding Protein Insulin-like Growth Factor mRNA-binding Protein 3 (IMP-3) Promotes Cell Survival via Insulin-like Growth Factor II Signaling after Ionizing Radiation*

    PubMed Central

    Liao, Baisong; Hu, Yan; Brewer, Gary

    2011-01-01

    Ionizing radiation (IR) induces proapoptotic gene expression programs that inhibit cell survival. These programs often involve RNA-binding proteins that associate with their mRNA targets to elicit changes in mRNA stability and/or translation. The RNA-binding protein IMP-3 is an oncofetal protein overexpressed in many human malignancies. IMP-3 abundance correlates with tumor aggressiveness and poor prognosis. As such, IMP-3 is proving to be a highly significant biomarker in surgical pathology. Among its many mRNA targets, IMP-3 binds to and promotes translation of insulin-like growth factor II (IGFII) mRNA. Our earlier studies showed that reducing IMP-3 abundance with siRNAs reduced proliferation of human K562 chronic myeloid leukemia cells because of reduced IGF-II biosynthesis. However, the role of IMP-3 in apoptosis is unknown. Here, we have used IR-induced apoptosis of K562 cells as a model to explore a role for IMP-3 in cell survival. Knockdown of IMP-3 with siRNA increased susceptibility of cells to IR-induced apoptosis and led to reduced IGF-II production. Gene reporter assays revealed that IMP-3 acts through the 5′ UTR of IGFII mRNA during apoptosis to promote translation. Finally, culture of IR-treated cells with recombinant IGF-II partially reversed the effects of IMP-3 knockdown on IR-induced apoptosis. Together, these results indicate that IMP-3 acts in part through the IGF-II pathway to promote cell survival in response to IR. Thus, IMP-3 might serve as a new drug target to increase sensitivity of CML cells or other cancers to IR therapy. PMID:21757716

  7. RNA-binding protein insulin-like growth factor mRNA-binding protein 3 (IMP-3) promotes cell survival via insulin-like growth factor II signaling after ionizing radiation.

    PubMed

    Liao, Baisong; Hu, Yan; Brewer, Gary

    2011-09-09

    Ionizing radiation (IR) induces proapoptotic gene expression programs that inhibit cell survival. These programs often involve RNA-binding proteins that associate with their mRNA targets to elicit changes in mRNA stability and/or translation. The RNA-binding protein IMP-3 is an oncofetal protein overexpressed in many human malignancies. IMP-3 abundance correlates with tumor aggressiveness and poor prognosis. As such, IMP-3 is proving to be a highly significant biomarker in surgical pathology. Among its many mRNA targets, IMP-3 binds to and promotes translation of insulin-like growth factor II (IGFII) mRNA. Our earlier studies showed that reducing IMP-3 abundance with siRNAs reduced proliferation of human K562 chronic myeloid leukemia cells because of reduced IGF-II biosynthesis. However, the role of IMP-3 in apoptosis is unknown. Here, we have used IR-induced apoptosis of K562 cells as a model to explore a role for IMP-3 in cell survival. Knockdown of IMP-3 with siRNA increased susceptibility of cells to IR-induced apoptosis and led to reduced IGF-II production. Gene reporter assays revealed that IMP-3 acts through the 5' UTR of IGFII mRNA during apoptosis to promote translation. Finally, culture of IR-treated cells with recombinant IGF-II partially reversed the effects of IMP-3 knockdown on IR-induced apoptosis. Together, these results indicate that IMP-3 acts in part through the IGF-II pathway to promote cell survival in response to IR. Thus, IMP-3 might serve as a new drug target to increase sensitivity of CML cells or other cancers to IR therapy.

  8. IMP-3 EXPRESSION IN BENIGN MELANOCYTIC NEVI, DYSPLASTIC NEVI AND MALIGNANT MELANOMA: PRELIMINARY FINDINGS IN BULGARIAN PATIENTS.

    PubMed

    Chokoeva, A A; Ananiev, J; Wollina, U; Tana, C; Lotti, T; Cardoso, J C; Tchernev, G

    2015-01-01

    IMP-3 is generally considered as an oncofetal protein, which plays a critical role in regulation of cell proliferation via an IGF-II-dependent pathway in K562 leukemia cells. IMP-3 expression has been detected in malignancies with various origins, while its appearance in adult tissue is generally considered abnormal, with some exceptions. IMP3 is also considered a prognostic biomarker in patients with renal cell carcinoma and clear-cell type ovarian carcinoma, hepatocellular carcinoma, pancreatic ductal adenocarcinoma and in patients with poorly differentiated thyroid carcinoma and uterine cervical carcinomas, testicular cancer and malignant melanoma. To our knowledge, no more than 4 PubMed-indexed studies have investigated the expression of IMP-3 in melanocytic lesions, namely its role in the differentiation between benign and malignant neoplasms. We investigated the expression of IMP-3 in a small series of benign melanocytic lesions, dysplastic nevi and melanomas, aiming to establish its significance as a marker for their distinction, comparing the results with those from the literature. IMP- 3 immunostaining was performed in 30 melanocytic lesions: 10 malignant melanomas, 10 dysplastic nevi and 10 benign melanocytic nevi. Our results revealed expression in 20% of dysplastic lesions and 40% of melanoma cases, while none of the benign nevi showed positive expression. These data contradict some of the results from other studies and raise some questions regarding the correlation between IMP- 3 and the degree of dysplasia of melanocytic nevi, as well as its potential relationship with prognostic parameters in melanoma, including tumor thickness and mitotic rate. Our results suggest that IMP-3 expression could be only an auxiliary marker for differentiation between dysplastic nevi and benign nevi, since although it is not expressed in all dysplastic lesions, staining correlates with the degree of dysplasia/atypia. It seems that IMP-3 expression is not a useful

  9. IMP3 expression in lesions of the biliary tract: a marker for high-grade dysplasia and an independent prognostic factor in bile duct carcinomas.

    PubMed

    Riener, Marc-Oliver; Fritzsche, Florian R; Clavien, Pierre-Alain; Pestalozzi, Bernhard C; Probst-Hensch, Nicole; Jochum, Wolfram; Kristiansen, Glen

    2009-10-01

    The oncofetal protein IMP3 (insulin-like growth factor II mRNA binding protein 3) is expressed during embryogenesis and carcinogenesis. Various tumor types have been analyzed for IMP3 expression, which was exclusively found in tumor cells and correlated with increased tumor aggressiveness and reduced overall survival. To our knowledge, IMP3 expression has not been investigated in bile duct carcinomas. Using large tissue sections from resection specimens of the extrahepatic biliary tract, we analyzed IMP3 in normal bile ducts (n = 36), bile ducts with acute inflammation and reactive epithelial changes (n = 26), low-grade dysplasia (n = 9), and high-grade dysplasia (n = 11). Furthermore, IMP3 expression was assessed in bile duct carcinoma (n = 115) using clinically well-characterized tissue microarrays. The findings were correlated with clinical-pathologic parameters including survival. High-grade dysplasia was strongly positive for IMP3 in all cases studied compared with no or weak expression in normal, inflamed, and low-grade dysplastic bile ducts. Of the bile duct carcinomas 58.3% (67/115) were strongly positive for IMP3, which was associated with a higher proliferation rate (P = .004) and p53 positivity (P = .022). Patients with strong IMP3 expression had significantly reduced overall survival (P = .037) similarly to the subgroup of pT3 carcinomas (P = .007). In multivariate analysis, IMP3 expression was an independent prognostic factor for overall survival (P = .040, RR = 1.809). This comprehensive study shows that IMP3 is an independent prognostic biomarker in bile duct carcinoma. In addition, it may be a marker for high-grade dysplasia in the extrahepatic biliary tract.

  10. Migration-stimulating factor (MSF) is over-expressed in non-small cell lung cancer and promotes cell migration and invasion in A549 cells over-expressing MSF

    SciTech Connect

    Deng, Xuefeng; Ma, Qunfeng; Zhang, Bo; Jiang, Hong; Zhang, Zhipei; Wang, Yunjie

    2013-10-15

    Migration-stimulating factor (MSF), an oncofetal truncated isoform of fibronectin, is a potent stimulator of cell invasion. However, its distribution and motogenic role in non-small cell lung cancer (NSCLC) have never been identified. In this study, real-time PCR and immunohistochemical staining (IHC) were performed to detect MSF mRNA and protein levels in tumor tissues and matched adjacent tumor-free tissues. Furthermore, to examine the effect of MSF on invasiveness, MSF was upregulated in A549 cells. The invasiveness and viability of A549 cells were then determined using a transwell migration assay and the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) viability assays, respectively. The expression level of MSF in NSCLC tissue was markedly higher than in matched adjacent tumor-free tissue. Additionally, the level of MSF protein expression in stage III and IV NSCLC samples was higher than in stage I and II NSCLC samples. More importantly, we also demonstrated that migration and invasion of A549 cells increased substantially after upregulating MSF, although proliferation remained unchanged. Meanwhile, we found no correlation between increasing motility and invasiveness of MSF-overexpressing cells and expression levels and activities of matrix metalloprotease MMP-2 and MMP-9. Our current study shows that MSF plays a role in migration and invasion of A549 cells and suggests that MSF may be a potential biomarker of NSCLC progression. - Highlights: • MSF expression was upregulated in NSCLC and correlated with TNM stages. • MSF may be a new biomarker for NSCLC progression. • MSF promoted migration and invasion in A549 cells, independent of MMP-2/MMP-9 expression.

  11. Tumor-derived alpha-fetoprotein impairs the differentiation and T cell stimulatory activity of human dendritic cells

    PubMed Central

    Pardee, Angela D.; Shi, Jian; Butterfield, Lisa H.

    2014-01-01

    Several tumor-derived factors have been implicated in DC dysfunction in cancer patients. Alpha-fetoprotein (AFP) is an oncofetal antigen that is highly expressed in abnormalities of prenatal development and several epithelial cancers, including hepatocellular carcinoma (HCC). In HCC patients exhibiting high levels of serum AFP, we have observed a lower ratio of myeloid-to-plasmacytoid circulating DC compared to patients with low serum AFP levels and healthy donors. To test the effect of AFP on DC differentiation in vitro, peripheral blood monocytes from healthy donors were cultured in the presence of cord blood-derived normal AFP (nAFP) or HCC tumor-derived AFP (tAFP), and DC phenotype and function was assessed. Although the nAFP and tAFP isoforms only differ at one carbohydrate group, low (physiological) levels of tAFP, but not nAFP, significantly inhibited DC differentiation. tAFP-conditioned DC expressed diminished levels of DC maturation markers, retained a monocyte-like morphology, exhibited limited production of inflammatory mediators, and failed to induce robust T cell proliferative responses. Mechanistic studies revealed that the suppressive activity of tAFP is dependent on the presence of low molecular weight (LMW) species that i) co-purify with tAFP, and ii) function equivalently to the LMW fractions of both tumor and non-tumor cell lysates. These data reveal the unique ability of tAFP to serve as a chaperone protein for LMW molecules, both endogenous and ubiquitous in nature, which function cooperatively to impair DC differentiation and function. Therefore, novel therapeutic approaches that antagonize the regulatory properties of tAFP will be critical to enhance immunity and improve clinical outcomes. PMID:25355916

  12. Tumor-derived α-fetoprotein impairs the differentiation and T cell stimulatory activity of human dendritic cells.

    PubMed

    Pardee, Angela D; Shi, Jian; Butterfield, Lisa H

    2014-12-01

    Several tumor-derived factors have been implicated in dendritic cell (DC) dysfunction in cancer patients. α-fetoprotein (AFP) is an oncofetal Ag that is highly expressed in abnormalities of prenatal development and several epithelial cancers, including hepatocellular carcinoma (HCC). In HCC patients exhibiting high levels of serum AFP, we observed a lower ratio of myeloid/plasmacytoid circulating DCs compared with patients with low serum AFP levels and healthy donors. To test the effect of AFP on DC differentiation in vitro, peripheral blood monocytes from healthy donors were cultured in the presence of cord blood-derived normal AFP (nAFP) or HCC tumor-derived AFP (tAFP), and DC phenotype and function were assessed. Although the nAFP and tAFP isoforms only differ at one carbohydrate group, low (physiological) levels of tAFP, but not nAFP, significantly inhibited DC differentiation. tAFP-conditioned DCs expressed diminished levels of DC maturation markers, retained a monocyte-like morphology, exhibited limited production of inflammatory mediators, and failed to induce robust T cell proliferative responses. Mechanistic studies revealed that the suppressive activity of tAFP is dependent on the presence of low molecular mass (LMM) species that copurify with tAFP and function equivalently to the LMM fractions of both tumor and nontumor cell lysates. These data reveal the unique ability of tAFP to serve as a chaperone protein for LMM molecules, both endogenous and ubiquitous in nature, which function cooperatively to impair DC differentiation and function. Therefore, novel therapeutic approaches that antagonize the regulatory properties of tAFP will be critical to enhance immunity and improve clinical outcomes.

  13. A pancreatic tumor-specific biomarker characterized in humans and mice as an immunogenic onco-glycoprotein is efficient in dendritic cell vaccination

    PubMed Central

    Collignon, Aurélie; Perles-Barbacaru, Adriana Teodora; Robert, Stéphane; Silvy, Françoise; Martinez, Emmanuelle; Crenon, Isabelle; Germain, Sébastien; Garcia, Stéphane; Viola, Angèle; Lombardo, Dominique

    2015-01-01

    Oncofetal fucose-rich glycovariants of the pathological bile salt-dependent lipase (pBSDL) appear during human pancreatic oncogenesis and are detected by themonoclonal antibody J28 (mAbJ28). We aimed to identify murine counterparts onpancreatic ductal adenocarcinoma (PDAC) cells and tissue and investigate the potential of dendritic cells (DC) loaded with this unique pancreatic tumor antigen to promote immunotherapy in preclinical trials. Pathological BSDLs purified from pancreatic juices of patients with PDAC were cleaved to generate glycosylated C-terminal moieties (C-ter) containing mAbJ28-reactive glycoepitopes. Immunoreactivity of the murine PDAC line Panc02 and tumor tissue to mAbJ28 was detected by immunohistochemistry and flow cytometry. C-ter-J28+ immunization promoted Th1-dominated immune responses. In vitro C-ter-J28+-loaded DCskewed CD3+ T-cells toward Th1 polarization. C-ter-J28+-DC-vaccinations selectively enhanced cell immunoreactivity to Panc02, as demonstrated by CD4+- and CD8+-T-cell activation, increased percentages of CD4+- and CD8+-T-cells and NK1.1+ cells expressing granzyme B, and T-cell cytotoxicity. Prophylactic and therapeutic C-ter-J28+-DC-vaccinations reduced ectopic Panc02-tumor growth, provided long-lasting protection from Panc02-tumor development in 100% of micebut not from melanoma, and attenuated progression of orthotopic tumors as revealed by MRI. Thusmurine DC loaded with pancreatic tumor-specific glycoepitope C-ter-J28+ induce efficient anticancer adaptive immunity and represent a potential adjuvant therapy for patients afflicted with PDAC. PMID:26405163

  14. Sweet and Sour: The Impact of Differential Glycosylation in Cancer Cells Undergoing Epithelial–Mesenchymal Transition

    PubMed Central

    Freire-de-Lima, Leonardo

    2014-01-01

    Glycosylation changes are a feature of disease states. One clear example is cancer cells, which commonly express glycans at atypical levels or with different structural attributes than those found in normal cells. Epithelial–mesenchymal transition (EMT) was initially recognized as an important step for morphogenesis during embryonic development, and is now shown to be one of the key steps promoting tumor metastasis. Cancer cells undergoing EMT are characterized by significant changes in glycosylation of the extracellular matrix (ECM) components and cell-surface glycoconjugates. Current scientific methodology enables all hallmarks of EMT to be monitored in vitro and this experimental model has been extensively used in oncology research during the last 10 years. Several studies have shown that cell-surface carbohydrates attached to proteins through the amino acids, serine, or threonine (O-glycans), are involved in tumor progression and metastasis, however, the impact of O-glycans on EMT is poorly understood. Recent studies have demonstrated that transforming growth factor-beta (TGF-β), a known EMT inducer, has the ability to promote the up-regulation of a site-specific O-glycosylation in the IIICS domain of human oncofetal fibronectin, a major ECM component expressed by cancer cells and embryonic tissues. Armed with the knowledge that cell-surface glycoconjugates play a major role in the maintenance of cell homeostasis and that EMT is closely associated with glycosylation changes, we may benefit from understanding how unusual glycans can govern the molecular pathways associated with cancer progression. This review initially focuses on some well-known changes found in O-glycans expressed by cancer cells, and then discusses how these alterations may modulate the EMT process. PMID:24724053

  15. Binding of galectin-1 to breast cancer cells MCF7 induces apoptosis and inhibition of proliferation in vitro in a 2D- and 3D- cell culture model.

    PubMed

    Geiger, Pamina; Mayer, Barbara; Wiest, Irmi; Schulze, Sandra; Jeschke, Udo; Weissenbacher, Tobias

    2016-11-08

    Galectin-1 (gal-1) belongs to the family of β-galactoside-binding proteins which primarily recognizes the Galβ1-4GlcNAc sequences of oligosaccharides associated with several cell surface glycoconjugates. The lectin recognizes correspondent glycoepitopes on human breast cancer cells. Galectin-1 is expressed both in normal and malignant tissues. Lymphatic organs naturally possessing high rates of apoptotic cells, express high levels of Galectin-1. Furthermore galectin-1 can initiate T cell apoptosis. Binding of galectin-1 to trophoblast tumor cells presenting the oncofetal Thomsen-Friedenreich (TF) carbohydrate antigen inhibits tumor cell proliferation. In this study we examined the impact galectin-1 has in vitro on cell proliferation, apoptotic potential and metabolic activity of MCF-7 and T-47D breast cancer cells in dependence to their expression of the Thomsen-Friedenreich (TF) tumor antigen. For proliferation and apoptosis assays cells were grown in presence of 10, 30 and 60 μg gal-1/ml medium. Cell proliferation was determined by a BrdU uptake ELISA. Detection of apoptotic cells was done by M30 cyto death staining, in situ nick translation and by a nucleosome ELISA method. Furthermore we studied the impact galectin-1 has on the metabolic activity of MCF-7 and T-47D cells in a homotypic three-dimensional spheroid cell culture model mimicking a micro tumour environment. Gal-1 inhibited proliferation of MCF-7 cells (strong expression of the TF epitope) but did not significantly change proliferation of T-47D cells (weak expression of the TF epitope). The incubation of MCF-7 cells with gal-1 raised number of apoptotic cells significantly. Treating the spheroids with 30 μg/ml galectin-1 in addition to standard chemotherapeutic regimes (FEC, TAC) resulted in further suppression of the metabolic activity in MCF-7 cells whereas T-47D cells were not affected. Our results demonstrate that galectin-1 can inhibit proliferation und metabolic cell activity and induce

  16. Expression of serum albumin and of alphafetoprotein in murine normal and neoplastic primitive embryonic structures.

    PubMed

    Trojan, J; Naval, X; Johnson, T; Lafarge-Frayssinet, C; Hajeri-Germond, M; Farges, O; Pan, Y; Uriel, J; Abramasky, O; Ilan, J

    1995-12-01

    corresponding morphologically neoplastic tissues of the teratocarcinomas. The same SA:AFP relationship constitutes an oncofetal marker of primitive neuroectoblastic structures.

  17. Characterization of H type 1 and type 1 N-acetyllactosamine glycan epitopes on ovarian cancer specifically recognized by the anti-glycan monoclonal antibody mAb-A4.

    PubMed

    Choo, Matthew; Tan, Heng Liang; Ding, Vanessa; Castangia, Roberto; Belgacem, Omar; Liau, Brian; Hartley-Tassell, Lauren; Haslam, Stuart M; Dell, Anne; Choo, Andre

    2017-04-14

    Cancer-specific glycans of ovarian cancer are promising epitopes for targeting with monoclonal antibodies (mAb). Despite their potential, structural characterization of these glycan epitopes remains a significant challenge in mAb preclinical development. Our group generated the monoclonal antibody mAb-A4 against human embryonic stem cells (hESC), which also bound specifically to N-glycans present on 11 of 19 ovarian cancer (OC) and 8 of 14 breast cancer cell lines tested. Normal cell lines and tissue were unstained by mAb-A4. To characterize the N-linked glycan epitopes on OC cell lines targeted by mAb-A4, we used glycosidases, glycan microarray, siRNA, and advanced high sensitivity matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The mAb-A4 epitopes were found to be Fucα1-2Galβ1-3GlcNAcβ (H type 1) and Galβ1-3GlcNAcβ (type 1 LacNAc). These structures were found to be present on multiple proteins from hESC and OC. Importantly, endo-β-galactosidase coupled with MALDI-MS allowed these two epitopes, for the first time, to be directly identified on the polylactosamines of N-glycans of SKOV3, IGROV1, OV90, and OVCA433. Furthermore, siRNA knockdown of B3GALT5 expression in SKOV3 demonstrated that mAb-A4 binding was dependent on B3GALT5, providing orthogonal evidence of the epitopes' structures. The recognition of oncofetal H type 1 and type 1 LacNAc on OC by mAb-A4 is a novel and promising way to target OC and supports the theory that cancer can acquire stem-like phenotypes. We propose that the orthogonal framework used in this work could be the basis for advancing anti-glycan mAb characterization. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Characterization of H type 1 and type 1 N-acetyllactosamine glycan epitopes on ovarian cancer specifically recognized by the anti-glycan monoclonal antibody mAb-A4

    PubMed Central

    Choo, Matthew; Tan, Heng Liang; Ding, Vanessa; Castangia, Roberto; Belgacem, Omar; Liau, Brian; Hartley-Tassell, Lauren; Haslam, Stuart M.; Dell, Anne; Choo, Andre

    2017-01-01

    Cancer-specific glycans of ovarian cancer are promising epitopes for targeting with monoclonal antibodies (mAb). Despite their potential, structural characterization of these glycan epitopes remains a significant challenge in mAb preclinical development. Our group generated the monoclonal antibody mAb-A4 against human embryonic stem cells (hESC), which also bound specifically to N-glycans present on 11 of 19 ovarian cancer (OC) and 8 of 14 breast cancer cell lines tested. Normal cell lines and tissue were unstained by mAb-A4. To characterize the N-linked glycan epitopes on OC cell lines targeted by mAb-A4, we used glycosidases, glycan microarray, siRNA, and advanced high sensitivity matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). The mAb-A4 epitopes were found to be Fucα1–2Galβ1–3GlcNAcβ (H type 1) and Galβ1–3GlcNAcβ (type 1 LacNAc). These structures were found to be present on multiple proteins from hESC and OC. Importantly, endo-β-galactosidase coupled with MALDI-MS allowed these two epitopes, for the first time, to be directly identified on the polylactosamines of N-glycans of SKOV3, IGROV1, OV90, and OVCA433. Furthermore, siRNA knockdown of B3GALT5 expression in SKOV3 demonstrated that mAb-A4 binding was dependent on B3GALT5, providing orthogonal evidence of the epitopes' structures. The recognition of oncofetal H type 1 and type 1 LacNAc on OC by mAb-A4 is a novel and promising way to target OC and supports the theory that cancer can acquire stem-like phenotypes. We propose that the orthogonal framework used in this work could be the basis for advancing anti-glycan mAb characterization. PMID:28167527

  19. Deregulation of MYCN, LIN28B and LET7 in a Molecular Subtype of Aggressive High-Grade Serous Ovarian Cancers

    PubMed Central

    Cowin, Prue A.; Johnstone, Cameron N.; House, Colin M.; Sheppard, Karen E.; Etemadmoghadam, Dariush; Melnyk, Nataliya; Rustgi, Anil K.; Phillips, Wayne A.; Johnsen, Hilde; Holm, Ruth; Kristensen, Gunnar B.; Birrer, Michael J.; Pearson, Richard B.; Børresen-Dale, Anne-Lise; Huntsman, David G.; deFazio, Anna; Creighton, Chad J.; Smyth, Gordon K.; Bowtell, David D. L.

    2011-01-01

    Molecular subtypes of serous ovarian cancer have been recently described. Using data from independent datasets including over 900 primary tumour samples, we show that deregulation of the Let-7 pathway is specifically associated with the C5 molecular subtype of serous ovarian cancer. DNA copy number and gene expression of HMGA2, alleles of Let-7, LIN28, LIN28B, MYC, MYCN, DICER1, and RNASEN were measured using microarray and quantitative reverse transcriptase PCR. Immunohistochemistry was performed on 127 samples using tissue microarrays and anti-HMGA2 antibodies. Fluorescence in situ hybridisation of bacterial artificial chromosomes hybridized to 239 ovarian tumours was used to measure translocation at the LIN28B locus. Short interfering RNA knockdown in ovarian cell lines was used to test the functionality of associations observed. Four molecular subtypes (C1, C2, C4, C5) of high-grade serous ovarian cancers were robustly represented in each dataset and showed similar pattern of patient survival. We found highly specific activation of a pathway involving MYCN, LIN28B, Let-7 and HMGA2 in the C5 molecular subtype defined by MYCN amplification and over-expression, over-expression of MYCN targets including the Let-7 repressor LIN28B, loss of Let-7 expression and HMGA2 amplification and over-expression. DICER1, a known Let-7 target, and RNASEN were over-expressed in C5 tumours. We saw no evidence of translocation at the LIN28B locus in C5 tumours. The reported interaction between LIN28B and Let-7 was recapitulated by siRNA knockdown in ovarian cancer cell lines. Our results associate deregulation of MYCN and downstream targets, including Let-7 and oncofetal genes, with serous ovarian cancer. We define for the first time how elements of an oncogenic pathway, involving multiple genes that contribute to stem cell renewal, is specifically altered in a molecular subtype of serous ovarian cancer. By defining the drivers of a molecular subtype of serous ovarian cancers we

  20. IMP-1 displays crosstalk with K-Ras and modulates colon cancer cell survival through the novel pro-apoptotic protein CYFIP2

    PubMed Central

    Mongroo, Perry S.; Noubissi, Felicite K.; Cuatrecasas, Miriam; Kalabis, Jiri; King, Catrina E.; Johnstone, Cameron N.; Bowser, Mark J.; Castells, Antoni; Spiegelman, Vladimir S.; Rustgi, Anil K.

    2011-01-01

    Insulin-like growth factor 2 mRNA-binding protein-1 (IMP-1) is an oncofetal protein that binds directly to and stabilizes oncogenic c-Myc and regulates in turn its post-transcriptional expression and translation. In contrast to normal adult tissue, IMP-1 is re-expressed and/or overexpressed in human cancers. We demonstrate that knock-down of c-Myc in human colon cancer cell lines increases the expression of mature let-7 miRNA family members and downregulates several of its mRNA targets: IMP-1, Cdc34, and K-Ras. We further demonstrate that loss of IMP-1 inhibits Cdc34, Lin-28B, and K-Ras, and suppresses SW-480 cell proliferation and anchorage-independent growth, and promotes caspase and lamin-mediated cell death. We also found that IMP-1 binds to the coding region and 3′UTR of K-Ras mRNA. RNA microarray profiling and validation by reverse transcription PCR reveals that the p53-inducible pro-apoptotic protein, CYFIP2, is upregulated in IMP-1 knock-down SW480 cells, a novel finding. We also show that overexpression of IMP-1 increases c-Myc and K-Ras expression, and LIM2405 cell proliferation. Furthermore, we show that loss of IMP-1 induces Caspase-3 and Parp–mediated apoptosis, and inhibits K-Ras expression in SW480 cells, which is rescued by CYFIP2 knock-down. Importantly, analysis of 228 patients with colon cancers reveals that IMP-1 is significantly upregulated in differentiated colon tumors (p ≤ 0.0001) and correlates with K-Ras expression (r=0.35, p ≤ 0.0001) relative to adjacent normal mucosa. These findings indicate that IMP-1, interrelated with c-myc, acts upstream of K-Ras to promote survival through a novel mechanism that may be important in colon cancer pathogenesis. PMID:21252116