- Email: [email protected]
P53 METABOLIC AND ONCOGENIC CONTRIBUTIONS OF MHC-AP IN OBESITY-ASSOCIATED HEPATOCELLULAR CARCINOMA
POSTERS is an urgent need for new systemic therapies. Although sorafenib, a multikinase inhibitor with antiangiogenic and antiproliferative effects, currently sets the new standard for advanced HCC, tumor response rates are usually quite low. An understanding of the underlying mechanisms for sorafenib resistance is critical if outcomes are to be improved. Methods: In this study, MTS or transwell assay was conducted to examine the ability of cell survival or migration. Lentivirusmediated siRNA targeted for hypoxia inducible factor (HIF)-2a or HPPCn was introdued to HCC cells. Results: In this study, we found that hypoxia protected HCC cells against sorafenib and HIF-2a was required for the process. The combination of lentivirus-mediated HIF-2a shRNA and sorafenib showed synergistically effects against tumor growth and metastasis. HPPCn, a growth factor from hepatic stimulator substance, was demonstrated to stimulate cell growth and metastasis in HCC. We found that HPPCn level was increased under hypoxia and treatment with recombinant human (rh) HPPCn promoted sorafenib resistance by increasing HIF-2a accumulation in HCC. HPPCn knockdown by lentivirus mediated shRNA alleviated sorafenib resistance through inhibiting HIF-2a accumulation under hypoxia. Additionally, DeSUMOylation of HIF-2a by SUMO-specific protease 1 (SENP1) increased HIF-2a accumulation, and HPPCn upregulated the expression of SENP1 by increasing phosphorylation of Akt under hypoxia. Conclusions: Hypoxia induced by sustained sorafenib treatment confers sorafenib resistance to HCC through HIF-2a. HPPCn promoted sorafenib resistance through increasing HIF-2a level. Aktmediated SENP1 upregulation is responsible for HPPCn-induced HIF-2a accumulation under hypoxia. P52 HEPATITIS B VIRUS X PROTEIN MODULATES TUMOR MICRO-ENVIRONMENT BY MEANS OF EXOSOMES N.R. Kapoor. ICGEB, New Delhi, India E-mail: [email protected] Background and Aims: Hepatis B Virus X Protein (HBx) is a pleiotropic transactivator and has been closely associated with development of liver cancer in mouse models and in clinical cases. We sought to determine the modulation of exosomal biogenesis and secretion pathway by HBx protein in regulating naive parenchymal and non parenchymal cells. Methods: Exosome partcle count from Huh7 cells expressing HBx along with mRNA and activity of neutral sphingomyelinase II, a key enzyme involved in exosome biogenesis was investigated. Purified exosomes from HBx expressing hepatocytes were co-cultured with Hepatic stellate cell line (LX-2) or Immortalized primary hepatocyte cell line (IHH). The relative mRNA levels of molecules involved in stellate cell activation like collagen, alpha smooth muscle actin were monitored. Reactivation of notch ignaling in IHH cells was also monitored by qRT PCR. Results: Our data points towards role of HBx in modulating exosomal biogenesis and secretion. We observed a 1.5 fold rise in exosome secretion from HBx expressing cells along with increase in nSMNASeII mRNA and activity.we also demonstrate that HBx modulated the exosomal content to stimulate quiescent HSC to undergo activation as indicated by incresed mRNA levels of clo1A, col 3A and aSMA. HBx exosomes lead to a reactivation of Notch signaling pathway in IHH cells indicate by increase in mRNA levels of Jag-1 and Hes-1. Conclusions: HBx protein modulates exosomal content leading to activation of naive hepatocytes and the stromal cells.
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P53 METABOLIC AND ONCOGENIC CONTRIBUTIONS OF MHC-AP IN OBESITY-ASSOCIATED HEPATOCELLULAR CARCINOMA A.L. Eheim1,2,3 , M. Berriel Diaz1,2,3 , B. Meissburger1,2,3 , S. Herzig1,2,3 . 1 Joint Research Division, Molecular Metabolic Control, German Cancer Research Center (DKFZ), 2 Center for Molecular Biology (ZMBH), University of Heidelberg, 3 Network Aging Research, University Hospital Heidelberg, Heidelberg, Germany E-mail: [email protected] Background and Aims: Hepatic manifestations of metabolic syndrome, specifically NAFLD, are significant risk factors for hepatocellular carcinoma (HCC), with HCC incidence projected to rise in accordance with obesity prevalence. In contrast to historical risk factors (viral hepatitis/ALD), molecular mechanisms underlying progression of NAFLD/NASH to HCC are largely unknown; therefore we sought to identify novel factors regulating tumorigenesis in a setting of obesity-associated HCC. Methods: DEN-induced carcinogenesis was paired with dietinduced obesity/NAFLD using C3H mice. Expression profiling of tumor and non-tumor tissue from low-fat (LFD) or high-fat (HFD) diet fed-mice was performed and screened against hepatic gene expression data from genetic/diet-induced obesity and NASH. Upon confirmation in human data-sets, selected candidates were characterized for metabolic and oncogenic function in vitro. Results: As expected, HCC development was significantly accelerated in HFD-fed mice. Hepatic expression of Major Histocompatibility Complex-Associated Protein (MHC-AP) expression was elevated in tumor-bearing mice fed LFD and in HFDfed controls. Interestingly, tumors derived from HFD-fed mice were characterized by significantly higher MHC-AP expression vs. tumors from LFD-fed mice, indicating MHC-AP may be involved in the pathogenesis of obesity-associated HCC. siRNA-mediated knockdown of MHC-AP significantly decreased cell proliferation (~60%) in hepatoma cells (Hepa1–6, HepG2) and induced significant changes in oxygen consumption and extracellular acidification rates. In vitro data also indicate MHC-AP expression is subject to regulation by bioactive phospholipids and insulin simulation. Conclusions: These data demonstrate both metabolic and oncogenic functionality of MHC-AP, supporting a functional relevance in obesity-associated HCC. Further investigation of MHC-AP at the mechanistic level may expand metabolocentric approaches of HCC treatment. P54 TEMPORAL DYNAMICS AND THERAPEUTIC POTENTIAL OF THE UNFOLDED PROTEIN RESPONSE IN HEPATOCELLULAR CARCINOMA Y.-P. Vandewynckel1 , D. Laukens1 , A. Geerts1 , E. Bogaerts1 , A. Paridaens1 , X. Verhelst1 , C. Van Steenkiste1 , B. Descamps2 , C. Vanhove2 , L. Libbrecht3 , R. De Rycke4 , B. Lambrecht5 , S. Janssens5 , H. Van Vlierberghe1 . 1 Ghent University Hospital, 2 Infinity Imaging Lab, Ghent University, 3 Pathology, Ghent University Hospital, 4 Ghent University, 5 Unit Immunoregulation and Mucosal Immunology, VIB Inflammation Research Center, Ghent, Belgium E-mail: [email protected] Background and Aims: Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related mortality. The unfolded protein response (UPR) is involved in the consecutive steps of carcinogenesis. Little is known about the temporal alterations of the UPR during carcinogenesis. We examined the kinetics and effect of UPR modulation in HCC. Methods: The UPR was evaluated in a diethylnitrosamine-induced mouse model. Mice were sacrificed every 5 weeks until week 30. Hematoxylin/eosin, Sirius red and reticulin staining and electron microscopy were performed. Expression analysis was performed in a cohort of human HCC. The effect of UPR modulation on cell
Journal of Hepatology 2014 vol. 60 | S67–S214
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P53 METABOLIC AND ONCOGENIC CONTRIBUTIONS OF MHC-AP IN OBESITY-ASSOCIATED HEPATOCELLULAR CARCINOMA A.L. Eheim1,2,3 , M. Berriel Diaz1,2,3 , B. Meissburger1,2,3 , S. Herzig1,2,3 . 1 Joint Research Division, Molecular Metabolic Control, German Cancer Research Center (DKFZ), 2 Center for Molecular Biology (ZMBH), University of Heidelberg, 3 Network Aging Research, University Hospital Heidelberg, Heidelberg, Germany E-mail: [email protected] Background and Aims: Hepatic manifestations of metabolic syndrome, specifically NAFLD, are significant risk factors for hepatocellular carcinoma (HCC), with HCC incidence projected to rise in accordance with obesity prevalence. In contrast to historical risk factors (viral hepatitis/ALD), molecular mechanisms underlying progression of NAFLD/NASH to HCC are largely unknown; therefore we sought to identify novel factors regulating tumorigenesis in a setting of obesity-associated HCC. Methods: DEN-induced carcinogenesis was paired with dietinduced obesity/NAFLD using C3H mice. Expression profiling of tumor and non-tumor tissue from low-fat (LFD) or high-fat (HFD) diet fed-mice was performed and screened against hepatic gene expression data from genetic/diet-induced obesity and NASH. Upon confirmation in human data-sets, selected candidates were characterized for metabolic and oncogenic function in vitro. Results: As expected, HCC development was significantly accelerated in HFD-fed mice. Hepatic expression of Major Histocompatibility Complex-Associated Protein (MHC-AP) expression was elevated in tumor-bearing mice fed LFD and in HFDfed controls. Interestingly, tumors derived from HFD-fed mice were characterized by significantly higher MHC-AP expression vs. tumors from LFD-fed mice, indicating MHC-AP may be involved in the pathogenesis of obesity-associated HCC. siRNA-mediated knockdown of MHC-AP significantly decreased cell proliferation (~60%) in hepatoma cells (Hepa1–6, HepG2) and induced significant changes in oxygen consumption and extracellular acidification rates. In vitro data also indicate MHC-AP expression is subject to regulation by bioactive phospholipids and insulin simulation. Conclusions: These data demonstrate both metabolic and oncogenic functionality of MHC-AP, supporting a functional relevance in obesity-associated HCC. Further investigation of MHC-AP at the mechanistic level may expand metabolocentric approaches of HCC treatment. P54 TEMPORAL DYNAMICS AND THERAPEUTIC POTENTIAL OF THE UNFOLDED PROTEIN RESPONSE IN HEPATOCELLULAR CARCINOMA Y.-P. Vandewynckel1 , D. Laukens1 , A. Geerts1 , E. Bogaerts1 , A. Paridaens1 , X. Verhelst1 , C. Van Steenkiste1 , B. Descamps2 , C. Vanhove2 , L. Libbrecht3 , R. De Rycke4 , B. Lambrecht5 , S. Janssens5 , H. Van Vlierberghe1 . 1 Ghent University Hospital, 2 Infinity Imaging Lab, Ghent University, 3 Pathology, Ghent University Hospital, 4 Ghent University, 5 Unit Immunoregulation and Mucosal Immunology, VIB Inflammation Research Center, Ghent, Belgium E-mail: [email protected] Background and Aims: Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related mortality. The unfolded protein response (UPR) is involved in the consecutive steps of carcinogenesis. Little is known about the temporal alterations of the UPR during carcinogenesis. We examined the kinetics and effect of UPR modulation in HCC. Methods: The UPR was evaluated in a diethylnitrosamine-induced mouse model. Mice were sacrificed every 5 weeks until week 30. Hematoxylin/eosin, Sirius red and reticulin staining and electron microscopy were performed. Expression analysis was performed in a cohort of human HCC. The effect of UPR modulation on cell
Journal of Hepatology 2014 vol. 60 | S67–S214