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Exploiting cross-talk between lipid metabolism and oncogenic signaling for treatment of ovarian cancer
EACR24 Poster Sessions / European Journal of Cancer 61, Suppl. 1 (2016) S9–S218 Conclusions: In patients, 95% of metastatic cases occur in the liver, however, the reason for this preference is unknown. The results presented here suggest that BAP1 LOF alters numerous properties of human uveal melanoma cell lines, including adhesion, proliferation a cellular signalling, which may drive this preference. No conflict of interest. 313 SIRT1 in bladder cancer E. Chapman1 , E. Baxter2 , J.A. Roulson3 , M. Sanchez-Carbayo4 . 1 Leeds Institute of Cancer and Pathology St. James’s University Hospital, St James’s University Hospital, Leeds, United Kingdom, 2 University of Leeds, Leeds Institute of Rheumatic and Musculoskeletal Medicine, Leeds, United Kingdom, 3 Leeds Teaching Hospitals NHS Trust, Department of Cellular Pathology, Leeds, United Kingdom, 4 University of the Basque Country, Bladder Cancer Group- Lascaray Research Center, Vitoria-Gasteiz, Spain Introduction: The role of histone deacetylase, Sirtuin 1 (SIRT1) in cancer is unclear. In some tumour types over-expression of SIRT1 correlates with worse prognosis and chemoresistance. In others, expression is lost in tumours and a tumour suppressor role is suggested. Either induction or inhibition of SIRT1 have been proposed as cancer treatment, yet its role in many cancer types is currently not understood. Immortalization of normal human urothelial cells with hTERT, the catalytic subunit of telomerase, led to an increase in SIRT1 expression. In these cells, SIRT1 is implicated in mediating changes in gene expression during the early stages of tumorigenesis. For the first time, we investigated SIRT1 expression and phenotypic effects in primary urothelial carcinoma (UC). Material and Methods: SIRT1 transcript was quantified in 44 UC cell lines by expression array analysis. Immunohistochemistry was performed on 2 independent panels of UC and sections were scored for nuclear and/or cytoplasmic staining. Stable retroviral-mediated transduction with shRNA was used to inhibit expression in UC cell lines in vitro. Results: Over-expression of SIRT1 was detected in UC cell lines and primary UC. Preliminary data indicates that nuclear SIRT1 over-expression correlates with tumour stage and grade and recurrence free survival. Extra nuclear, cytoplasmic SIRT1 accumulation was also observed, consistent with the reported nuclear-cytoplasmic translocation. In this small panel, cytoplasmic SIRT1 appeared to correlate with a poor prognosis. SIRT1 was commonly co-expressed with polycomb repressor complex members BMI1 and EZH2. Attenuation of SIRT1 expression in UC cell lines in vitro, increased apoptosis and decreased cell migration. Anchorage independent growth of tumour cell lines in vitro was attenuated. We investigated the involvement of SIRT1dependent deacetylation of Cortactin in mediating phenotypic effects. Conclusions: Preliminary data indicates that SIRT1 is up-regulated in urothelial cancer and may be a potential therapeutic target. The significance of nuclear and cytoplasmic accumulation of SIRT1 merits investigation in a larger tumour set. Future studies will further investigate the mechanisms by which SIRT1 modulates tumorigenicity in UC. No conflict of interest. 314 Activation of ER stress and autophagy induced by pterostilbene via Akt/mTOR pathway in human hepatocellular carcinoma cells H.L. Chiou1 , C.L. Yu2 , Y.H. Hsieh3 . 1 Chung Shan Medical University-, School of Medical Laboratory and Biotechnology, Taichung, Taiwan, 2 Chung Shan Medical University-, Institute of Medicine, Taichung, Taiwan, 3 Chung Shan Medical University-, Department of Biochemistry- School of Medicine, Taichung, Taiwan Introduction: Hepatocellular carcinoma (HCC) is the one of the most common cancers worldwide and the leading cause of cancer-related death. Due to the poor response rate and severe side effect of current treatment, new effective therapy strategies are urgently needed. Pterostilbene, a natural analogue of resveratrol, is known to have diverse pharmacologic activities including antioxidation, anti-inflammation and anti-proliferation. The aim of this study was to verify the feasibility of pterostilbene being an anticancer agent and related molecular mechanisms. Material and Method: MTT assay was used to examine the effect of pterostilbene on cell viability of two human HCC cell lines, Huh 7 and SK-Hep 1. Flow cytometry assays with acridine orange staining, DCFDA staining and JC-1 staining, were used to identify the source of cellular stress. Immunoblotting was also used to clarify the effect of pterostilbene on the expressions of related proteins. Results and Discussion: Results of this study indicated that pterostilbene may inhibit cell growth and induce autophagy of HCC cells, as evidenced by increased acridine orange staining, accumulation of LC3-II and upregulated p62/SQSTM1 expression. Subsequent analysis by flow cytometry and western blotting have revealed that pterostilbene increased the formation of reactive oxygen species (ROS) and the expressions of endoplasmic reticulum (ER) stress related-proteins (ATF4, Bip, PERK, IRE1a and CHOP)
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in cells. Moreover, pterostilbene-induced autophagy was suggested by the decreased expression of phosphorylated Akt/mTOR family proteins. Conclusion: Overall, our results demonstrated that pterostilbene could induce ER stress and autophagy through Akt/mTOR inactivation in HCC cells while autophagy inhibition enhanced pterostilben-induced cell death. These findings could potentially contribute to the beneficial effect of pterostilbene in HCC treatments. No conflict of interest. 315 Exploiting cross-talk between lipid metabolism and oncogenic signaling for treatment of ovarian cancer 2 R. Wagner1 , G. Stubiger ¨ , P. Lanzerstorfer3 , J. Weghuber3 , E. Karteris4 , ´ ıguez6 , K. Nowikovsky1 , N. Wilfinger1 , R. Colomer5 , M.L. Lopez-Rodr´ T.W. Grunt7 . 1 Medical University Vienna- Comprehensive Cancer Center, Department of Medicine I- Division of Oncology, Vienna, Austria, 2 Medical University Vienna- Comprehensive Cancer Center, Department of Biomedical Imaging and Imageguided Therapy, Vienna, Austria, 3 University of Applied Sciences Upper Austria, School of Engineering and Environmental Sciences, Wels, Austria, 4 Brunel University London, Department of BiosciencesSchool of Health Sciences & Social Care, Uxbridge, United Kingdom, 5 Hospital Universitario La Princesa and Spanish National Cancer Research Center CNIO, Department of Medical Oncology, Madrid, Spain, 6 Universidad ´ Complutense de Madrid, Dpto Qu´ımica Organica I- Fac. CC. Qu´ımicas, Madrid, Spain, 7 Medical University Vienna- Comprehensive Cancer CenterLudwig Boltzmann Cluster Oncology, Department of Medicine I- Division of Oncology, Vienna, Austria
Background: Fatty acid synthase (FASN), a regulator of lipid biosynthesis, is overexpressed in ovarian cancer and indicative of poor prognosis. FASNinhibition abolishes cancer cell proliferation and survival. Control of FASN by phosphatidylinositol(PI)3-kinase(PI3K)-mechanistic target of rapamycin complex 1(mTORC1) signaling is well documented, whereas reverse interaction from FASN towards PI3K-mTORC1 is only poorly understood and underlying molecular mechanisms are still elusive. However, since malignancy is driven by PI3K-mTORC1, control of this pathway by FASN is highly relevant for therapy prompting an in-depth analysis. Materials and Methods: To this end, SKOV3, OVCAR-3 or HOC-7 cells were exposed to FASN-antagonists (C75, G28UCM) and analyzed by thin-layer chromatography, matrix-assisted laser desorption/ionization timeof-flight tandem-mass spectrometry, ELISA, Western blotting, quantitative micropatterning and growth assays. Results: We demonstrate that FASN-blockade normalizes cell membranes by preferential incorporation of polyunsaturated fatty acids such as arachidonic acid and diminishes diacylglycerols. Inhibition of FASN affects PI- and lipid raft-composition, which impairs upstream EGF-receptor/ERBB/HER complex function/expression and Grb2 recruitment causing PI3K-silencing represented by depletion of its product phosphatidylinositol (3,4,5)-trisphosphate and inactivation of downstream AKT. Moreover, FASN-blockers rapidly induce cell stress responses characterized by up-regulation of HIF-1a and its targetgene REDD1(RTP801/DIG2/DDIT4), followed by activation of the energysensor AMPK. REDD1 and AMPK in turn act as mTORC1-repressors blocking ribosomal S6 protein. Thus, blockade of malignant lipogenesis efficaciously impairs receptor-PI3K-mTORC1 at multiple molecular levels. Interestingly, concurrent targeting of PI3K/mTOR with dactolisib/NVP-BEZ235 does not augment FASN-inhibitor efficacy indicating that FASN-blockade fully silenced PI3K-mTORC1 already on its own. In contrast, FASN-inhibitor-mediated silencing of receptor-PI3K-mTORC1 releases cross-repression of MEK-ERK thereby causing its concurrent trans-activation. Accordingly, co-treatment with MEK-inhibitor selumetinib/AZD6244 significantly enhances FASN-inhibitor action. Conclusions: This is to the best of our knowledge the first characterization of PI3K-mTORC1 cross-silencing by FASN-inhibitors, which may be crucial for the anticancer effects of these drugs. Moreover, our data encourage therapeutic approaches using FASN-antagonists together with MEK-ERK-inhibitors. No conflict of interest. 316 Platycodon grandiflorum-induced autophagy and growth inhibition in human lung cancer cells C. Park1 , J.W. Jeong2 , E.O. Choi3 , M.H. Han4 , Y.H. Choi2 , S.H. Hong2 . 1 Department of Molecular Biology- College of Natural Sciences and Human Ecology, Dongeui University, Busan, Korea, 2 Department of Biochemistry, Dongeui University College of Korean Medicine, Busan, Korea, 3 Anti-Aging Research Center and Blue-Bio Industry Regional Innovation Center, Dongeui University, Busan, Korea, 4 Natural Products Research Team- Converging Research Division, National Marine Biodiversity institute of Korea, Seocheon, Korea Introduction: Autophagy is a fundamental cellular process for the degradation of damaged organelles and proteins. Accumulation of recent studies showed that and defects in autophagy lead to a number of diseases including lung
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in cells. Moreover, pterostilbene-induced autophagy was suggested by the decreased expression of phosphorylated Akt/mTOR family proteins. Conclusion: Overall, our results demonstrated that pterostilbene could induce ER stress and autophagy through Akt/mTOR inactivation in HCC cells while autophagy inhibition enhanced pterostilben-induced cell death. These findings could potentially contribute to the beneficial effect of pterostilbene in HCC treatments. No conflict of interest. 315 Exploiting cross-talk between lipid metabolism and oncogenic signaling for treatment of ovarian cancer 2 R. Wagner1 , G. Stubiger ¨ , P. Lanzerstorfer3 , J. Weghuber3 , E. Karteris4 , ´ ıguez6 , K. Nowikovsky1 , N. Wilfinger1 , R. Colomer5 , M.L. Lopez-Rodr´ T.W. Grunt7 . 1 Medical University Vienna- Comprehensive Cancer Center, Department of Medicine I- Division of Oncology, Vienna, Austria, 2 Medical University Vienna- Comprehensive Cancer Center, Department of Biomedical Imaging and Imageguided Therapy, Vienna, Austria, 3 University of Applied Sciences Upper Austria, School of Engineering and Environmental Sciences, Wels, Austria, 4 Brunel University London, Department of BiosciencesSchool of Health Sciences & Social Care, Uxbridge, United Kingdom, 5 Hospital Universitario La Princesa and Spanish National Cancer Research Center CNIO, Department of Medical Oncology, Madrid, Spain, 6 Universidad ´ Complutense de Madrid, Dpto Qu´ımica Organica I- Fac. CC. Qu´ımicas, Madrid, Spain, 7 Medical University Vienna- Comprehensive Cancer CenterLudwig Boltzmann Cluster Oncology, Department of Medicine I- Division of Oncology, Vienna, Austria
Background: Fatty acid synthase (FASN), a regulator of lipid biosynthesis, is overexpressed in ovarian cancer and indicative of poor prognosis. FASNinhibition abolishes cancer cell proliferation and survival. Control of FASN by phosphatidylinositol(PI)3-kinase(PI3K)-mechanistic target of rapamycin complex 1(mTORC1) signaling is well documented, whereas reverse interaction from FASN towards PI3K-mTORC1 is only poorly understood and underlying molecular mechanisms are still elusive. However, since malignancy is driven by PI3K-mTORC1, control of this pathway by FASN is highly relevant for therapy prompting an in-depth analysis. Materials and Methods: To this end, SKOV3, OVCAR-3 or HOC-7 cells were exposed to FASN-antagonists (C75, G28UCM) and analyzed by thin-layer chromatography, matrix-assisted laser desorption/ionization timeof-flight tandem-mass spectrometry, ELISA, Western blotting, quantitative micropatterning and growth assays. Results: We demonstrate that FASN-blockade normalizes cell membranes by preferential incorporation of polyunsaturated fatty acids such as arachidonic acid and diminishes diacylglycerols. Inhibition of FASN affects PI- and lipid raft-composition, which impairs upstream EGF-receptor/ERBB/HER complex function/expression and Grb2 recruitment causing PI3K-silencing represented by depletion of its product phosphatidylinositol (3,4,5)-trisphosphate and inactivation of downstream AKT. Moreover, FASN-blockers rapidly induce cell stress responses characterized by up-regulation of HIF-1a and its targetgene REDD1(RTP801/DIG2/DDIT4), followed by activation of the energysensor AMPK. REDD1 and AMPK in turn act as mTORC1-repressors blocking ribosomal S6 protein. Thus, blockade of malignant lipogenesis efficaciously impairs receptor-PI3K-mTORC1 at multiple molecular levels. Interestingly, concurrent targeting of PI3K/mTOR with dactolisib/NVP-BEZ235 does not augment FASN-inhibitor efficacy indicating that FASN-blockade fully silenced PI3K-mTORC1 already on its own. In contrast, FASN-inhibitor-mediated silencing of receptor-PI3K-mTORC1 releases cross-repression of MEK-ERK thereby causing its concurrent trans-activation. Accordingly, co-treatment with MEK-inhibitor selumetinib/AZD6244 significantly enhances FASN-inhibitor action. Conclusions: This is to the best of our knowledge the first characterization of PI3K-mTORC1 cross-silencing by FASN-inhibitors, which may be crucial for the anticancer effects of these drugs. Moreover, our data encourage therapeutic approaches using FASN-antagonists together with MEK-ERK-inhibitors. No conflict of interest. 316 Platycodon grandiflorum-induced autophagy and growth inhibition in human lung cancer cells C. Park1 , J.W. Jeong2 , E.O. Choi3 , M.H. Han4 , Y.H. Choi2 , S.H. Hong2 . 1 Department of Molecular Biology- College of Natural Sciences and Human Ecology, Dongeui University, Busan, Korea, 2 Department of Biochemistry, Dongeui University College of Korean Medicine, Busan, Korea, 3 Anti-Aging Research Center and Blue-Bio Industry Regional Innovation Center, Dongeui University, Busan, Korea, 4 Natural Products Research Team- Converging Research Division, National Marine Biodiversity institute of Korea, Seocheon, Korea Introduction: Autophagy is a fundamental cellular process for the degradation of damaged organelles and proteins. Accumulation of recent studies showed that and defects in autophagy lead to a number of diseases including lung