MP73-05 ACTIVATION OF SERINE BIOSYNTHESIS PATHWAY IN HIF2? ANTAGONIST-RESISTANT RENAL CELL CARCINOMA CELLS AFTER ACQUIRING SUNITINIB RESISTANCE: RESULTS FROM THE GENOME EDITING TECHNOLOGY

Abstract

You have accessJournal of UrologyKidney Cancer: Basic Research & Pathophysiology III1 Apr 2017MP73-05 ACTIVATION OF SERINE BIOSYNTHESIS PATHWAY IN HIF2? ANTAGONIST-RESISTANT RENAL CELL CARCINOMA CELLS AFTER ACQUIRING SUNITINIB RESISTANCE: RESULTS FROM THE GENOME EDITING TECHNOLOGY Hideki Enokida, Hirofumi Yoshino, Kazutaka Miyamoto, Masaya Yonemori, Satoru Sugita, Takashi Sakaguchi, and Masayuki Nakagawa Hideki EnokidaHideki Enokida More articles by this author , Hirofumi YoshinoHirofumi Yoshino More articles by this author , Kazutaka MiyamotoKazutaka Miyamoto More articles by this author , Masaya YonemoriMasaya Yonemori More articles by this author , Satoru SugitaSatoru Sugita More articles by this author , Takashi SakaguchiTakashi Sakaguchi More articles by this author , and Masayuki NakagawaMasayuki Nakagawa More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.3350AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES ontinuous activation of hypoxia-inducible factor (HIF) is important for renal cell carcinoma (RCC) progression as well as resistance acquisition against mTOR and VEGFR inhibitors. Recently a HIF2α antagonist (PT2385) has been developed, and is currently being investigated in a phase I clinical trial for advanced or metastatic clear cell RCC (ccRCC) patients who had prior tyrosine kinase inhibitors. However, resistant mechanisms against the HIF2α antagonist seems to be a next problem in the near future. The aim of this study was to find activated signals involved in the HIF2α antagonist-resistance after acquiring sunitinib resistance. METHODS First, we established sunitinib resistant 786-o (SU-R-786-o) cells in vivo by oral gavage treatment. Instead of the HIF2α antagonist administration, we completely knocked out the HIF2α gene expression in SU-R-786-o cells by using a recent genome editing technology, CRISPR/Cas9 system, with two different single guide RNAs against HIF2α. We identified their characteristics in cell proliferation assay and western blot. These cells were also analyzed in proteomics and RNA sequence analyses to elucidate their activated signals. RESULTS By using CRISPR/Cas9 system, we succeeded in establishing HIF2α knockout sunitinib resistant 786-o (HIF2α-KO-SU-R-786-o) cells. Cell morphology was changed from spindle to round shape in HIF2α-KO-SU-R-786-o cells. In addition, cell proliferation in HIF2α-KO-SU-R-786-o cells became significantly slower than in the parental cells as well as SU-R-786-o cells (P < 0.01). After recovering cell proliferation in HIF2α-KO-SU-R-786-o cells, proteomics and RNA sequence revealed that CXCR4 expression was dramatically up-regulated in SU-R-786-o cells compared to the parental cells, however, it was decreased in HIF2α-KO-SU-R-786-o cells compared to SU-R-786-o cells. Another surprise was that serine biosynthesis pathway was accelerated in HIF2α-KO-SU-R-786-o cells compared to SU-R-786-o cells. CONCLUSIONS Based on the proteomics and RNA sequence analyses, we found that CXCR4 upregulation or serine biosynthesis might be critical for acquiring resistance against multi kinase inhibitors or HIF2α antagonist. Even though both drugs target angiogenesis pathway, the activated signals were altered in the different therapeutic stage by the different drugs administration. The inhibitions of serine biosynthesis pathways might be potential targeted therapy for advanced or metastatic ccRCC which shows resistance to HIF2α agonist after acquiring resistance to multi kinase inhibitors. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e965 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Hideki Enokida More articles by this author Hirofumi Yoshino More articles by this author Kazutaka Miyamoto More articles by this author Masaya Yonemori More articles by this author Satoru Sugita More articles by this author Takashi Sakaguchi More articles by this author Masayuki Nakagawa More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...