Mp04-09 novel inhibitor of pro-survival protein phosphatase 5 (pp5) leads to apoptosis in clear cell renal cell carcinoma

You have accessJournal of UrologyCME1 Apr 2023MP04-09 NOVEL INHIBITOR OF PRO-SURVIVAL PROTEIN PHOSPHATASE 5 (PP5) LEADS TO APOPTOSIS IN CLEAR CELL RENAL CELL CARCINOMA Elham Ahanin, Rebecca A. Sager, Sarah J. Backe, Natela Dushukyan, Michael Daneshvar, Gennady Bratslavsky, John Chisholm, Mark R. Woodford, Dimitra Bourboulia, and Mehdi Mollapour Elham AhaninElham Ahanin More articles by this author , Rebecca A. SagerRebecca A. Sager More articles by this author , Sarah J. BackeSarah J. Backe More articles by this author , Natela DushukyanNatela Dushukyan More articles by this author , Michael DaneshvarMichael Daneshvar More articles by this author , Gennady BratslavskyGennady Bratslavsky More articles by this author , John ChisholmJohn Chisholm More articles by this author , Mark R. WoodfordMark R. Woodford More articles by this author , Dimitra BourbouliaDimitra Bourboulia More articles by this author , and Mehdi MollapourMehdi Mollapour More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003215.09AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Despite good survival rates for localized renal cell carcinoma a proportion of patients present with or will progress to metastatic disease, which portends worse prognosis and poor overall survival. Survival has been improved with the use of new targeted therapies and immunotherapy regimens; many patients, however, ultimately develop resistance. There is therefore a need for identification and development of new therapeutics. Clear cell renal cell carcinoma (ccRCC), the most common histologic subtype, generally harbors loss of the tumor suppressor von Hippel Lindau (VHL). The serine/threonine protein phosphatase-5 (PP5) plays a role in the regulation of numerous signaling pathways essential for cancer growth. Previously we have shown that PP5 is targeted for ubiquitination and degradation by a VHL-containing E3 ubiquitin ligase complex in a hypoxia-independent manner. Expression and activity of PP5 is consequently increased in ccRCC, and it plays a pro-survival role. Knock-down of PP5 expression or targeting its activity through inhibition of the kinase casein kinase-1δ (CK1δ) caused apoptosis in VHL-null ccRCC. The objective of this study was to develop a novel PP5 inhibitor for ccRCC. METHODS: An in silico docking screen using the crystal structure of the PP5 active site and a large compound library was used to screen for candidate inhibitors. A series of candidates were then evaluated using both in vitro and cell-based assays for their ability to inhibit PP5 activity and cause apoptosis in ccRCC cells. In vitro PP5 activity was assessed using specific phospho-peptide substrates and in cells by immunoblotting to assess phosphorylation levels of known substrates. Two primary candidates were conjugated to biotin for PP5 pulldown experiments to assess specificity. RESULTS: Our in silico screen identified approximately 200 candidate inhibitors. Further screening of about 20 of these compounds identified two compounds, P5 and P13, which inhibit PP5 activity both in vitro and in ccRCC cells and were further refined based on structure. Pulldown using biotin-conjugated drugs demonstrated good inhibitor specificity. Furthermore, treatment with these inhibitors leads to apoptosis specifically in VHL-null ccRCC cell lines. CONCLUSIONS: PP5 promotes cell survival in ccRCC and knockdown or inhibition of PP5 leads to cellular apoptosis. We have identified a small molecule which specifically binds to and inhibits PP5. This novel PP5 inhibitor causes apoptosis in VHL-null ccRCC cells and may serve as a new therapeutic strategy for treatment of advanced ccRCC. Source of Funding: This work was supported by the National Institute of General Medical Sciences of the National Institutes of Health under award numbers R35GM139584 (M.M.) and DoD KC190038 (M.M.) © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e36 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Elham Ahanin More articles by this author Rebecca A. Sager More articles by this author Sarah J. Backe More articles by this author Natela Dushukyan More articles by this author Michael Daneshvar More articles by this author Gennady Bratslavsky More articles by this author John Chisholm More articles by this author Mark R. Woodford More articles by this author Dimitra Bourboulia More articles by this author Mehdi Mollapour More articles by this author Expand All Advertisement PDF downloadLoading ...