696 CHARACTERIZATION OF A C-TERMINAL HEAT SHOCK PROTEIN 90 INHIBITOR IN PROSTATE CANCER CELLS

Abstract

You have accessJournal of UrologyProstate Cancer: Basic Research V1 Apr 2010696 CHARACTERIZATION OF A C-TERMINAL HEAT SHOCK PROTEIN 90 INHIBITOR IN PROSTATE CANCER CELLS Heidi A. Penn, Craig A. Manthe, Shawna B. Comer, George A. Vielhauer, Roger J. Rajewski, Brian S.J. Blagg, and Jeffrey M. Holzbeierlein Heidi A. PennHeidi A. Penn Kansas City, KS More articles by this author , Craig A. MantheCraig A. Manthe Kansas City, KS More articles by this author , Shawna B. ComerShawna B. Comer Kansas City, KS More articles by this author , George A. VielhauerGeorge A. Vielhauer Kansas City, KS More articles by this author , Roger J. RajewskiRoger J. Rajewski Lawrence, KS More articles by this author , Brian S.J. BlaggBrian S.J. Blagg Lawrence, KS More articles by this author , and Jeffrey M. HolzbeierleinJeffrey M. Holzbeierlein Kansas City, KS More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2010.02.1095AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Heat shock protein 90 (Hsp90) assists in folding, activation, and maintenance of proteins, as well as protects from degradation in stressed cells. In cancer, it has been found to be constitutely over-expressed and involved in various signaling pathways to promote tumor survival. Because of this, HSP 90 is a viable drug target. Traditionally, Hsp90 inhibitors have targeted the N-terminal ATP-binding site and 17-AAG has demonstrated activity both in vitro and in vivo and clinical trials are currently ongoing. Unfortunately, significant toxicity and difficult pharmacokinetic properties have limited their use. A second ATP-binding site was identified in the carboxyl terminus of Hsp90 and the antibiotic novobiocin has been demonstrated to weakly bind to and inhibit, however it has very low potency. We evaluate the in vitro efficacy of one of our analogues, KU-174, in hormone dependent (LNCaP-LN3) and independent (PC3-MM2) prostate cancer cell lines and normal renal proximal tubule epithelial cells (RPTEC). METHODS Three cell lines, LNCaP-LN3, PC3-MM2, and RPTEC, were treated with KU-174, 17-AAG, and novobiocin and the effects compared. MTS anti-proliferation assays were used to determine potency. Western blot experiments determined the effect on Hsp90, Hsp70 and client protein expression and trypan blue exclusion assays assessed cell death. Additionally, KU-174 was analyzed in flow cytometry experiments for the induction of apoptosis and arrest of cell cycle. RESULTS KU-174 demonstrated anti-proliferative activity in both cancer cell lines and was more potent at earlier time points compared to novobiocin and 17-AAG. KU-174 induced the degradation of client proteins without inducing a heat-shock response. The percentage of viable cells was decreased significantly in both PC3–MM2 and LNCaP-LN3 lines with no effect on normal cells and KU-174 appears to be cytotoxic. In cell cycle experiments, KU-174 induced significant G2/M and S-phase arrest with the LNCaP-LN3 cell line demonstrating an increasing percentage of cells in the sub-G0/apoptotic fraction. In both cancer cell lines, the induction of apoptosis and cell cycle arrest increased over time in a dose-dependent manner with less of an effect on RPTEC cells. CONCLUSIONS KU-174 is specific for cancer cells and differs from N-terminal inhibitors in that it is cytotoxic versus cystostatic, it does not induce a heat-shock response, and is more effective at earlier times. KU-174 represents a new class of Hsp-90 inhibitors that may prove to be superior to N-terminal inhibitors for the treatment of prostate cancer. © 2010 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 183Issue 4SApril 2010Page: e271-e272 Advertisement Copyright & Permissions© 2010 by American Urological Association Education and Research, Inc.MetricsAuthor Information Heidi A. Penn Kansas City, KS More articles by this author Craig A. Manthe Kansas City, KS More articles by this author Shawna B. Comer Kansas City, KS More articles by this author George A. Vielhauer Kansas City, KS More articles by this author Roger J. Rajewski Lawrence, KS More articles by this author Brian S.J. Blagg Lawrence, KS More articles by this author Jeffrey M. Holzbeierlein Kansas City, KS More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...