Therapeutics, Targets, and Chemical Biology Cotargeting Androgen Receptor and Clusterin Delays Castrate-Resistant Prostate Cancer Progression by Inhibiting Adaptive Stress Response and AR Stability Hiroaki Matsumoto 1 , Yoshiaki Yamamoto 1 , Masaki Shiota 1 , Hidetoshi Kuruma 1 , Eliana Beraldi 1 , Hideyasu Matsuyama 2 , Amina Zoubeidi 1 , and Martin Gleave 1 Abstract Although androgen receptor (AR) pathway inhibitors prolong survival in castrate-resistant prostate cancer (CRPC), resistance rapidly develops and is often associated with increased stress-activated molecular chaperones like clusterin (CLU) and continued AR signaling. Because adaptive pathways activated by treatment facilitate development of acquired resistance, cotargeting the stress response, activated by AR inhibition and mediated through CLU, may create conditional lethality and improve outcomes. Here, we report that CLU is induced by AR antagonism and silencing using MDV3100 and antisense, respectively, to become highly expressed in castrate- and MDV3100-resistant tumors and cell lines. CLU, as well as AKT and mitogen-activated protein kinase (MAPK) signalosomes, increase in response to MDV3100-induced stress. Mechanistically, this stress response is coordinated by a feed-forward loop involving p90rsk (RPS6KA)-mediated phosphoactivation of YB-1 with subsequent induction of CLU. CLU inhibition repressed MDV3100-induced activation of AKT and MAPK pathways. In addition, when combined with MDV3100, CLU knockdown accelerated AR degradation and repressed AR transcriptional activity through mechanisms involving decreased YB-1–regulated expression of the AR cochaperone, FKBP52. Cotargeting the AR (with MDV3100) and CLU (with OGX-011) synergistically enhanced apoptotic rates over that seen with MDV3100 or OGX-011 monotherapy and delayed CRPC LNCaP tumor and prostate-specific antigen (PSA) progression in vivo. These data indicate that cotargeting adaptive stress pathways activated by AR pathway inhibitors, and mediated through CLU, creates conditional lethality and provides mechanistic and preclinical proof-of-principle to guide biologically rational combinatorial clinical trial design. Cancer Res; 73(16); 5206–17. Ó2013 AACR. Introduction Prostate cancer is the second leading cause of cancer deaths among males in western countries (1). Although early-stage disease is treated with curative surgery or radiotherapy, the mainstay of treatment for locally advanced, recurrent or metastatic prostate cancer is androgen deprivation therapy (ADT), which reduces serum testosterone to castrate levels and suppresses androgen receptor (AR) activity. Despite high initial response rates after ADT, progression to castrate-resistant prostate cancer (CRPC) occurs within 3 years (2–4). The AR is the principle driver of CRPC (5), and is supported by ADT- activated growth factor signaling pathways (6), survival genes (7), and cytoprotective chaperone networks (8). Docetaxel (9) was the first therapy to prolong survival in CRPC, stratifying the treatment landscape into pre and postchemotherapy states. More recently, the CYP17 inhibitor abiraterone (10) and the AR antagonist MDV3100 (11) have prolonged survival and are rapidly changing the CRPC treatment landscape. Despite significant responses (11, 12), these novel AR pathway inhibitors activate redundant survival pathways that adaptive- ly drive treatment resistance and recurrent CRPC progression. Realization of the full potential of AR pathway inhibition will require characterization of these stress-activated survival responses and rational combinatorial cotargeting strategies designed to abrogate them. Molecular chaperones play central roles in stress responses by maintaining protein homeostasis and regulating prosurvi- val signaling and transcriptional networks. Clusterin (CLU), a stress-activated chaperone transcriptionally regulated by HSF1 (13), YB-1 (14), and others (15), inhibits stress-induced apoptosis by suppressing protein aggregation (16), p53-acti- vating stress signals (17), and conformationally altered Bax (17, 18) while enhancing AKT phosphorylation (19) and transactivation of NF-kB and HSF-1 (13, 20). CLU is expressed in many human cancers (21, 22), including prostate where it increases following castration to become highly expressed in Authors' Affiliations: 1 The Vancouver Prostate Centre and Department of Urologic Sciences, University of British Columbia, Vancouver, British Columbia, Canada; and 2 Department of Urology, Graduate School of Medicine, Yamaguchi University, Ube, Japan Note: Supplementary data for this article are available at Cancer Research Online (http://cancerres.aacrjournals.org/). Corresponding Author: Martin E Gleave, The Vancouver Prostate Centre, 2660 Oak Street, Vancouver, British Columbia V6H 3Z6, Canada. Phone: 604-875-4818; Fax: 604-875-5654; E-mail: m.gleave@ubc.ca doi: 10.1158/0008-5472.CAN-13-0359 Ó2013 American Association for Cancer Research. Cancer Research Cancer Res; 73(16) August 15, 2013 5206 on August 31, 2021. © 2013 American Association for Cancer Research. cancerres.aacrjournals.org Downloaded from Published OnlineFirst June 20, 2013; DOI: 10.1158/0008-5472.CAN-13-0359