Human Cancer Biology Steroidogenic Enzyme AKR1C3 Is a Novel Androgen Receptor-Selective Coactivator that Promotes Prostate Cancer Growth Muralimohan Yepuru 1 , Zhongzhi Wu 1 , Anand Kulkarni 2 , Feng Yin 1 , Christina M. Barrett 1 , Juhyun Kim 1 , Mitchell S. Steiner 1 , Duane D. Miller 1 , James T. Dalton 1 , and Ramesh Narayanan 1 Abstract Purpose: Castration-resistant prostate cancer (CRPC) may occur by several mechanisms including the upregulation of androgen receptor (AR), coactivators, and steroidogenic enzymes, including aldo keto reductase 1C3 (AKR1C3). AKR1C3 converts weaker 17-keto androgenic precursors to more potent 17- hydroxy androgens and is consistently the major upregulated gene in CRPC. The studies in the manuscript were undertaken to examine the role of AKR1C3 in AR function and CRPC. Experimental Design: LNCaP cells stably transfected with AKR1C3 and VCaP cells endogenously expressing AKR1C3 were used to understand the effect of AKR1C3 on prostate cancer cell and tumor growth in nude mice. Chromatin immunoprecipitation, confocal microscopy, and co-immunoprecipita- tion studies were used to understand the recruitment of AKR1C3, intracellular localization of AKR1C3 and its interaction with AR in cells, tumor xenograft, and in Gleason sum 7 CRPC tissues. Cells were transiently transfected for AR transactivation. Novel small-molecule AKR1C3-selective inhibitors were synthesized and characterized in androgen-dependent prostate cancer and CRPC models. Results: We identified unique AR-selective coactivator- and prostate cancer growth-promoting roles for AKR1C3. AKR1C3 overexpression promotes the growth of both androgen-dependent prostate cancer and CRPC xenografts, with concomitant reactivation of androgen signaling. AKR1C3 interacted with AR in prostate cancer cells, xenografts, and in human CRPC samples and was recruited to the promoter of an androgen-responsive gene. The coactivator and growth-promoting functions of AKR1C3 were inhibited by an AKR1C3-selective competitive inhibitor. Conclusions: AKR1C3 is a novel AR-selective enzymatic coactivator and may represent the first of more than 200 known nuclear hormone receptor coactivators that can be pharmacologically targeted. Clin Cancer Res; 19(20); 5613–25. Ó2013 AACR. Introduction Prostate cancer is the most frequently diagnosed cancer in men with more than 200,000 new prostate cancer cases estimated for 2012 (American Cancer Society, Facts and Figures). The mainstay of therapy for advanced prostate cancer is the reduction of peripheral androgens to castrate levels. Androgen deprivation therapy creates an androgen- deficient state in which prostate cancer evolves into castra- tion-resistant prostate cancer (CRPC) and tumor progres- sion occurs (1). Newer therapies that target androgen metabolizing enzymes and/or androgen receptor (AR) have shown clinical efficacy, indicating the continued impor- tance of the androgen signaling axis in advanced prostate cancer (2). A variety of mechanisms are thought to contribute to the emergence of the androgen hypersensitivity that is observed in CRPC. Overexpression of AR and coactivators drives the growth and metastasis of CRPC even in response to low levels of androgens (3–5). Despite having castrate levels of serum testosterone, tumor samples obtained from men with CRPC have only a 60% reduction in intratumoral dihydrotestosterone (DHT). This suggests that intratumoral conversion of weak adrenal androgens or androgen pre- cursors produced de novo in the tumor fuel the growth of CRPC (6). Androgen biosynthesis enzyme inhibitors have been used in the treatment of advanced prostate cancer. Drugs such as ketoconazole and abiraterone acetate to treat Authors' Affiliations: 1 Preclinical Research and Development, GTx Inc.; and 2 Department of Pathology, University of Tennessee Health Science Center, Memphis, Tennessee Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). J.T. Dalton and R. Narayanan share senior authorship. Corresponding Authors: Ramesh Narayanan, Preclinical Research and Development, GTx Inc., 3 North Dunlap, Memphis, TN 38163. Phone: 901- 523-9700; Fax: 901-523-9772; E-mail: rnarayanan@gtxinc.com; and James T. Dalton, E-mail: jdalton@gtxinc.com doi: 10.1158/1078-0432.CCR-13-1151 Ó2013 American Association for Cancer Research. Clinical Cancer Research www.aacrjournals.org 5613 on February 9, 2016. © 2013 American Association for Cancer Research. clincancerres.aacrjournals.org Downloaded from Published OnlineFirst August 30, 2013; DOI: 10.1158/1078-0432.CCR-13-1151