Enzalutamide, an Androgen Receptor Signaling Inhibitor, InducesTumor Regression in a Mouse Model of Castration-Resistant Prostate Cancer Javier Guerrero, 1 Iv an E. Alfaro, 1 Francisco G omez, 1 Andrew A. Protter, 2 and Sebasti an Bernales 1,2 * 1 Fundaci on Ciencia & Vida, Santiago,Chile 2 Medivation, Inc., San Francisco,California BACKGROUND. Enzalutamide (formerly MDV3100 and available commercially as Xtandi ® ), a novel androgen receptor (AR) signaling inhibitor, blocks the growth of castration-resistant prostate cancer (CRPC) in cellular model systems and was shown in a clinical study to increase survival in patients with metastatic CRPC. Enzalutamide inhibits multiple steps of AR signaling: binding of androgens to AR, AR nuclear translocation, and association of AR with DNA. Here, we investigate the effects of enzalutamide on AR signaling, AR-dependent gene expression and cell apoptosis. METHODS. The expression of AR target gene prostate-specific antigen (PSA) was measured in LnCaP and C4-2 cells. AR nuclear translocation was assessed in HEK-293 cells stably transfected with AR-yellow fluorescent protein. The in vivo effects of enzalutamide were determined in a mouse xenograft model of CRPC. Differential gene expression in LNCaP cells was measured using Affymetrix human genome microarray technology. RESULTS. We found that unlike bicalutamide, enzalutamide lacked AR agonistic activity at effective doses and did not induce PSA expression or AR nuclear translocation. Additionally, it is more effective than bicalutamide at inhibiting agonist-induced AR nuclear translocation. Enzalutamide induced the regression of tumor volume in a CRPC xenograft model and apoptosis in AR-over-expressing prostate cancer cells. Finally, gene expression profiling in LNCaP cells indicated that enzalutamide opposes agonist-induced changes in genes involved in processes such as cell adhesion, angiogenesis, and apoptosis. CONCLUSIONS. These results indicate that enzalutamide efficiently inhibits AR signaling, and we suggest that its lack of AR agonist activity may be important for these effects. Prostate # 2013 Wiley Periodicals, Inc. KEY WORDS: androgen receptor; MDV3100; nuclear translocation; prostate cancer; microarray INTRODUCTION Prostate cancer is one of the most commonly diagnosed cancers in developed countries [1]. Growth and survival of early prostate tumors is highly depen- dent on levels of circulating androgens. Treatments such as androgen deprivation therapy (ADT), which typically includes suppression of testicular androgen by surgical castration or treatment with analogues of luteinizing hormone releasing hormone, are effective at slowing disease progression. In advanced disease, however, the cancer progresses despite low levels of circulating androgens that result from ADT. It had not Conflict of interest: Andrew A. Protter and Sebasti an Bernales are employees of Medivation, Inc., Javier Guerrero, Francisco G omez, and Iv an E. Alfaro were paid contractors to Medivation, Inc., in the development of this manuscript. Together Medivation, Inc. and Astellas Pharma are co-developing and commercializing enzaluta- mide, which was recently approved by the Food and Drug Administration for the treatment of advanced prostate cancer and is commercially available as Xtandi ® (enzalutamide) capsules.  Correspondence to: Sebasti an Bernales, Ph.D., Preclinical Develop- ment, Medivation, Inc., 525 Market Street, 36th Floor, San Francisco, CA 94105. E-mail: sebastian.bernales@medivation.com Received 12 November 2012; Accepted 25 March 2013 DOI 10.1002/pros.22674 Published online in Wiley Online Library (wileyonlinelibrary.com). The Prostate ß 2013 Wiley Periodicals, Inc.