Cancer Biology and Signal Transduction Inhibition of Stat5a/b Enhances Proteasomal Degradation of Androgen Receptor Liganded by Antiandrogens in Prostate Cancer

Although poorly understood, androgen receptor (AR) signaling is sustained despite treatment of prostate cancer with antiandrogens and potentially underlies development of incurable castrateresistant prostate cancer. However, therapies targeting the AR signaling axis eventually fail when prostate cancer progresses to the castrate-resistant stage. Stat5a/b, a candidate therapeutic target protein in prostate cancer, synergizes with AR to reciprocally enhance the signaling of both proteins. In this work, we demonstrate that Stat5a/b sequesters antiandrogen-liganded (MDV3100, bicalutamide, flutamide) AR in prostate cancer cells and protects it against proteasomal degradation in prostate cancer. Active Stat5a/b increased nuclear levels of both unliganded and antiandrogen-liganded AR, as demonstrated in prostate cancer cell lines, xenograft tumors, and clinical patient-derived prostate cancer samples. Physical interaction between Stat5a/b and AR in prostate cancer cellswasmediated by theDNA-bindingdomainof Stat5a/b and the N-terminal domain of AR. Moreover, active Stat5a/b increased AR occupancy of the prostate-specific antigen promoter and AR-regulated gene expression in prostate cancer cells. Mechanistically, both Stat5a/b genetic knockdown and antiandrogen treatment induced proteasomal degradation of AR in prostate cancer cells, with combined inhibition of Stat5a/b and AR leading to maximal loss of AR protein and prostate cancer cell viability. Our results indicate that therapeutic targeting of AR in prostate cancer using antiandrogens may be substantially improved by targeting of Stat5a/b. Mol Cancer Ther; 14(3); 713–26. 2014 AACR. Introduction Although curative therapies exist for organ-confined or locally advanced prostate cancer, treatment options are limited for metastatic castrate-resistant prostate cancer (CRPC). Progression to CRPC is defined by resistance to androgen-deprivation therapy, which typically occurs less than 3 years after initiation of androgen-deprivation therapy (1–3). The transition from androgendependent prostate cancer to CRPC is not fully understood. Androgen receptor (AR) signaling is known to be maintained despite low levels of circulatory androgens (1), which has been attributed to numerous mechanisms, including (i) AR gene amplification (4); (ii) AR ligand-binding domain mutations conferring ligand promiscuity (5, 6); (iii) intracrine androgen biosynthesis within prostate cancer cells from adrenal steroids and cholesterol (7, 8); (iv) ligand-independent, noncanonical AR transactivation by kinase signaling pathways (9, 10); and (v) upregulation of constitutively active, AR splice variants that do not require ligand to support prostate cancer growth (11, 12). Most therapeutic strategies for prostate cancer are directed at the AR signaling axis, with established antiandrogens such as Deparment of CancerBiology, SidneyKimmel CancerCenter,Thomas JeffersonUniversity, Philadelphia, Pennsylvania. DepartmentofMedical Oncology, H. Lee Moffit Cancer Center and Research Institute, University of South Florida,Tampa, Florida. Department of Biochemistry and Molecular Biology, Sidney Kimmel Cancer Center, Thomas JeffersonUniversity, Philadelphia, Pennsylvania. Deparment of Physiology, University of Turku, Turku, Finland. Center for Disease Modeling, Institute of Biomedicine, University of Turku,Turku, Finland. Division of Biostatistics, Department of Pharmacology and Experimental Therapeutics, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania. Department of Pathology, Haartman Institute, University of Helsinki and HUSLAB, Helsinki, Finland. Finnish Institute for Molecular Medicine (FIMM), University of Helsinki, Helsinki, Finland. Department of Urology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania. Department of Medical Oncology, Sidney Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania. Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/). D.T. Hoang and L. Gu contributed equally to this article. Current address for Feng Shen: Division of Oncology, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania; current address for Mateusz Koptyra, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania; current address for Shyh-HanTan, Department of Surgery, Center for ProstateDisease Research, Uniformed Services University of the Health Sciences, Rockville, Maryland; and current address for Ayush Dagvadorj, Department of Cytogenetics and Molecular Biology, Grandmed Hospital, Ulaanbaatar, Mongolia. Corresponding Author: Marja T. Nevalainen, Departments of Cancer Biology, Medical Oncology and Urology, Sidney Kimmel Cancer Center, Thomas Jefferson University, 233 South 10th Street, BLSB 309, Philadelphia, PA 19107. Phone: 215-503-9250; Fax: 215-503-9245; E-mail: [email protected] doi: 10.1158/1535-7163.MCT-14-0819 2014 American Association for Cancer Research. Molecular Cancer Therapeutics www.aacrjournals.org 713 on June 24, 2017. © 2015 American Association for Cancer Research. mct.aacrjournals.org Downloaded from Published OnlineFirst December 31, 2014; DOI: 10.1158/1535-7163.MCT-14-0819