Upregulation of constitutively-active androgen receptor splice variants (AR-Vs) has been implicated in AR-driven tumor progression in castration-resistant prostate cancer. To date, functional studies of AR-Vs have been focused mainly on their ability to regulate gene expression independent of the full-length AR (AR-FL). Here, we showed that AR-V7 and ARv567es, two major AR-Vs, both facilitated AR-FL nuclear localization in the absence of androgen and mitigated the ability of the antiandrogen enzalutamide to inhibit AR-FL nuclear trafficking. AR-V bound to the promoter of its specific target without AR-FL, but co-occupied the promoter of canonical AR target with AR-FL in a mutually-dependent manner. AR-V expression attenuated both androgen and enzalutamide modulation of AR-FL activity/cell growth, and mitigated the in vivo antitumor efficacy of enzalutamide. Furthermore, ARv567es levels were upregulated in xenograft tumors that had acquired enzalutamide resistance. Collectively, this study highlights a dual function of AR-Vs in mediating castration resistance. In addition to trans-activating target genes independent of AR-FL, AR-Vs can serve as a “rheostat” to control the degree of response of AR-FL to androgen-directed therapy via activating AR-FL in an androgen-independent manner. The findings shed new insights into the mechanisms of AR-V-mediated castration resistance and have significant therapeutic implications.

Constitutively-active androgen receptor splice variants (AR-V) lacking the ligand-binding domain have been implicated in the pathogenesis of castration-resistant prostate cancer and in mediating resistance to newer drugs that target the androgen axis. AR-V regulate expression of both canonical AR targets and a unique set of cancer-specific targets that are enriched for cell cycle functions. However, little is known about how AR-V control gene expression. Here we report that two major AR-V, termed AR-V7 and ARv567es, not only homodimerize and heterodimerize with each other but also heterodimerize with full-length androgen receptor (AR-FL) in an androgen-independent manner. We found that heterodimerization of AR-V and AR-FL was mediated by N- and C-terminal interactions and by the DNA-binding domain of each molecule, whereas AR-V homodimerization was mediated only by DNA-binding domain interactions. Notably, AR-V dimerization was required to transactivate target genes and to confer castration-resistant cell growth. Our results clarify the mechanism by which AR-V mediate gene regulation and provide a pivotal pathway for rational drug design to disrupt AR-V signaling, as a rational strategy for effective treatment of advanced prostate cancer.