266 Background: The persistent activation of the androgen receptor (AR) signaling axis plays a major role in castration-resistant prostate cancer (CRPC). This knowledge has driven the clinical development of new inhibitors of androgen production (e.g., abiraterone) and AR signaling (e.g., enzalutamide). However, in many patients, the efficacy is short-lived and eventually resistance appears. Reactivation of AR signaling occurs by several mechanisms in CRPC. In this study, we report the biological properties of TAS3681, a new AR antagonist with AR downregulating activity, and propose this concept as a potential new approach for the treatment of CRPC. Methods: For assay of AR transactivation, COS-7 cells were transiently transfected with androgen-responsive reporter gene construct and expression vectors encoding wt or mu ARs. The transfected cells were treated with TAS3681 and androgen in steroid depleted media for 24 h, and luciferase activity was measured. For an in vivo pharmacodynamic assay to confirm AR downregulation in tumor, TAS3681 was orally dosed in CRPC tumor xenograft model. Tumor was isolated and AR level in tumor was determined by Western blot. Results: TAS3681 suppressed the growth of AR positive prostate cancer (PCa) cells but did not affect that of AR-negative DU145 PCa cells, indicating a dependency on AR for efficacy. TAS3681 did not stimulate AR nuclear translocation and suppressed wt and mu ARs (including F876L) transactivation in cells, indicative of its pure AR antagonist profile. In contrast to enzalutamide, TAS3681 effectively suppressed androgen-independent AR transactivation by growth factors and cytokines via AR downregulating activity. Interestingly, in PCa cells which express full-length and splice variant AR, TAS3681 reduced the expression of both ARs. Moreover, TAS3681 treatment effectively decreased AR level in CRPC tumors in vivo. Conclusions: TAS3681 exerts an anti-androgenic effect via two mechanisms of action: pure AR antagonism and AR decreasing activity. It is expected that TAS3681 has a potential to overcome the resistance to current and 2nd-generation therapies targeting AR signaling.