Effect of a new type androgen receptor antagonist, TAS3681, on ligand-independent AR activation through its AR downregulation activity.

Abstract

199 Background: Two new therapies, enzalutamide and abiraterone, directed at the androgen receptor (AR) signaling axis, represent important advances in the management of castration-resistant prostate cancer (CRPC). However, eventually almost all of patients acquire resistance to these drugs by a variety of mechanisms. Ligand independent AR activation such as induction of AR splice variants and AR overexpression are major issues of current CRPC progression. In the present study, we report the biological characterization of TAS3681, which is a new AR antagonist with AR downregulation activity, and propose this concept as a potential new approach for the treatment of CRPC. Methods: For assay of AR transactivation, prostate cancer (PCa) cells were transiently transfected with androgen-responsive reporter gene construct. The transfected cells were treated with growth factor and cytokine in steroid-depleted media, and luciferase activity was measured. To evaluate the effect of TAS3681 on AR and c-Myc protein expression, PCa cells were treated with TAS3681 in steroid-depleted media. AR and c-Myc protein levels were determined by western blot. Real-time PCR was used to analyze the mRNA levels of c-Myc and c-Myc target gene. Chromatin immunoprecipitation was performed to determine the enrichment of AR at the element. Results: TAS3681 dose-dependently reduced AR protein levels in PCa cells. In contrast to enzalutamide, TAS3681 suppressed androgen-independent AR transactivation by growth factor and cytokine. In PCa cells which express full-length AR and splice variant AR-v7, TAS3681 suppressed AR-v7 target gene expression through downregulation of AR-v7. Moreover, TAS3681 reduced expression of c-Myc, critical driver of androgen-independent mechanisms of PCa progression, via AR downregulation activity. In addition, real-time PCR assay showed the transcriptional suppression of c-Myc and its target gene by TAS3681. Conclusions: TAS3681 exhibits suppressive effects on ligand-independent AR activation via AR decreasing activity. These finding suggest that TAS3681 could be a candidate of breakthrough therapy for resistance to current AR pathway target drugs.