Abstract 5220: Chronic exposure to a novel AR-NTD inhibitor induces resistance via a selective metabolism pathway

Abstract

Abstract Background: The androgen receptor (AR) is recognized as playing a crucial role in prostate cancer (PCa) maintenance and progression; therefore its inhibition has been the cornerstone of modern therapy for men who fail primary treatment. Current treatments are initially effective, however resistance ultimately develops and the disease progresses to a lethal form termed castration-resistant prostate cancer (CRPC). Our lab has discovered a new class of molecules (EPI) which inhibit the AR by binding to the N-terminal domain (NTD). The NTD interacts with transcriptional machinery, and its presence is vital for a transcriptionally functional receptor. We have previously shown that EPI-002 specifically inhibits both full-length AR and constitutively active AR splice variants. The efficacy of EPI-506, the prodrug of EPI-002, is currently being tested in a Phase I/II clinical trial for CRPC. Here we propose novel resistance mechanisms arising from sustained AR-NTD inhibition, with the goal of preemptively developing backup compounds. Methods: The androgen sensitive human PCa cell line LNCaP was used in all experiments. A resistant cell line (LNCaP-EPIR) was generated by passaging parental LNCaP cells weekly in media supplemented with EPI-002 beginning in September 2012. In vitro and in vivo studies using LNCaP and LNCaP-EPIR cells were employed to confirm biological resistance. A human affymetrix microarray identified possible resistance mechanisms and was validated using qRT-PCR, western blot and functional studies. LNCaP and LNCaP-EPIR cells were challenged with EPI-045, an EPI-analog predicted to remain effective in the context of EPI-002 resistance. Results: LNCaP-EPIR cells treated with 25 µM EPI-002 displayed similar growth rates to vehicle treatment, both in vitro and in vivo. Conversely, parental LNCaP cells showed significant growth inhibition in response to EPI-002. LNCaP-EPIR cells retained sensitivity to anti-androgens and AR knock-down by targeted siRNA, implying functional AR remains essential for growth. qRT-PCR data demonstrated that EPI-002 had reduced ability to block AR mediated gene transcription. Interrogation of microarray data revealed candidate genes (UGT2B family) which were specifically upregulated in the resistant line, and may function to metabolize EPI-002. Supporting this hypothesis EPI-045 which is predicted to be resistant to UGT2B metabolism, was able to significantly inhibit LNCaP-EPIR proliferation and AR transcriptional activity. Conclusions: Taken together these data suggest an EPI-specific mechanism of resistance, whereby EPI-002 is preferentially metabolized and removed from the cell. LNCaP-EPIR cells remain dependent upon AR signalling, and are sensitive to anti-androgens used clinically as well as a novel EPI-analog. This work highlights the potential for combination or sequential therapy in the context of drug-resistant CRPC. Citation Format: Jonathon K. Obst, Jun Wang, David Williams, Amy H. Tien, Nasrin R. Mawji, Yu Chi Yang, Raymond J. Andersen, Marianne D. Sadar. Chronic exposure to a novel AR-NTD inhibitor induces resistance via a selective metabolism pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5220. doi:10.1158/1538-7445.AM2017-5220