Identify Mechanisms Driving Resistance to AR Antagonists in CRPC

Beth Israel Deaconess Medical Center, Boston MA

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Abstract

PROJECT SUMMARY/ABSTRACT Prostate cancer (PCa) that recurs after standard androgen deprivation therapy (ADT) with a GnRH agonist or antagonist (castration-resistant prostate cancer, CRPC) is in most cases still dependent on androgen receptor (AR). AR activity in these patients can be further suppressed by agents such as abiraterone or by AR antagonists such as enzalutamide (ENZ), apalutamide, or darolutamide, but patients treated with these androgen signaling inhibitor (ASI) drugs still invariably progress. A subset of tumors that progress despite ADT combined with ASI drugs become AR independent, but the majority continue to have high levels of AR and AR transcriptional activity. Therefore, a major focus of our efforts has been to determine the basis for this persistent AR activity. Our approach during the current P01 funding period has been to trace the progression to ENZ-resistance in VCaP cells, which express a wild-type amplified AR. ENZ treatment initially suppresses AR activity and proliferation, but these cells adapt over 1-2 months with restoration of AR activity despite culture in 16 µM ENZ (VCaP16 cells). Notably, AR activity in these cells is driven by an AR splice variant 7 (ARv7, the most common AR splice variant), and is not dependent on the full length AR (ARfl). Moreover, we find that ARv7 activity is dependent on adaptations that enhance chromatin accessibility. Sites with increased chromatin accessibility were strongly associated with binding sites for the NFI family of transcription factors, and NFI depletion decreased ARv7 chromatin binding and mimicked the effects of ARv7 depletion on the transcriptome. Aim 1 is to test the hypothesis that NFI transcription factors are major contributors to establishing and maintaining the AR cistrome and transcriptome, and to ENZ-resistance. In the ENZ-resistant VCaP16 cells we further found that FOXA1 binding was increased at ARv7 binding sites, and by mass spectrometry identified increased phosphorylation at sites that may enhance nucleosome binding. These findings are pursued in Aim 2. Aim 3 then focuses on identification of additional adaptations that are required to support reactivation of the AR transcriptional program in response to AR targeted therapies. The specific aims are 1) to determine the role of NFI transcription factors in PCa progression and ARv7 function, 2) to determine the role of FOXA1 posttranslational modifications in supporting ARv7 function, and 3) to identify further adaptations that are critical to support ARv7 function. This project makes extensive use of the PDX models and expertise in Core B and the genomic capabilities and expertise in Core C. Moreover, our focus on how AR activity at all or subsets of genes is highly related to the fundamental question addressed in Project 1, which is asking what are the critical functions of AR in PCa. We will also have extensive interactions with Project 3, which will address the role of PARP in ARv7 functions. Finally, our use of shared models and real-time sharing of data (facilitated by Core A) has during the current funding cycle (and will continue to) lead to novel insights and new approaches and directions.