Covalent Destabilizing Degrader of AR and AR-V7 in Androgen-Independent Prostate Cancer Cells

Abstract Androgen-independent prostate cancers, correlated with heightened aggressiveness and poor prognosis, are caused by mutations or deletions in the androgen receptor (AR) or expression of truncated variants of AR that are constitutively activated. Currently, drugs and drug candidates against AR target the steroid-binding domain to antagonize or degrade AR. However, these compounds cannot therapeutically access largely intrinsically disordered truncated splice variants of AR, such as AR-V7, that only possess the DNA binding domain and are missing the ligand binding domain. Targeting intrinsically disordered regions within transcription factors has remained challenging and is considered “undruggable”. Herein, we leveraged a cysteine-reactive covalent ligand library in a cellular screen to identify degraders of AR and AR-V7 in androgen-independent prostate cancer cells. We identified a covalent compound EN1441 that selectively degrades AR and AR-V7 in a proteasome-dependent manner through direct covalent targeting of an intrinsically disordered cysteine C125 in AR and AR-V7. EN1441 causes significant and selective destabilization of AR and AR-V7, leading to aggregation of AR/AR-V7 and subsequent proteasome-mediated degradation. Consistent with targeting both AR and AR-V7, we find that EN1441 completely inhibits total AR transcriptional activity in androgen-independent prostate cancer cells expressing both AR and AR-V7 compared to AR antagonists or degraders that only target the ligand binding domain of full-length AR, such as enzalutamide and ARV-110. Our results put forth a pathfinder molecule EN1441 that targets an intrinsically disordered cysteine within AR to destabilize, degrade, and inhibit both AR and AR-V7 in androgen-independent prostate cancer cells and highlights the utility of covalent ligand discovery approaches in directly targeting, destabilizing, inhibiting, and degrading classically undruggable transcription factor targets. Competing Interest Statement DKN is a co-founder, shareholder, and scientific advisory board member for Frontier Medicines, Vicinitas Therapeutics, and Zenith Therapeutics. DKN is a member of the board of directors for Vicinitas Therapeutics. DKN is also on the scientific advisory board of and receives payment and/or holds shares in The Mark Foundation for Cancer Research, Photys Therapeutics, Apertor Pharmaceuticals, Oerth Bio, Ten30 Biosciences, and Deciphera Pharmaceuticals. DKN is also an Investment Advisory Partner for a16z Bio, an Advisory Board member for Droia Ventures, and an iPartner for The Column Group. CN, PRP, and YL are employees and stockholders of Amgen.