Presentation Authors: Chengfei Liu*, Wei Lou, Joy C. Yang, Liangren Liu, Cameron M. Armstrong, Alan P. Lombard, Ruining Zhao, Onika DV Noel, Clifford G. Tepper, Mark Dall’Era, Hong-Wu Chen, Christopher P. Evans, Allen C. Gao, Sacramento, CA
Introduction: Proteostasis is a complementary process by which cells control the protein biosynthesis, folding, trafficking and degradation. Evidence suggests that proteomic instability, such as protein misfolding and aggregation plays pivotal roles in cancer cell survival and progression. AR-V7 lacking the ligand binding domain confers resistance to enzalutamide and abiraterone in prostate cancer, targeting AR-V7 is an logical strategy to overcome the resistance. However, the mechanisms of AR-V7 proteostasis have not been fully studied so far. The objectives of this project are to investigate the roles of proteostasis in regulating AR-V7 expression and sensitivity to enzalutamide and abiraterone treatment.
Methods: Expression of HSP70 and STUB1 was determined by qRT-PCR and western blot. Expression of HSP70 and STUB1 was downregulated using specific siRNA. HSP70/STUB1 and AR-V7 interaction was determined by co-immunoprecipitation and dual immunofluorescence. The gene regulating mechanisms underlying the HSP70 inhibition in drug resistant prostate cancer cells was determined by RNA sequencing analyses. The effects of HSP70 inhibition on enzalutamide sensitivity were examined in vitro and in vivo. The correlation between HSP70 and AR-V7 in high Gleason score prostate tumors was determined by qRT-PCR.
Results: In the present study, we analyzed enzalutamide and abiraterone resistant prostate cancer cells and found ubiquitin mediated proteolysis pathway was suppressed, and E3 ubiquitin ligases STUB1 is down regulated in enzalutamide and abiraterone resistant prostate cancer cells. STUB1 binds to AR-V7, degrades AR-V7 expression and suppresses its activity. HSP70, the STUB1 binding protein, also binds to AR-V7 and enhances AR-V7 transcriptional activity. Mechanistically, STUB1 disassociates HSP70 from AR-V7 binding and increases AR-V7 ubiquitination and degradation. Targeting HSP70 by siRNA or small molecular inhibitors (Apoptozole and Ver155008) significantly suppressed prostate cancer growth and improved enzalutamide and abiraterone treatment through AR-V7 inhibition in vitro and in vivo. Additionally, HSP70 expression is upregulated in mCRPC tumors and correlates with AR-V7 levels in high Gleason score and metastatic prostate tumor specimens.
Conclusions: Enzalutamide and abiraterone treatment induces the imbalance of AR-V7 proteostasis through the ubiquitin-proteolysis alteration. HSP70/STUB1 complex controls AR and AR variants proteostasis. Targeting HSP70 could be a valuable strategy to overcome the next generation anti-androgen resistance and improve their therapy.
Source of Funding: Grant Support: This work was supported in part by grants NIH/NCI CA168601, CA179970, DOD PC150229, and the U.S. Department of Veterans Affairs, Office of Research & Development BL&D grant number I01BX0002653 (A.C. G) and by resources from the VA Northern