Genome-wide flow sorting CRISPRi screen identify PTGES3 as a novel therapeutic target of AR

Comprehensively understanding the gene regulatory network that modulates the biology of the Androgen Receptor (AR) oncogene is a central goal of prostate cancer research. To study regulators of AR, we knock-in a split neon green fluorescent protein onto the AR in prostate cancer cells creating an endogenous AR reporter cell line. Using our endogenous AR reporter, we performed a genome-scale fluorescence-activated cell sorting based CRISPRi screen to comprehensively identify genes that modulate AR protein levels. We identify and validate known AR protein regulators including HOXB13, GATA2, GRHL2 as well as unexpected top hits including PTGES3, SAE1 and CSNK2A1. Loss of PTGES3 in multiple models of ARSI-resistant Pca resulted in loss of AR protein and thus AR’s oncogenic activity. Mechanistically, we demonstrate a novel nuclear co-factor function of PTGES3 facilitating AR-mediated transcription aside from its canonical co-chaperone function regulating AR protein levels. Lastly using a disulfide tethering chemical screen, we developed a PTGES3 inhibitor demonstrating PTGES3 is a therapeutically tractable target for the treatment of AR-addicted mCRPC.