FACS-based whole-genome CRISPRi screen to uncover STEAP1 regulators in prostate cancer.

e17084 Background: Six-Transmembrane Epithelial Antigen of the Prostate 1 (STEAP1) is a transmembrane protein frequently overexpressed in prostate cancer (PC). Antibody-based therapies targeting STEAP1 have shown promising preclinical and clinical outcomes. Despite its widespread and PC-specific overexpression, the molecular regulation and biological function of STEAP1 remain poorly understood. Elucidating the mechanisms that govern STEAP1 expression and its role in PC progression could provide valuable insights into disease mechanisms, improve patient selection for STEAP1-targeted therapies, and inform combination or sequential treatment strategies. Methods: To investigate STEAP1 regulation and function, we first compared its expression to established PC driver genes and observed a correlation between STEAP1 and the androgen receptor (AR) as well as several AR-regulated genes. Using pathway analyses in the Decipher (localized prostate cancer) and West Coast Dream Team (WCDT, metastatic prostate cancer) clinical cohorts, we examined dysregulated pathways in patients with high STEAP1 expression. Additionally, we performed a FACS-based whole-genome CRISPR interference (CRISPRi) screen in C4-2B cells using an anti-STEAP1 antibody to identify regulators by comparing STEAP1-high and STEAP1-low expressing subsets. Results: In the WCDT dataset, STEAP1 was highly expressed in AR-positive metastatic prostate cancer subtypes, irrespective of neuroendocrine differentiation, but was significantly lower in AR-negative subtypes. STEAP1 expression strongly correlated with several AR-regulated genes, including STEAP2, KLK2, NKX3-1, KLK3, ARHGAP6, SLC45A3, and PMEPA1. Pathway analyses in both localized and metastatic PC cohorts revealed dysregulated metabolic pathways, particularly lipid metabolism, in STEAP1-high patients. The CRISPRi screen identified several potential targetable regulators of STEAP1, including MYC, SWI/SNF chromatin remodeling complexes. Conclusions: Our study establishes a strong association between STEAP1 and AR-driven metastatic castration-resistant prostate cancer subtypes. Furthermore, patient data suggest that STEAP1 is linked to dysregulated metabolic pathways, particularly those involving lipid metabolism. The identification of STEAP1 regulators through the CRISPRi screen provides valuable insights into its regulation and biological function, offering potential avenues for developing combination therapies, strategies to prevent treatment resistance, and improved patient stratification for STEAP1-targeted treatments.