PROJECT ABSTRACT Prostate cancer (PCa) is the second leading cause of cancer related death in men in the United States in 2022. A limit number of PCa cell lines and patient-derived xenograft (PDX) models hinders research to improve disease outcome. To address this unmet need, we have developed multiple PDX models, conditional reprogramed cultures (CRCs) and organoids from patients with advanced disease. AKR1C3, also named HSD17B5, is one of the most important genes involved in androgen metabolism and elevated expression of this enzyme is associated with PCa progression and failure to androgen receptor pathway inhibitors (ARPIs) treatments. We have reported that upregulation of AKR1C3, concurrent with elevated testosterone and its precursors in prostate cancer cells and xenograft tumors that are resistant to anti-androgen treatments. This proposal will use blood AKR1C3 and plasma steroid levels as biomarkers to interrogate intracrine steroidogenesis activation in ARPIs treated patients to signify drug resistance. We will also test a novel AKR1C3 inhibitor in the newly established PDX and CRC models. The inhibitor has great potential to increase efficacy of ARPIs treatments in advanced PCa which might translate into the clinical trial directly. The goal of this program is to identify novel biomarkers and develop new pharmaceutical approaches to provide co- targeting neoadjuvant with ARPIs to treat CRPC patients. Manipulation of key enzymes contributing to the production of androgen potentially offers a novel targeted therapy for the advanced prostate cancer treatment, which will have a meaningful impact on patients’ lives. The data acquired from this project will establish a clinically relevant decision making for patient treatment and pave the way to future precision medicine in prostate cancer.