Abstract Bladder cancer (BLCA) is the fourth most common cancer diagnosed in American men. In the U.S., men are four times more likely to be diagnosed with bladder cancer than women. Our research group has previously identified alterations in xenobiotic metabolism in BLCA. In this study, using a metabolomics approach, we identified alterations in the arachidonic acid pathway in bladder tumors from male versus female patients. Included within this altered arachidonic acid pathway was downregulation of EPHX2, an enzyme that converts epoxyeicosatrienoic acids (EETs) to dihydroxyeicosatrienoic acids (DiHETs). Consistent with this, levels of EETs were significantly elevated in bladder tumors derived from male versus female patients. Cox proportional hazard regression analysis further revealed that reduced expression of EPHX2 was significantly associated with poor clinical outcome across multiple publicly available datasets only in male BLCA patients but not in females. Mechanistic studies revealed that EPHX2 over-expression in male-derived BLCA cell line reduced cell growth in vitro and tumor growth in vivo. Overall, these studies nominate EPHX2 as a potential tumor suppressor in male patients with BLCA. Citation Format: Mohammed Khurshidul Hassan, Roshan Borkar, Karthik Reddy Kami Reddy, Danthasinghe Waduge Badrajee Piyarathna, Chandra Shekar Amara, Allison Bellman, ChandraShekar R. Ambati, Vasanta Putluri, Abu Hena Mostafa Kamal, Roni J. Bollag, Martha K. Terris, Leomar Y. Ballester, Yair Lotan, Cristian Coarfa, Arun Sreekumar, Nagireddy Putluri. Gender-specific metabolome in bladder cancer: Role of EPHX2 in bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2379.