Hsu_2026_Toxicol.Lett_422_113147

The soluble epoxide hydrolase (sEH) is widely studied for its therapeutic potential in inflammation, cardiovascular disease, and pain through stabilization of endogenous epoxyeicosatrienoic acid (EET). Here, we evaluated the impact of several urea-type sEH inhibitors on steroid hormone synthesis; another pathway in which sEH may be involved. Steroidogenesis assays employing the adrenocortical carcinoma cell line H295R demonstrated that TPPU and several other urea-type sEH inhibitors induce dose-dependent suppression of multiple steroid hormones without cytotoxicity, whereas the amide-based GSK2256294A had no effect on steroid production or cell survival. Substrate-to-product ratio analysis suggested selective modulation of CYP17, CYP21, and 17beta-hydroxysteroid dehydrogenase activities, with only partial transcriptional changes in select steroidogenic genes. Exogenous 11,12-epoxyeicosatrienoic acid (EET) did not reproduce these effects, and TPPU maintained suppression under dbcAMP stimulation, indicating an off-target mechanism downstream of cAMP signaling. Collectively, these findings reveal that urea-based sEH inhibitors suppress adrenal steroidogenesis via scaffold-dependent, off-target mechanisms independent of sEH. In addition, it suggests that sEH is not involved directly in steroid metabolism.