11β-Hydroxyandrostenedione, the product of androstenedione metabolism in the adrenal, is metabolized in LNCaP cells by 5α-reductase yielding 11β-hydroxy-5α-androstanedione

Abstract

11β-Hydroxyandrostenedione (11OHA4), which is unique to the adrenal, was first isolated from human adrenal tissue in the fifties. It was later shown in the sixties that 11β-hydroxytestosterone (11OHT) was also produced by the human adrenal. Attention has shifted back to these adrenal androgens once more, as improved analytical techniques have enabled more accurate detection of steroid hormones. In this paper, we investigated the origin of these metabolites as well as their subsequent metabolism and examined a possible physiological role for 11OHA4 in prostate cancer cells. In H295R cells treated with forskolin and trilostane, etomidate, a reported cytochrome P450 11β-hydroxylase (CYP11B1) inhibitor, blocked the production of corticosterone, cortisol, 11OHA4 and 11OHT. The metabolism of androstenedione and testosterone by CYP11B1 and aldosterone synthase (CYP11B2) was assayed. Androstenedione was converted by CYP11B1, while the conversion by CYP11B2 was negligible. Both enzymes readily converted testosterone. The metabolism of these 11β-hydroxylated metabolites by 11β-hydroxysteroid dehydrogenase (11βHSD) types 1 and 2 was subsequently investigated. 11βHSD2 catalyzed the conversion of both 11OHA4 and 11OHT to their respective keto-steroids, while 11βHSD1 catalyzed the conversion of 11-ketoandrostenedione and 11-ketotestosterone to their respective hydroxy-steroids in Chinese hamster ovary cells. Investigating a functional role, steroid 5α-reductase types 1 and 2 converted 11OHA4 to 11β-hydroxy-5α-androstanedione (11OH-5α-dione), identified by accurate mass detection. UPLC-MS/MS analyses of 11OHA4 metabolism in LNCaP androgen-dependent prostate cancer cells, identified the 5α-reduced metabolite as well as 11-ketoandrostenedione and 11-ketotestosterone, with the latter indicating conversion by 17β-hydroxysteroid dehydrogenase. Downstream metabolism by 11βHSD2 and by 5α-reductase may therefore indicate a physiological role for 11OHA4 and/or 11OH-5α-dione in normal and prostate cancer cells.