%0 Journal Article %A Abdul Basit %A John K. Amory %A Vijaya Saradhi Mettu %A Cindy Yanfei Li %A Scott Heyward %A Parth B. Jariwala %A Matthew R. Redinbo %A Bhagwat Prasad %T Relevance of Human Aldoketoreductases and Microbial β-Glucuronidases in Testosterone Disposition %D 2023 %R 10.1124/dmd.122.000975 %J Drug Metabolism and Disposition %P DMD-AR-2022-000975 %X Testosterone exhibits high variability in pharmacokinetics and glucuronidation after oral administration. While testosterone metabolism has been studied for decades, the impact of UGT2B17 gene deletion and the role of gut bacterial β-glucuronidases on its disposition are not well characterized. We first performed an exploratory study to investigate the effect of UGT2B17 gene deletion on the global liver proteome, which revealed significant increases in proteins from multiple biological pathways. The most upregulated liver proteins were aldoketoreductases (AKR1D1, AKR1C4, AKR7A3, AKR1A1, 7-dehydrocholesterol reductase (DHCR7)) and alcohol or aldehyde dehydrogenases (ADH6, ADH1C, ALDH1A1, ALDH9A1, and ALDH5A). In vitro assays revealed that AKR1D1 and AKR1C4 inactivate testosterone to 5β-dihydrotestosterone (5β-DHT) and 3α, 5β-tetrahydrotestosterone (3α, 5β-THT), respectively. These metabolites also appeared in human hepatocytes treated with testosterone and in human serum collected after oral testosterone dosing in men. Second, we evaluated the fate of testosterone glucuronide (TG) secreted into the intestinal lumen and its potential reactivation into testosterone. Incubation of TG with purified gut microbial β-glucuronidase (GUS) enzymes and with human fecal extracts confirmed testosterone reactivation by gut bacterial enzymes. Our exploratory study suggests that testosterone is metabolized by AKRs to 5β-DHT and 3α, 5β-THT in individuals harboring a UGT2B17 deletion, and these metabolites are then eliminated through glucuronidation by another UGT isoforms, UGT2B7. Both testosterone metabolic switching and variable testosterone activation by gut microbial enzymes are important mechanisms for explaining disposition of orally administered testosterone, and appear essential to unraveling the molecular mechanisms underlying UGT2B17-associated pathophysiological conditions. Significance Statement We investigated the association of UGT2B17 gene deletion and the role of gut bacterial β-glucuronidases on testosterone disposition in vitro. The experiment revealed upregulation of AKR1D1 and AKR1C4 and their role to inactivate testosterone to 5β-dihydrotestosterone and 3α, 5β-tetrahydrotestosterone, respectively. Key gut bacterial species responsible for testosterone glucuronide activation were identified. These data are important for explaining disposition of exogenously administered testosterone and appear essential to unraveling the molecular mechanisms underlying UGT2B17-associated pathophysiological conditions. %U https://dmd.aspetjournals.org/content/dmd/early/2023/01/06/dmd.122.000975.full.pdf