Characterization of testosterone 11 beta-hydroxylation catalyzed by human liver microsomal cytochromes P450.
A combination of accelerator mass spectrometry (AMS) and liquid chromatography-tandem mass spectrometry has been used to clarify some new aspects of testosterone metabolism. The main pathway of testosterone oxidative metabolism by human liver microsomes is the formation of 1beta-, 2alpha-/beta-, 6beta-, 15beta-, and 16beta-hydroxytestosterones, mainly catalyzed by cytochromes P450 2C9, 2C19, and 3A4. We now report the first determination that 11beta-hydroxytestosterone (11beta-OHT) can also be formed by human liver microsomal fractions. The structures of five hydroxylated metabolites of testosterone (2beta-, 6beta-, 11beta-, 15beta-, and 16beta-OHT) and the C-17 oxidative metabolite androstenedione were determined by liquid chromatography with UV detection at 240 nm and liquid chromatography-tandem mass spectrometry. Corresponding results were obtained by high-performance liquid chromatography-AMS analysis of incubations of [4-14C]testosterone with human liver microsomes. 6beta-Hydroxylation was always the dominant metabolic pathway, but 2beta-, 15beta-, and 16beta-OHT, and androstenedione were also formed. The previously undetected hydroxytestosterone, 11beta-OHT, was found to be a minor metabolite formed by human liver microsomal enzymes. It was formed more readily by CYP3A4 than by either CYP2C9 or CYP2C19. 11beta-Hydroxylation was inhibited by ketoconazole (IC50 = 30 nM) at concentrations similar to the IC50 (36 nM) for 6beta-hydroxylation Therefore, CYP3A4 could be mainly responsible for testosterone 11beta-hydroxylation in the human liver. These findings identify human hepatic biotransformation of testosterone to 11beta-OHT as a previously unrecognized extra-adrenal metabolic pathway.
Choi
Man Ho
MH
Biological Engineering Division, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139, USA.
Skipper
Paul L
PL
Wishnok
John S
JS
Tannenbaum
Steven R
SR
D002384:Catalysis / Q000187:drug effects; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006900:Hydroxylation / Q000187:drug effects; D006915:Hydroxytestosterones / Q000737:chemistry / Q000378:metabolism*; D007654:Ketoconazole / Q000494:pharmacology; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*
D006915:Hydroxytestosterones; D003577:Cytochrome P-450 Enzyme System; D007654:Ketoconazole