TY - JOUR T1 - Pathways of Carbamazepine Bioactivation in Vitro I. Characterization of Human Cytochromes P450 Responsible for the Formation of 2- and 3-Hydroxylated Metabolites JF - Drug Metabolism and Disposition JO - Drug Metab Dispos SP - 1170 LP - 1179 DO - 10.1124/dmd.30.11.1170 VL - 30 IS - 11 AU - Robin E. Pearce AU - Gangadhara R. Vakkalagadda AU - J. Steven Leeder Y1 - 2002/11/01 UR - http://dmd.aspetjournals.org/content/30/11/1170.abstract N2 - In vitro studies were conducted to identify the cytochromes P450 (P450s) involved in the formation of 2- and 3-hydroxycarbamazepine, metabolites that may serve as precursors in the formation of protein-reactive metabolites. Human liver microsomes (HLMs) converted carbamazepine (30–300 μM) to 3-hydroxycarbamazepine at rates >25 times those of 2-hydroxycarbamazepine. Both the 2- and 3-hydroxylation of carbamazepine appeared to conform to monophasic Michaelis-Menten kinetics in HLMs (apparentKm values, ∼1640 and ∼217 μM; apparentVmax values, ∼5.71 and ∼46.9 pmol/mg of protein/min, respectively). Rates of carbamazepine 2- and 3-hydroxylation correlated strongly with CYP2B6 activity (r ≥ 0.757) in a panel of HLMs (n = 8). Carbamazepine 3-hydroxylation also correlated significantly with CYP2C8 activity at a carbamazepine concentration of 30 μM. Formation of 2- and 3-hydroxycarbamazepine did not correlate significantly with any other P450 activities. The chemical inhibitors ketoconazole (CYP3A) and 7-EFC (CYP2B6) inhibited both 2- and 3-hydroxycarbamazepine formation whereas 4-methylpyrazole (CYP2E1) markedly decreased 2-hydroxycarbamazepine formation. Several recombinant P450s catalyzed carbamazepine 2- and 3-hydroxylation, but after adjustment for relative hepatic abundance, CYP3A4 and CYP2B6 appeared to be the major catalysts of carbamazepine 3-hydroxylase activity, and at least five P450s were significant contributors to 2-hydroxycarbamazepine formation; CYP2E1 made the greatest contribution to the Clint of carbamazepine 2-hydroxylation (∼30%), but P450s CYP1A2, 2A6, 2B6, and 3A4 also made significant contributions (∼13–18%). These results suggest that CYP2B6 and CYP3A4 are largely responsible for the formation of 3-hyrdoxycarbamazepine, whereas multiple P450s (CYP1A2, 2A6, 2B6, 2E1, and 3A4) contributed to 2-hydroxycarbamazepine formation. The American Society for Pharmacology and Experimental Therapeutics ER -