title: Pathways of carbamazepine bioactivation in vitro. III. The role of human cytochrome P450 enzymes in the formation of 2,3-dihydroxycarbamazepine.
abstract: Conversion of the carbamazepine metabolite 3-hydroxycarbamazepine (3-OHCBZ) to the catechol 2,3-dihydroxycarbamazepine (2,3-diOHCBZ) followed by subsequent oxidation to a reactive o-quinone species has been proposed as a possible bioactivation pathway in the pathogenesis of carbamazepine-induced hypersensitivity. Initial in vitro phenotyping studies implicated CYP3A4 as a primary catalyst of 2,3-diOHCBZ formation: 2-hydroxylation of 3-OHCBZ correlated significantly (r(2) > or = 0.929, P < 0.001) with CYP3A4/5 activities in a panel of human liver microsomes (n = 14) and was markedly impaired by CYP3A inhibitors (>80%) but not by inhibitors of other cytochrome P450 enzymes (< or = 20%). However, in the presence of troleandomycin, the rate of 2,3-diOHCBZ formation correlated significantly with CYP2C19 activity (r(2) = 0.893, P < 0.001) in the panel of human liver microsomes. Studies with a panel of cDNA-expressed enzymes revealed that CYP2C19 and CYP3A4 were high (S50 = 30 microM) and low (S50 = 203 microM) affinity enzymes, respectively, for 2,3-diOHCBZ formation and suggested that CYP3A4, but not CYP2C19, might be inactivated by a metabolite formed from 3-OHCBZ. Subsequent experiments demonstrated that preincubation of 3-OHCBZ with human liver microsomes or recombinant CYP3A4 led to decreased CYP3A4 activity, which was both preincubation time- and concentration-dependent, but not inhibited by inclusion of glutathione or N-acetylcysteine. CYP3A4, CYP3A5, CYP3A7, CYP2C19, and CYP1A2 converted [14C]3-OHCBZ into protein-reactive metabolites, but CYP3A4 was the most catalytically active enzyme. The results of this study suggest that CYP3A4-dependent secondary oxidation of 3-OHCBZ represents a potential carbamazepine bioactivation pathway via formation of reactive metabolites capable of inactivating CYP3A4, potentially generating a neoantigen that may play a role in the etiology of carbamazepine-induced idiosyncratic toxicity.
authors.affiliation: Section of Developmental Pharmacology and Experimental Therapeutics, Division of Pediatric Pharmacology and Medical Toxicology, Children's Mercy Hospitals and Clinics, 2401 Gillham Road, Kansas City, MO 64108, USA. rpearce@cmh.edu
authors.forename: Robin E
authors.initials: RE
authors.lastname: Pearce
authors.forename: Wei
authors.initials: W
authors.lastname: Lu
authors.forename: Yongqiang
authors.initials: Y
authors.lastname: Wang
authors.forename: Jack P
authors.initials: JP
authors.lastname: Uetrecht
authors.forename: Maria Almira
authors.initials: MA
authors.lastname: Correia
authors.forename: J Steven
authors.initials: JS
authors.lastname: Leeder
mesh_terms: D000818:Animals; D000927:Anticonvulsants / Q000493:pharmacokinetics*; D001711:Biotransformation; D002220:Carbamazepine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000493:pharmacokinetics*; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D051379:Mice; D008862:Microsomes, Liver / Q000201:enzymology
chemical_list: C531583:2,3-dihydroxycarbamazepine; D000927:Anticonvulsants; D002220:Carbamazepine; D003577:Cytochrome P-450 Enzyme System