title: Expression of the human CYP3A4 gene in the small intestine of transgenic mice: in vitro metabolism and pharmacokinetics of midazolam.
abstract: Human cytochrome P450 3A4 (CYP3A4) is the most abundant hepatic and intestinal phase I drug-metabolizing enzyme, and participates in the oxidative metabolism of approximately 50% of drugs on the market. In the present study, a transgenic-CYP3A4 (Tg-CYP3A4) mouse model that expresses CYP3A4 in the intestine and is phenotypically normal was generated, which was genotyped by both polymerase chain reaction and Southern blotting. Intestinal microsomes prepared from Tg-CYP3A4 mice metabolized midazolam (MDZ) to 1'-hydroxymidazolam about 2 times, and to 4-hydroxymidazolam around 3 times faster than that from wild-type (WT) mice. These increased MDZ hydroxylation activities were completely inhibited by an anti-CYP3A4 monoclonal antibody. The time course of plasma MDZ and its metabolite concentrations was measured after intravenous (0.25 mg/kg) and oral (2.5 mg/kg) administration of MDZ, and pharmacokinetic parameters were estimated by fitting to a noncompartmental model. Pretreatment with ketoconazole increased orally dosed MDZ maximum plasma concentration (C(max)), time of the maximum concentration, area under the plasma concentration-time curve from zero to infinity (AUC(0- infinity)), and elimination half-life (t(1/2)) to 3.2-, 1.7-, 7.7-, 2-fold, and decreased MDZ apparent oral clearance about 8-fold in Tg-CYP3A4 mice. The ratios of MDZ C(max), AUC(0- infinity), t(1/2) and bioavailability between Tg-CYP3A4 and WT mice after the oral dose of MDZ were 0.3, 0.6, 0.5, and 0.5, respectively. These results suggest that this Tg-CYP3A4 mouse would be an appropriate in vivo animal model for the evaluation of human intestine CYP3A4 metabolism of drug candidates and potential food-drug and drug-drug interactions in preclinical drug development.
authors.affiliation: Laboratory of Metabolism, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Camille P
authors.initials: CP
authors.lastname: Granvil
authors.forename: Ai-Ming
authors.initials: AM
authors.lastname: Yu
authors.forename: Guillermo
authors.initials: G
authors.lastname: Elizondo
authors.forename: Taro E
authors.initials: TE
authors.lastname: Akiyama
authors.forename: Connie
authors.initials: C
authors.lastname: Cheung
authors.forename: Lionel
authors.initials: L
authors.lastname: Feigenbaum
authors.forename: Kristopher W
authors.initials: KW
authors.lastname: Krausz
authors.forename: Frank J
authors.initials: FJ
authors.lastname: Gonzalez
mesh_terms: D000284:Administration, Oral; D000818:Animals; D019540:Area Under Curve; D001682:Biological Availability; D015139:Blotting, Southern; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006207:Half-Life; D006801:Humans; D066298:In Vitro Techniques; D007421:Intestine, Small / Q000378:metabolism*; D007654:Ketoconazole / Q000494:pharmacology; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008861:Microsomes / Q000187:drug effects / Q000378:metabolism; D008874:Midazolam / Q000031:analogs & derivatives* / Q000097:blood / Q000378:metabolism / Q000493:pharmacokinetics*; D016133:Polymerase Chain Reaction; D012293:Rifampin / Q000494:pharmacology; D013997:Time Factors
chemical_list: D004791:Enzyme Inhibitors; C042940:4-hydroxymidazolam; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System; C040081:1-hydroxymethylmidazolam; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D008874:Midazolam; D007654:Ketoconazole; D012293:Rifampin