title: Human cytochrome P4502B6: interindividual hepatic expression, substrate specificity, and role in procarcinogen activation.
abstract: The level of expression and interindividual variation in human hepatic microsomal cytochrome P450 (CYP) 2B6 was characterized using a polyclonal antibody (WB-2B6) raised against rat CYP2B1. Immunoblot analysis using cDNA-expressed human CYPs revealed strong cross-reactivity of this antibody with CYP2B6 (limit of detection < 0.05 pmol) and only minor cross-reactivities with human CYP2A6, CYP2D6, and CYP2E1, all of which could be resolved from CYP2B6 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Analysis of human liver microsomes using this antibody revealed immunodetectable CYP2B6 protein in a majority of individual liver samples, with levels up to 74 pmol/mg protein in the CYP2B6-positive samples. Kinetic analysis of cDNA-expressed CYPs identified many of these enzymes as catalysts of 7-ethoxy-4-trifluoromethylcoumarin (7EFC) O-deethylation, but with significantly different apparent K(M) values (CYP1A2 < CYP2B6 approximately CYP1A1 < CYP2C19 < CYP2C9 < CYP2E1 < CYP2A6). By assaying liver microsomal 7EFC O-deethylase activity at a low 7EFC concentration (5 microM) and preincubating human liver microsomes with anti-CYP1A, anti-CYP2C, and anti-CYP2E1 antibodies, we were able to monitor CYP2B6-dependent 7EFC O-deethylase activity in a panel of 17 human liver microsomes and observe a significant correlation (r2 = 0.80) between this activity and CYP2B6 protein content. The ability of CYP2B6 to activate prodrugs and procarcinogens was examined using gene locus mutation assays in CYP2B6-expressing human lymphoblast cells. CYP2B6-expressing cells were found to be more sensitive than control cells to the cytotoxicity and mutagenicity of cyclophosphamide, aflatoxin B1, and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone. CYP2B6 is thus a widely expressed human liver microsomal CYP that can contribute to a broad range of drug metabolism and procarcinogen activation reactions.
authors.affiliation: GENTEST Corporation, Woburn, MA 01801, USA.
authors.forename: E L
authors.initials: EL
authors.lastname: Code
authors.forename: C L
authors.initials: CL
authors.lastname: Crespi
authors.forename: B W
authors.initials: BW
authors.lastname: Penman
authors.forename: F J
authors.initials: FJ
authors.lastname: Gonzalez
authors.forename: T K
authors.initials: TK
authors.lastname: Chang
authors.forename: D J
authors.initials: DJ
authors.lastname: Waxman
mesh_terms: D000293:Adolescent; D000328:Adult; D000818:Animals; D000918:Antibody Specificity; D001189:Aryl Hydrocarbon Hydroxylases; D001711:Biotransformation; D002273:Carcinogens / Q000493:pharmacokinetics*; D002648:Child; D002675:Child, Preschool; D065702:Cytochrome P-450 CYP2B6; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology / Q000378:metabolism*; D018076:DNA, Complementary; D005260:Female; D006801:Humans; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D008875:Middle Aged; D010089:Oxidoreductases, N-Demethylating / Q000276:immunology / Q000378:metabolism*; D011355:Prodrugs / Q000493:pharmacokinetics*; D051381:Rats; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D002273:Carcinogens; D018076:DNA, Complementary; D011355:Prodrugs; D011994:Recombinant Proteins; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; D065702:Cytochrome P-450 CYP2B6; D010089:Oxidoreductases, N-Demethylating