Roles of individual human cytochrome P-450 enzymes in the bioactivation of benzo(a)pyrene, 7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene, and other dihydrodiol derivatives of polycyclic aromatic hydrocarbons

Abstract

Human liver microsomes oxidized 7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene [B(a)P-7,8-diol] to products that yield DNA adduct formation and umu gene expression in the tester system Salmonella typhimurium TA1535/pSK1002. The umu response is correlated to levels of microsomal cytochrome P-450NF (P-450NF) and nifedipine oxidation in different human liver samples used for activation, and both the (+)- and (-)-enantiomers of B(a)P-7,8-diol gave similar results in these and other assays. The microsomal umu response was inhibited by antibodies raised against P-450NF. 7,8-Benzoflavone stimulated the B(a)P-7,8-diol-dependent umu response observed with purified P-450NF and human liver and lung microsomes. Thus, P-450NF appears to be the major enzyme involved in the activation of B(a)P-7,8-diol in human liver and possibly lung. Similar results were obtained for the activation of trans-9,10-dihydroxy-9,10-dihydrobenzo(b)fluoranthene and trans-3,4-dihydroxy-3,4-dihydro-7,12-dimethylbenz(a)anthracene, compounds that are known to form highly tumorigenic diol-epoxides. The major product of the oxidation of (+)-B(a)P-7,8-diol was the cis-syn isomer of benzo(a)pyrene-7,8,9,10-tetraol[7 beta, 8 alpha, 9 beta, 10 beta-tetrahydroxy-7,8,9,10-tetrahydrobenzo(a)pyrene]. Studies on the nature of the human liver enzymes involved in the formation of B(a)P-7,8-diol [from benzo(a)pyrene] indicate that neither P-450NF, P-450PA, P-450j, P-450DB, nor P-450MP is involved. The correlation of 7,8-diol formation with phenacetin O-deethylation in a set of liver samples and the partial inhibition of the reaction by 7,8-benzoflavone and anti-rat P-450 beta NF-B suggest that the enzyme involved may be P1-450, the human ortholog of rat P-450 beta NF-B, which catalyzes both the formation of B(a)P-7,8-diol and its subsequent oxidation in tissues of polycyclic hydrocarbon-treated rats. The differential effects of inhibitors indicate that benzo(a)pyrene 3-hydroxylation, 4,5-epoxidation, and 9,10-epoxidation are catalyzed by an enzyme(s) distinct from that which forms the 7,8-epoxide. The roles of the human P-450 enzymes differ from the rodent orthologs in the paradigm for bioactivation of polycyclic hydrocarbons; further, flavones appear to have opposing effects on diol formation and further epoxidation in both human liver and lung.