Specificity of substrate and inhibitor probes for human cytochromes P450 1A1 and 1A2.

Kinetic and inhibitor studies using cDNA-expressed enzymes and human liver microsomes have characterized the specificity of a range of cytochrome P450 (CYP) 1A substrate and inhibitor probes towards the two isoforms comprising this subfamily. Expressed CYP1A1 and CYP1A2 both catalyzed the O-deethylation of phenacetin, although the apparent Km was about 4-fold lower for CYP1A2 (25 vs. 108 microM). Phenacetin O-deethylation exhibited biphasic kinetics in human liver microsomes, and the apparent Km for the high-affinity component (9 +/- 6 microM) was consistent with the involvement of CYP1A2 in this reaction. The prototypic CYP1A xenobiotic inhibitor and substrate probes alpha-naphthoflavone, ellipticine, 7-ethoxycoumarin and 7-ethoxyresorufin all inhibited CYP1A1- and CYP1A2-mediated phenacetin O-deethylation as well as the high-affinity component of human liver phenacetin O-deethylase activity. alpha-Naphthoflavone and 7-ethoxycoumarin were, however, approximately 10-fold more potent as inhibitors of CYP1A2 than CYP1A1. Other putative human CYP1A xenobiotic substrates and inhibitors, including caffeine, 5- and 8-methoxypsoralen, nifedipine, paraxanthine, propranolol and theophylline similarly inhibited CYP1A1- and 1A2-catalyzed phenacetin O-deethylation and the high-affinity human liver phenacetin O-deethylase. In contrast, the monoclonal antibody MAb 1-7-1, raised against 3-methylcholanthrene-inducible rat cytochromes 450, almost abolished CYP1A1-mediated phenacetin O-deethylation, but had no effect on human liver microsomal- or CYP1A2-catalyzed phenacetin dealkylation. Together with previous data, the results indicate that the majority of human CYP1A xenobiotic inhibitor and substrate probes are nonspecific in their recognition of CYP1A1 and CYP1A2, although selectivity is apparent for some compounds.