Study of Human P450 Enzymes in Drug Oxidations using Recombinant P450s and Human Liver Microsomes

Abstract

Multiple forms cytochrome P450 (P450 or CYP) enzymes play important roles in the oxidation of structurally diverse xenobiotics and endobiotics. P450s are not self-sufficient enzymes and require a reductase as an electron carrier to function as monooxygenases. Recently recombinant P450 enzymes from different sources, e.g., microsomes of human lymphoblastoid cells, of yeast, and insect cells infected with baculovirus systems, and Escherichia coli membranes containing P450 and reductase coexpressed, have been widely used for drug metabolism research. However, the marker activities or kinetic parameters of human P450 enzymes reported from different research laboratories and commercial enzymes manufacturers are not always similar. These differences may be a critical factor for understanding roles of human P450 enzymes involved in drug metabolism. Troglitazone, a new oral antidiabetic drug, is reported not to be oxidized by P450 enzymes. However, cDNA-expressed CYP2C8 and CYP3A4 were active in catalyzing formation of a quinonetype metabolite. Intensity of inhibitory effects of specific P450 inhibitors and anti-P450 antibodies on the quinone-type metabolite formation depended on human liver samples and their P450 status. Azelastine, an antiallergy drug, was N-demethylated by CYP2D6, CYP3A4 and CYP1A2 in human liver microsomes biphasically. Human intrinsic clearance was predictable from P450 contents or kinetic parameters using cDNA-expressed P450 enzymes. The results suggest that different P450 enzymes in human liver have major roles in quinone-type metabolite formation from troglitazone and azelastine N-demethylation and that the hepatic contents of these P450 forms determine which P450 enzymes play the major roles in drug metabolism in individual humans.