title: Human cytochrome P450 3A4-catalyzed testosterone 6 beta-hydroxylation and erythromycin N-demethylation. Competition during catalysis.
abstract: Cytochrome P450 3A4 is known to catalyze the metabolism of both endogenous substrates (such as the 6 beta-hydroxylation of testosterone) and many important therapeutic agents, including the N-demethylation of erythromycin. However, erythromycin and testosterone have been reported to have little or no effect on the metabolism of each other by recombinant CYP3A4. In an effort to understand the basis of these observations, we studied the N-demethylation of erythromycin and the 6 beta-hydroxylation of testosterone in human liver microsomes and in microsomes from cells containing recombinant human CYP3A4 and P450 reductase under a variety of experimental conditions. In both human liver microsomal and recombinant CYP3A4 systems, erythromycin inhibited testosterone 6 beta-hydroxylation in a concentration dependent manner, and vice versa. However, the inhibition mechanism was complex. At low substrate concentrations, testosterone and erythromycin acted as competitive inhibitors to each other. Under these experimental conditions, an apparent competitive inhibition of testosterone 6 beta-hydroxylation by erythromycin was observed, with Ki values similar to that of the K(m) values for erythromycin. When the rates of testosterone 6 beta-hydroxylation and erythromycin N-demethylation were determined in microsomal incubations containing both substrates at lower concentrations, the observed rates for each reaction were in good agreement with the calculated rates based on the rate equation describing simultaneous metabolism of two substrates by a single enzyme. However, at high substrate concentrations, the kinetic results could be best explained by a mechanism involving partial competitive inhibition. We conclude from these studies that testosterone and erythromycin mutually inhibit the metabolism of each other, consistent with the fact that CYP 3A4 catalyzes the metabolism of both substrates.
authors.affiliation: Department of Drug Metabolism, Merck Research Laboratories, Rahway, NJ 07065, USA.
authors.forename: R W
authors.initials: RW
authors.lastname: Wang
authors.forename: D J
authors.initials: DJ
authors.lastname: Newton
authors.forename: T D
authors.initials: TD
authors.lastname: Scheri
authors.forename: A Y
authors.initials: AY
authors.lastname: Lu
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases; D001667:Binding, Competitive; D002384:Catalysis; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006801:Humans; D006900:Hydroxylation; D007700:Kinetics; D008745:Methylation; D008862:Microsomes, Liver / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D010089:Oxidoreductases, N-Demethylating; D013250:Steroid Hydroxylases / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C022657:steroid hormone 6-beta-hydroxylase; C510163:CYP3A4 protein, human; D010089:Oxidoreductases, N-Demethylating