title: Metabolism of cyclosporin A. II. Implication of the macrolide antibiotic inducible cytochrome P-450 3c from rabbit liver microsomes.
abstract: The in vitro metabolism of cyclosporin A (CsA) was investigated by rabbit liver microsomes in order to identify the form(s) of cytochrome P-450 responsible for its biotransformation. Metabolites including monohydroxy-, N-demethylated, dihydroxy- and dihydroxy-N-demethylated derivatives were detected and quantified by HPLC from incubates of liver microsomes, CsA, and NADPH. Kinetic data indicated that monohydroxy- and N-demethylated derivatives were first generated and then served as substrates for production of dihydroxylated derivatives. Liver microsomes from phenobarbital-, beta-naphthoflavone-, triacetyloleandomycin-, erythromycin-, or rifampicin-treated and untreated rabbits were investigated, but only microsomes from animals treated with macrolide antibiotics (specific inducers of form P-450 3c) exhibited a type I binding spectrum upon CsA addition (Ks = 1.5 +/- 0.5 microM) and extensively metabolized the drug to all groups of derivatives (Km = 5.0 +/- 0.5 microM, Vmax = 1.0 +/- 0.2 nmol/mg/min). A linear correlation existed between CsA oxidase activity and P-450 3c specific content. Antibodies to P-450 3c strongly inhibited CsA oxidase activity of microsomes from macrolide antibiotic-induced animals, whereas antibodies to other forms, including P-450 2, 3b, 4, and 6, did not. When highly purified forms of P-450, including P-450 2, 3b, 3c, and 4, were assayed in a reconstituted system, only P-450 3c exhibited type I binding spectrum upon CsA addition (Ks = 1.4 +/- 0.5 microM) and extensively metabolized the drug to all derivatives. We conclude that the macrolide antibiotic-inducible form P-450 3c (or P-450 3c related from(s)) is responsible for the major part of CsA metabolism by rabbit liver microsomes.
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D003524:Cyclosporins / Q000378:metabolism*; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis* / Q000276:immunology; D004790:Enzyme Induction / Q000187:drug effects; D004917:Erythromycin / Q000494:pharmacology; D007074:Immunoglobulin G / Q000276:immunology; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D010088:Oxidoreductases / Q000032:analysis; D011817:Rabbits; D013057:Spectrum Analysis
chemical_list: D000900:Anti-Bacterial Agents; D003524:Cyclosporins; D007074:Immunoglobulin G; D004917:Erythromycin; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases