Identification of the rabbit and human cytochromes P-450IIIA as the major enzymes involved in the N-demethylation of diltiazem.
Oxidative metabolism of diltiazem (DTZ), a calcium channel blocker, was investigated in rabbit and human liver microsomes as well as in primary cultures of human hepatocytes. DTZ N-demethylation, the major metabolic pathway in man, was strongly increased by treatment of animals, patients, and hepatocyte cultures with rifampicin and other inducers of the P-450IIIA subfamily. In a reconstituted system with purified forms of P-450 and NADPH cytochrome P-450 reductase, P-450IIIA7 exhibited the highest DTZ N-demethylase activity. In both rabbit and human liver microsomes, this activity was highly correlated with erythromycin demethylase, a characteristic substrate of P-450IIIA, or with an immunoquantitated level of P-450IIIA, and was specifically inhibited by anti-P-450IIIA7 polyclonal and monoclonal antibodies. Cyclosporin A, another specific substrate of P-450IIIA in rabbit and human, competitively inhibited DTZ N-demethylase in both species. In primary cultures of human hepatocytes treated with various inducers, including rifampicin, dexamethasone, phenobarbital, phenylbutazone or beta-naphthoflavone, the rate of release of N-demethyl-DTZ in the extracellular medium was highly correlated with the intracellular level of P-450IIIA, which appeared to be strongly induced by rifampicin and phenobarbital and to a lesser extent by dexamethasone and phenylbutazone. In aggregate, these results are consistent with the view that in both rabbit and human, cytochromes P-450 from the P-450IIIA subfamily are the major enzymes involved in the N-demethylation of DTZ. Accordingly, drugs which may be specific substrates or inducers of this P-450 are likely to influence both the side effects and the efficacy of this molecule.
Pichard
L
L
Institut National de la Santé et de la Recherche Médicale, Centre National de la Recherche Scientifique, Montpellier, France.
Gillet
G
G
Fabre
I
I
Dalet-Beluche
I
I
Bonfils
C
C
Thenot
J P
JP
Maurel
P
P
D000328:Adult; D000368:Aged; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D015153:Blotting, Western; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D003524:Cyclosporins / Q000494:pharmacology; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000378:metabolism*; D003640:Dealkylation; D004110:Diltiazem / Q000378:metabolism*; D004790:Enzyme Induction / Q000187:drug effects; D005260:Female; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000096:biosynthesis / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D006899:Mixed Function Oxygenases / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000096:biosynthesis / Q000378:metabolism; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism; D011817:Rabbits
D003524:Cyclosporins; D007527:Isoenzymes; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D001189:Aryl Hydrocarbon Hydroxylases; D051544:Cytochrome P-450 CYP3A; D010089:Oxidoreductases, N-Demethylating; D009251:NADPH-Ferrihemoprotein Reductase; D004110:Diltiazem