Involvement of the macrolide antibiotic inducible cytochrome P-450 LM3c in the metabolism of midazolam by microsomal fractions prepared from rabbit liver.
This report characterizes the cytochrome P-450 isozyme involved in midazolam metabolism. This study was undertaken into liver microsomal fractions prepared from untreated rabbits or animals treated with drugs known to specifically induce various cytochrome P-450 isozymes such as form LM2 by phenobarbital, LM4 and LM6 by 3-methylcholanthrene and beta-naphthoflavone, LM3a by ethyl alcohol and acetone, and LM3c by macrolide antibiotics (rifampicin, erythromycin and triacetyloleandomycin). Among this library of characterized microsomal preparations, only those obtained from macrolide antibiotic-treated rabbits exhibited a Type I binding spectrum upon addition of midazolam (Ks = 3.2-5.3 micrograms/ml; 10.6-17.5 microM) and significantly metabolized midazolam to its various hydroxylated metabolites (Km = 2.52 +/- 0.22 micrograms/ml; 8.32 +/- 0.73 microM and Vmax = 20 micrograms metabolites formed/min/mg proteins; 66 nmoles metabolites formed/min/mg proteins). The following observations further confirmed the specific involvement of the cytochrome P-450 LM3c isozyme: (i) only anti-cytochrome P-450 LM3c isozyme antibodies intensively inhibited midazolam metabolism, (ii) incubation of microsomes, prepared from TAO-treated rabbits, with midazolam in the presence of potassium ferricyanide which restored the functional cytochrome P-450 LM3c isozyme, increased midazolam metabolism to a similar extent, and (iii) in the presence of Cyclosporin A, a specific substrate of the rabbit cytochrome P-450 LM3c isozyme, midazolam metabolism was inhibited in a concentration-dependent manner. These data demonstrated that the rabbit cytochrome P-450 LM3c isozyme was predominantly involved in midazolam metabolism.
Fabre
G
G
INSERM U 278, Laboratoire Hospitalo-Universitaire de Pharmacocinétique et de Toxicocinétique, Marseille, France.
Crevat-Pisano
P
P
Dragna
S
S
Covo
J
J
Barra
Y
Y
Cano
J P
JP
D000818:Animals; D003524:Cyclosporins / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis / Q000276:immunology / Q000502:physiology*; D004790:Enzyme Induction / Q000187:drug effects; D004917:Erythromycin / Q000494:pharmacology*; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000096:biosynthesis / Q000276:immunology / Q000502:physiology*; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism*; D008874:Midazolam / Q000378:metabolism*; D011817:Rabbits; D012293:Rifampin / Q000494:pharmacology*; D014217:Troleandomycin / Q000494:pharmacology*
D003524:Cyclosporins; D007527:Isoenzymes; D004917:Erythromycin; D003577:Cytochrome P-450 Enzyme System; D014217:Troleandomycin; D008874:Midazolam; D012293:Rifampin