CYP2C19 participates in tolbutamide hydroxylation by human liver microsomes.
Tolbutamide is a sulfonylurea-type oral hypoglycemic agent whose action is terminated by hydroxylation of the tolylsulfonyl methyl moiety catalyzed by cytochrome P-450 (CYP) enzymes of the human CYP2C subfamily. Although most studies have implicated CYP2C9 as the exclusive catalyst of hepatic tolbutamide hydroxylation in humans, there is evidence that other CYP2C enzymes (e.g., CYP2C19) may also participate. To that end, we used an immunochemical approach to assess the role of individual CYP2Cs in microsomal tolbutamide metabolism. Polyclonal antibodies were raised to CYP2C9 purified from human liver, and were then back-adsorbed against recombinant CYP2C19 coupled to a solid-phase support. Western blotting revealed that the absorbed anti-human CYP2C9 preparation reacted with only recombinant CYP2C9 and the corresponding native protein in hepatic microsomes, and no longer recognized CYP2C19 and CYP2C8. Monospecific anti-CYP2C9 not only retained the ability to inhibit CYP2C9-catalyzed reactions, as evidenced by its marked (90%) inhibition of diclofenac 4'-hydroxylation by purified CYP2C9 and by human liver microsomes, but also exhibited metabolic specificity, as indicated by its negligible (<15%) inhibitory effect on S-mephenytoin 4'-hydroxylation by purified CYP2C19 or hepatic microsomes containing CYP2C19. Monospecific anti-CYP2C9 was also found to inhibit rates of tolbutamide hydroxylation by 93 +/- 4 and 78 +/- 6% in CYP2C19-deficient and CYP2C19-containing human liver microsomes, respectively. Taken together, our results indicate that both CYP2C9 and CYP2C19 are involved in tolbutamide hydroxylation by human liver microsomes, and that CYP2C19 underlies at least 14 to 22% of tolbutamide metabolism. Although expression of CYP2C19 in human liver is less than that of CYP2C9, it may play an important role in tolbutamide disposition in subjects expressing either high levels of CYP2C19 or a catalytically deficient CYP2C9 enzyme.
Wester
M R
MR
Toxicology Program, University of New Mexico, College of Pharmacy, Albuquerque, New Mexico, USA.
Lasker
J M
JM
Johnson
E F
EF
Raucy
J L
JL
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000302:isolation & purification / Q000494:pharmacology; D001189:Aryl Hydrocarbon Hydroxylases; D003429:Cross Reactions; D065731:Cytochrome P-450 CYP2C19; D065727:Cytochrome P-450 CYP2C8; D065729:Cytochrome P-450 CYP2C9; D003577:Cytochrome P-450 Enzyme System / Q000276:immunology / Q000378:metabolism*; D004008:Diclofenac / Q000378:metabolism; D006801:Humans; D006900:Hydroxylation / Q000187:drug effects; D008297:Male; D008617:Mephenytoin / Q000378:metabolism; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000276:immunology / Q000378:metabolism*; D011817:Rabbits; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000276:immunology / Q000378:metabolism; D014044:Tolbutamide / Q000378:metabolism*
D000911:Antibodies, Monoclonal; C400322:cytochrome P-450 CYP2C subfamily; D004008:Diclofenac; D003577:Cytochrome P-450 Enzyme System; D014044:Tolbutamide; D006899:Mixed Function Oxygenases; D013250:Steroid Hydroxylases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C045793:CYP2C19 protein, human; C450259:CYP2C8 protein, human; D065731:Cytochrome P-450 CYP2C19; D065727:Cytochrome P-450 CYP2C8; D039181:Steroid 16-alpha-Hydroxylase; D008617:Mephenytoin