title: In vitro characterization of the cytochrome P450 isoforms involved in the metabolism of 6-methoxy-2-napthylacetic acid, an active metabolite of the prodrug nabumetone.
abstract: The cytochrome P450 (CYP) isoforms that catalyze the oxidation metabolism of 6-methoxy-2-napthylacetic acid (6-MNA), an active metabolite of nabumetone, were studied in rats and humans. Using an extractive reversed-phase HPLC assay with fluorescence detection, monophasic Michaelis-Menten kinetics was obtained for the formation of 6-hydroxy-2-naphthylacetic acid (6-HNA) in liver microsomes of rats and humans, and kinetic analysis showed that the K(m) and V(max) values for the formation of 6-HNA in humans and rats were 640.0 ± 30.9 and 722.9 ± 111.7 µM, and 1167.5 ± 33.0 and 1312.7 ± 73.8 pmol min⁻¹ mg protein⁻¹, respectively. The CYPs responsible for metabolism of 6-MNA in liver microsomes of rats and humans were identified using correlation study, recombinant CYP supersomes, and specific CYP inhibitors and antibodies. Recombinant human CYP2C9 exhibited appreciable catalytic activity with respect to 6-HNA formation from 6-MNA. Among 14 recombinant rat CYPs examined, CYP2C6, CYP2C11 and CYP1A2 were involved in the metabolism of 6-MNA. Sulfaphenazole (a selective inhibitor of CYP2C9) inhibited the formation of 6-HNA in pooled human microsomes by 89%, but failed to inhibit this reaction in rat liver microsomes. The treatment of pooled human liver microsomes with an antibody against CYP2C9 inhibited the formation of 6-HNA by about 80%. The antibody against CYP2C11 suppressed the activity by 20 to 30% in rat microsomes, whereas that of CYP1A2 microsomes did not show drastic inhibition. These findings suggest that CYP2C9 has the highest catalytic activity of 6-MNA metabolism in humans. In contrast, metabolism of 6-MNA is suggested to be mediated mainly by CYP2C6 and CYP2C11 in rats.
authors.affiliation: Faculty of Pharmaceutical Sciences, Josai International University, Chiba, Japan.
authors.forename: Kaori
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Eiichi
authors.initials: E
authors.lastname: Nemoto
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Hasegawa
authors.forename: Masayuki
authors.initials: M
authors.lastname: Akimoto
authors.forename: Kenji
authors.initials: K
authors.lastname: Sugibayashi
mesh_terms: D000818:Animals; D002074:Butanones / Q000378:metabolism / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000378:metabolism*; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology; D000077430:Nabumetone; D009280:Naphthaleneacetic Acids / Q000378:metabolism*; D011355:Prodrugs / Q000378:metabolism / Q000494:pharmacology*; D051381:Rats
chemical_list: D002074:Butanones; D007527:Isoenzymes; D009280:Naphthaleneacetic Acids; D011355:Prodrugs; D003577:Cytochrome P-450 Enzyme System; C044612:6-methoxy-2-naphthylacetic acid; D000077430:Nabumetone