title: Human liver CYP2B6-catalyzed hydroxylation of RP 73401.
abstract: RP 73401 is a potent inhibitor of cyclic nucleotide phosphodiesterase type IV. RP 73401 is metabolized by human liver microsomes almost exclusively by transhydroxylation of the cyclopentyl group to RPR 113406. Liquid chromatography/mass spectrometry/mass spectrometry analysis of plasma from patients given RP 73401 also revealed a molecular ion and fragmentation consistent with RPR 113406. Thus, the objective was to determine the oxidative enzyme(s) responsible for RP 73401 hydroxylation. Kinetic constants of RP 113406 formation ranged from 8 to 26 MM and 0.83 to 5.99 nmol/min/mg protein for K(m) and V(max), respectively (n = 3). Enzyme activity varied 23-fold among 15 human liver microsome samples and correlated with CYP2A6-catalyzed coumarin hydroxylase (r2 = 0.85, P < .01) and CYP2B6-catalyzed 7-ethoxytrifluoromethylcoumarin O-deethylase (r2 = 0.82, P < .01) activities. Chemical inhibition studies showed a 63% decrease in RP 73401 hydroxylation by 500 microM orphenadrine. Coumarin (10 microM), however, did not inhibit RP 73401 hydroxylation. Also, anti-CYP2B1 IgG produced 85% inhibition of RP 73401 hydroxylation, but only a negligible decline in coumarin hydroxylase activity. Of the 10 expressed P450 forms studied, only CYP2B6 catalyzed RP 73401 hydroxylation. Finally, expressed CYP2B6 showed a high affinity (K(m) = 22.5 microM) for RP 73401 hydroxylation, similar to the human liver microsome studies.
authors.affiliation: Department of Drug Metabolism and Pharmacokinetics, Rhône-Poulenc Rorer, Collegeville, Pennsylvania 19426, USA.
authors.forename: J C
authors.initials: JC
authors.lastname: Stevens
authors.forename: R B
authors.initials: RB
authors.lastname: White
authors.forename: S H
authors.initials: SH
authors.lastname: Hsu
authors.forename: M
authors.initials: M
authors.lastname: Martinet
mesh_terms: D001189:Aryl Hydrocarbon Hydroxylases; D001549:Benzamides / Q000378:metabolism*; D065702:Cytochrome P-450 CYP2B6; D003577:Cytochrome P-450 Enzyme System / Q000502:physiology*; D006801:Humans; D006900:Hydroxylation; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D010089:Oxidoreductases, N-Demethylating / Q000502:physiology*; D010726:Phosphodiesterase Inhibitors / Q000378:metabolism*; D011725:Pyridines / Q000378:metabolism*
chemical_list: D001549:Benzamides; D010726:Phosphodiesterase Inhibitors; D011725:Pyridines; D003577:Cytochrome P-450 Enzyme System; D001189:Aryl Hydrocarbon Hydroxylases; C585599:CYP2B6 protein, human; D065702:Cytochrome P-450 CYP2B6; D010089:Oxidoreductases, N-Demethylating; C087566:piclamilast