Contribution of human cytochrome P-450 isoforms to the metabolism of the simplest phenothiazine neuroleptic promazine

1 The aim of the present study was to identify human cytochrome P-450 isoforms (CYPs) involved in 5-sulphoxidation and N-demethylation of the simplest phenothiazine neuroleptic promazine in human liver. 2 The experiments were performed in the following in vitro models: (A) a study of promazine metabolism in liver microsomes-(a) correlations between the rate of promazine metabolism and the level and activity of CYPs; (b) the effect of specific inhibitors on the rate of promazine metabolism (inhibitors: CYPlA2-furafylline, CYP2D6-quinidine, CYP2A6+CYP2EI-diethyldithiocarbamic acid,, CYP2C9-sulfaphenazole, CYP2C19-ticlopidine, CYP3A4-ketoconazole), (B) promazine biotransformation by cDNA-expressed human CYPs (Supersomes 1A1, 1A2, 2A6, 2136, 2C9, 2C19, 2E1, 3A4); (C) promazine metabolism in a primary culture of human hepatocytes treated with specific inducers (rifampicin-CYP3A4, CYP2136 and CYP2C inducer, 2,3,7,8-tetrachlordibenzeno-p-dioxin (TCDD )-CYP1A1/1A2 inducer). 3 In human liver microsomes, the formation of promazine 5-sulphoxide and N-desmethylpromazine was significantly correlated with the level of CYP1A2 and ethoxyresorufin O-deethylase and acetanilide 4-hydroxylase activities, as well as with the level of CYP3A4 and cyclosporin A oxidase activity. Moreover, the formation of N-desmethylpromazine was correlated well with S-mephenytoin 4'-hydroxylation. 4 Furafylline (a CYP1A2 inhibitor) and ketoconazole (a CYP3A4 inhibitor) significantly decreased the rate of promazine 5-sulphoxidation, while furafylline and ticlopidine (a CYP2C19 inhibitor) significantly decreased the rate of promazine N-demethylation in human liver microsomes. 5 The cDNA-expressed human CYPs generated different amounts of promazine metabolites, but the rates of CYP isoforms to catalyse promazine metabolism at therapeutic concentration (10 mum) was as follows: 1A1 >2B6> 1A2>2C9> 3A4>2EI > 2A6>2D6>2C19 for 5-sulphoxidation and 2C19>2B6>1A1>1A2>2D6>3A4>2C9>2EI>2A6 for N-demethylation. The highest intrinsic clearance (V-max /K-m) was found for CYP1A subfamily, CYP3A4 and CYP2136 in the case of 5-sulphoxidation, and for CYP2C19, CYP1A subfamily and CYP2136 in the case of N-demethylation. 6 In a primary culture of human hepatocytes, TCDD (a CYP1A subfamily inducer), as well as rifampicin (mainly a CYP3A4 inducer) induced the formation of promazine 5-sulphoxide and N-desmethylpromazine. 7 Regarding the relative expression of various CYPs in human liver, the obtained results indicate that CYP1A2 and CYP3A4 are the main isoforms responsible for 5-sulphoxidation, while CYP1A2 and CYP2C 19 are the basic isoforms that catalyse N-demethylation of promazine in human liver. Of the other isoforms studied, CYP2C9 and CYP3A4 contribute to a lesser degree to promazine 5-sulphoxidation and N-demethylation, respectively. The role of CYP2A6, CYP2136, CYP2D6 and CYP2E1 in the investigated metabolic pathways of promazine seems negligible.