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Transformation of escitalopram (S-CT), the pharmacologically active S-enantiometer of citalopram, to S-desmethyl-CT (S-DCT), and of S-DCT to S-didesmethyl-CT (S-DDCT), was studied in human liver microsomes and in expressed cytochromes (CYPs). Biotransformation of the R-enantiomer (R-CT) was studied in parallel. S-CT was transformed to S-DCT by CYP2C19 (K(m) = 69 microM), CYP2D6 (K(m) = 29 microM), and CYP3A4 (K(m) = 588 microM). After normalization for hepatic abundance, relative contributions to net intrinsic clearance were 37% for CYP2C19, 28% for CYP2D6, and 35% for CYP3A4. At 10 microM S-CT in liver microsomes, S-DCT formation was reduced to 60% of control by 1 microM ketoconazole, and to 80 to 85% of control by 5 microM quinidine or 25 microM omeprazole. S-DDCT was formed from S-DCT only by CYP2D6; incomplete inhibition by quinidine in liver microsomes indicated participation of a non-CYP pathway. Based on established index reactions, S-CT and S-DCT were negligible inhibitors (IC(50) > 100 microM) of CYP1A2, -2C9, -2C19, -2E1, and -3A, and weakly inhibited CYP2D6 (IC(50) = 70-80 microM). R-CT and its metabolites, studied using the same procedures, had properties very similar to those of the corresponding S-enantiomers. Thus S-CT, biotransformed by three CYP isoforms in parallel, is unlikely to be affected by drug interactions or genetic polymorphisms. S-CT and S-DCT are also unlikely to cause clinically important drug interactions via CYP inhibition.

The kinetics of flunitrazepam (FNTZ) N-demethylation to desmethylflunitrazepam (DM FNTZ), and 3-hydroxylation to 3-hydroxyflunitrazepam (3-OH FNTZ), were studied in human liver microsomes and in microsomes containing heterologously expressed individual human CYPs. FNTZ was N-demethylated by cDNA-expressed CYP2A6 (K(m) = 1921 microM), CYP2B6 (K(m) = 101 microM), CYP2C9 (K(m) = 50 microM), CYP2C19 (K(m) = 60 microM), and CYP3A4 (K(m) = 155 microM), and 3-hydroxylated by cDNA-expressed CYP2A6 (K(m) = 298 microM) and CYP3A4 (K(m) = 286 microM). The 3-hydroxylation pathway was predominant in liver microsomes, accounting for more than 80% of intrinsic clearance compared with the N-demethylation pathway. After adjusting for estimated relative abundance, CYP3A accounted for the majority of intrinsic clearance via both pathways. This finding was supported by chemical inhibition studies in human liver microsomes. Formation of 3-OH FNTZ was reduced to 10% or less of control values by ketoconazole (IC(50) = 0.11 microM) and ritonavir (IC(50) = 0.041 microM). Formation of DM FNTZ was inhibited to 40% of control velocity by 2.5 microM ketoconazole and to 30% of control by 2.5 microM ritonavir. Neither 3-OH FNTZ nor DM FNTZ formation was inhibited to less than 85% of control activity by alpha-naphthoflavone (CYP1A2), sulfaphenazole (CYP2C9), omeprazole (CYP2C19), or quinidine (CYP2D6). Thus, CYP-dependent FNTZ biotransformation, like that of many benzodiazepine derivatives, is mediated mainly by CYP3A. Clinical interactions of FNTZ with CYP3A inhibitors can be anticipated.

The metabolism of the antidepressant mirtazapine (MIR) was investigated in vitro using human liver microsomes (HLM) and recombinant enzymes. Mean K(m) values (+/-S.D., n = 4) were 136 (+/-44) microM for MIR-hydroxylation, 242 (+/-34) microM for N-demethylation, and 570 (+/-281) microM for N-oxidation in HLM. Based on the K(m) and V(max) values, MIR-8-hydroxylation, N-demethylation, and N-oxidation contributed 55, 35, and 10%, respectively, to MIR biotransformation in HLM at an anticipated in vivo liver MIR concentration of 2 microM. Recombinant CYP predicted a 65% contribution of CYP2D6 to MIR-hydroxylation at 2 microM MIR, decreasing to 20% at 250 microM. CYP1A2 contribution increased correspondingly from 30 to 50%. In HLM, quinidine and alpha-naphthoflavone reduced MIR-hydroxylation to 75 and 45% of control, respectively, at 250 microM MIR. A >50% contribution of CYP3A4 to MIR-N-demethylation at <1 microM MIR was indicated by recombinant enzymes. In HLM, ketoconazole (1 microM) reduced N-desmethylmirtazapine formation rates to 60% of control at 250 microM. Twenty percent of MIR-N-oxidation was accounted for by CYP3A4 at 2 microM MIR, increasing to 85% at 250 microM, while CYP1A2 contribution decreased from 80 to 15%. Ketoconazole reduced MIR-N-oxidation to 50% of control at 250 microM. MIR did not substantially inhibit CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP1E2, and CYP3A4 activity in vitro. Induction/inhibition or genetic polymorphisms of CYP2D6, CYP1A2, and CYP3A4 may affect MIR metabolism, but involvement of several enzymes in different metabolic pathways may prevent large alterations in in vivo drug clearance.

The in vitro biotransformation of bupropion to hydroxybupropion was studied in human liver microsomes and microsomes containing heterologously expressed human cytochromes P450 (CYP). The mean (+/-S.E.) K(m) in four human liver microsomes was 89 (+/-14) microM. In microsomes containing cDNA-expressed CYPs, hydroxybupropion formation was mediated only by CYP2B6 at 50 microM bupropion (K(m) 85 microM). A CYP2B6 inhibitory antibody produced more than 95% inhibition of bupropion hydroxylation in four human livers. Bupropion hydroxylation activity at 250 microM was highly correlated with S-mephenytoin N-demethylation activity (yielding nirvanol), another CYP2B6-mediated reaction, in a panel of 32 human livers (r = 0.94). The CYP2B6 content of 12 human livers highly correlated with bupropion hydroxylation activity (r = 0.96). Thus bupropion hydroxylation is mediated almost exclusively by CYP2B6 and can serve as an index reaction reflecting activity of this isoform. IC(50) values for inhibition of a CYP2D6 index reaction (dextromethorphan O-demethylation) by bupropion and hydroxybupropion were 58 and 74 microM, respectively. This suggests a low inhibitory potency versus CYP2D6, the clinical importance of which is not established. Since bupropion is frequently coadministered with other antidepressants, IC(50) values (microM) for inhibition of bupropion hydroxylation were determined as follows: paroxetine (1.6), fluvoxamine (6.1), sertraline (3.2), desmethylsertraline (19.9), fluoxetine (59.5), norfluoxetine (4.2), and nefazodone (25.4). Bupropion hydroxylation was only weakly inhibited by venlafaxine, O-desmethylvenlafaxine, citalopram, and desmethylcitalopram. The inhibition of bupropion hydroxylation in vitro by a number of newer antidepressants suggests the potential for clinical drug interactions.

Eighteen healthy volunteers (10 men and 8 women) participated in a single-dose, double-blind, three-way crossover pharmacokinetic and pharmacodynamic study. Treatment conditions were 0.25 mg of triazolam, a full-agonist benzodiazepine ligand; 10 mg of zolpidem, an imidazopyridine having relative selectivity for the type 1 benzodiazepine receptor subtype; and placebo. Weight-normalized clearance of triazolam was higher in women than in men (8.7 versus 5. 5 ml/min/kg), but the difference was not significant. In contrast, zolpidem clearance was lower in women than in men (3.5 versus 6.7 ml/min/kg, P <.06). Compared to placebo, both active medications produced significant benzodiazepine agonist-like pharmacodynamic effects: sedation, impaired psychomotor performance, impaired information recall, and increased electroencephalographic beta-amplitude. Effects of triazolam and zolpidem in general were comparable and less than 8 h in duration. There was no evidence of a substantial or consistent sex difference in pharmacodynamic effects or in the kinetic-dynamic relationship, although subtle differences could not be ruled out due to low statistical power. The complete dependence of triazolam clearance on CYP3A activity, as opposed to the mixed CYP participation in zolpidem clearance, may explain the differing sex effects on clearance of the two compounds.

The in vitro biotransformation of sildenafil to its major circulating metabolite, UK-103,320, was studied in human liver microsomes and in microsomes containing heterologously expressed human cytochromes. In human liver microsomes, the mean K(m) (+/-S.E. ) was 14.4 +/- 2.0 microM. A screen of the chemical inhibitors omeprazole (10 microM), quinidine (10 microM), sulfaphenazole (10 microM), and ketoconazole (2.5 microM) only revealed detectable inhibition with ketoconazole. Sildenafil biotransformation (36 microM) was inhibited by increasing concentrations of ketoconazole and ritonavir (IC(50) values less than 0.02 microM), which are established cytochrome P450 (CYP) 3A4 inhibitors. Using microsomes containing cDNA-expressed cytochromes, UK-103,320 formation was found to be mediated by four cytochromes: CYP3A4, -2C9, -2C19, and -2D6. Estimated relative contributions to net intrinsic clearance were 79% for CYP3A4 and 20% for CYP2C9; for CYP2C19 and -2D6, estimated contributions were less than 2%. These results demonstrate that CYP3A4 is the primary cytochrome mediating UK-103,320 formation and that drugs that inhibit CYP3A4 are likely to impair sildenafil biotransformation.