O- and N-demethylation of venlafaxine in vitro by human liver microsomes and by microsomes from cDNA-transfected cells: effect of metabolic inhibitors and SSRI antidepressants.

Abstract The biotransformation of venlafaxine (VF) into its two major metabolites, O-desmethylvenlafaxine (ODV) and N-desmethylvenlafaxine (NDV) was studied in vitro with human liver microsomes and with microsomes containing individual human cytochromes from cDNA-transfected human lymphoblastoid cells. VF was coincubated with selective cytochrome P450 (CYP) inhibitors and several selective serotonin reuptake inhibitors (SSRIs) to assess their inhibitory effect on VF metabolism. Formation rates for ODV incubated with human microsomes were consistent with Michaelis-Menten kinetics for a single-enzyme mediated reaction with substrate inhibition. Mean parameters determined by non-linear regression were: V max = 0.36 nmol/min/mg protein, K m = 41 μM, and K s 22901 μM (K s represents a constant which reflects the degree of substrate inhibition). Quinidine (QUI) was a potent inhibitor of ODV formation with a K i of 0.04 μM, and paroxetine (PX) was the most potent SSRI at inhibiting ODV formation with a mean K i value of 0.17 μM. Studies using expressed cytochromes showed that ODV was formed by CYP2C9, −2C19, and −2D6. CYP2D6 was dominant with the lowest K m , 23.2 μM, and highest intrinsic clearance (V max /K m ratio). No unique model was applicable to the formation of NDV for all four livers tested. Parameters determined by applying a single-enzyme model were V max = 2.14 nmol/min/mg protein, and K m = 2504 μM. Ketoconazole was a potent inhibitor of NDV production, although its inhibitory activity was not as great as observed with pure 3A substrates. NDV formation was also reduced by 42% by a polyclonal rabbit antibody against rat liver CYP3A1. Studies using expressed cytochromes showed that NDV was formed by CYP2C9, −2C19, and −3A4. The highest intrinsic clearance was attributable to CYP2C19 and the lowest to CYP3A4. However the high in vivo abundance of 3A isoforms will magnify the importance of this cytochrome. Fluvoxamine (FX), at a concentration of 20 μM, decreased NDV production by 46% consistent with the capacity of FX to inhibit CYP3A, 2C9, and 2C19. These results are consistent with previous studies that show CYP2D6 and −3A4 play important roles in the formation of ODV and NDV, respectively. In addition we have shown that several other CYPs have important roles in the biotransformation of VF.