The role of CYP2D6 and CYP3A4 in first-pass intestinal drug metabolism

CYP3A4 and CYP2D6 are the two predominant CYP enzymes in the human intestinal mucosa. Many therapeutically important drugs that exhibit low oral bioavailability due to extensive first-pass metabolism are substrates of CYP3A4 and CYP2D6. This thesis research has a two-fold objective: (1) compared the catalytic kinetics of CYP2D6 and CYP3A4 in microsomes isolated from human liver and small intestine, using metoprolol and terfenadine as the respective prototype substrates, and (2) evaluated the proportionate contribution of the intestinal and hepatic enzymes towards first-pass drug extraction. Kinetic studies with microsomes from select human livers (n = 6) and jejunal mucosa (n = 5) showed nearly comparable average {dollar}rm Ksb{lcub}M{rcub}{dollar} values for CYP-2D6 mediated oxidation of metoprolol. The mean total {dollar}rm Vsbmax{dollar} for {dollar}alpha{dollar}-hydroxylation and O-desmethylation of metoprolol was 14-fold higher for the liver microsomes compared to the jejunal microsomes. A comparison of different intestinal regions showed that CYP2D6 protein content and metoprolol oxidative activity were in the order of jejunum {dollar}>{dollar} duodenum {dollar}>{dollar} ileum. Pharmacokinetic prediction of in vivo hepatic and intestinal first-pass extraction ratio of metoprolol based on in vitro microsomal kinetic data suggested minimal contribution of intestinal mucosal metabolism to the first-pass effect of metoprolol. The small intestinal is not a critical site of first-pass metabolism for most CYP2D6 substrates.; A sensitive LC/MS assay for two metabolites of terfenadine; azacyclonol (AZ) and terfenadine alcohol (TOH) was developed. This allowed detail kinetic characterization of CYP3A4-mediated terfenadine metabolite formation in five human livers and five intestinal microsomes. Non-hyperbolic or biphasic saturation kinetics were observed, which can be explained by substrate inhibition. The mean kinetic parameters for TOH formation were similar in the liver and intestinal microsomes {dollar}rm(Ksb{lcub}M{rcub}sim 0.2 mu M, Ksb{lcub}i,s{rcub},{dollar} 15 {dollar}mu{dollar}M, and {dollar}rm Vsbmax sim 250{dollar} pmol/min/mg). Kinetic parameters differed between the two tissues for AZ formation. The {dollar}rm Ksb{lcub}M{rcub}{dollar} was {dollar}sim{dollar}9-fold higher in the liver (3.27 vs. 0.38 {dollar}mu{dollar}M) and {dollar}rm Vsbmax{dollar} was {dollar}sim{dollar}35% lower (132 vs. 179 pmol/min/mg). Our prediction based on the in vitro intrinsic clearances indicated on average a 96% first-pass extraction at the intestinal mucosa and a 27% extraction by the liver. Small intestine appears to be the principal site of terfenadine first-pass metabolism.