The metabolism of indinavir, a human immune deficiency virus (HIV) protease inhibitor, has been characterized extensively in rats and humans. All oxidative metabolites found in vivo were formed when indinavir was incubated with NADPH-fortified hepatic and intestinal microsomes obtained from rats and humans. In vitro kinetic studies revealed that Vmax/Km values (microL/min/mg protein) in rat and human liver microsomes were approximately 8- and 2-fold greater than those in the intestinal microsomes of the corresponding species (55.8 and 6.7 for the liver and intestine, respectively, in rats; 16.5 and 7.7 for the liver and intestine, respectively, in humans). However, when Vmax/Km was scaled up to intrinsic clearance (mL/min/kg body weight), hepatic intrinsic clearance was much greater than the intestinal clearance by 50- to 200-fold. These results suggest that the liver plays a much greater role in first-pass metabolism of indinavir than the intestine in both species. Consistently, ketoconazole, a selective inhibitor for CYP3A, and an anti-rat CYP3A1 antibody strongly inhibited hepatic and intestinal metabolism of indinavir in both rats and humans, suggesting the involvement of CYP3A isoforms in both organs. Oral treatment of rats with dexamethasone (50 mg/kg/day for 4 days), a potent CYP3A inducer, increased both hepatic and intestinal metabolism of indinavir by a factor of 7 and 3, respectively. Furthermore, indinavir selectively inhibited 6beta-hydroxylase activity of testosterone, a CYP3A marker activity, in rat and human liver microsomes; the interactions between testosterone and indinavir were competitive with Ki values of < 1.0 microM.