Received for publication February 27, 2006.
Revised June 13, 2006.
Accepted for publication June 14, 2006.

IN VITRO AND IN VIVO CYP3A64 INDUCTION AND INHIBITION STUDIES IN RHESUS MONKEYS: A preclinical approach for CYP3A-mediated drug interaction studies

Abstract

In this study, induction and inhibition of rhesus monkey CYP3A64 versus human CYP3A4 were characterized in vitro, and the corresponding pharmacokinetic consequences were evaluated in rhesus monkeys. In monkey hepatocytes, rifampin markedly induced CYP3A64 mRNA (EC₅₀ = 0.5 µM; E_max = 6-fold) and midazolam (MDZ) 1'-hydroxylase activity (EC₅₀ = 0.2 µM; E_max = 2-fold). Compound A, a known potent and mechanism-based inhibitor of CYP3A4, strongly inhibited the formation of 1'-hydroxy MDZ by recombinant CYP3A64 in a concentration- and time-dependent manner (K_I = 0.25 µM; k_inact = 0.4 min^-1). Similar corresponding results also were obtained with human CYP3A4 in the presence of rifampin or compound A. In rhesus monkeys, MDZ exhibited a relatively high metabolic clearance (primarily via 1'-hydroxylation followed by glucuronidation), and a low hepatic availability (F_h = 16%). Consistent with the induction of hepatic metabolism of a high clearance compound, pretreatment with rifampin (18 mg/kg p.o. for 5 days) did not significantly affect the i.v. kinetics of MDZ, but caused a pronounced reduction (~10-fold) in the systemic exposure to MDZ and consequently its F_h following intra-hepatic portal vein (i.pv.) administration of MDZ. A comparable extent of the pharmacokinetic interaction also was obtained after a 1.8 mg/kg rifampin dose. Also consistent with the in vitro CYP3A64 inhibition finding, compound A (6 mg/kg i.v.) markedly increased (10-fold) the i.pv. administered MDZ exposure. At the doses studied, plasma concentrations of rifampin or compound A reached or exceeded their respective in vitro EC₅₀ or K_I values. These findings suggest the potential applicability of the in vitro - in vivo relationship approach in rhesus monkeys for studying CYP3A-mediated interactions in humans.

Key words: CYP induction, CYP inhibition, CYP3A, cytochrome P450, drug development, drug-drug interactions, hepatocytes

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