Received for publication August 12, 2004.
Revised January 25, 2005.
Accepted for publication January 26, 2005.

DIFFERENTIAL MECHANISM-BASED INHIBITION OF CYP3A4 AND CYP3A5 BY VERAPAMIL

Abstract

The genetic basis for polymorphic expression of CYP3A5 has been recently identified but the significance of CYP3A5 expression is unclear. The purpose of this study is to quantify the capability of verapamil, a mechanism-based inhibitor of CYP3A, and its metabolites to inhibit the activities of CYP3A4 and CYP3A5, and to determine if CYP3A5 expression in human liver microsomes alters the inhibitory potency of verapamil. Testosterone 6-hydroxylation or midazolam 1'-hydroxylation was used to quantify CYP3A activity. The possibility that verapamil and its metabolites form metabolic intermediate complex (MIC) with CYP3A was assessed using dual beam spectrophotometry. Verapamil and N-desalkylverapamil (D617) were found to have little inhibitory effect on cDNA-expressed CYP3A5 activity and not form MIC with cDNA-expressed CYP3A5 as indicated by the appearance of the characteristic peak at 455nm. At 50 µM, norverapamil showed time-dependent inhibition on CYP3A5 (30 %) but to a much less extent compared to that on CYP3A4 (80 %). The estimated values of the inactivation parameters k_inact and K_I of norverapamil were 4.53 µM and 0.07 min^-1 for the cDNA expressed CYP3A5, and 10.3 µM and 0.30 min^-1 for the cDNA-expressed CYP3A4. Human liver microsomes that expressed CYP3A5 were less inhibited by both verapamil and norverapamil. The inactivation efficiency of verapamil and norverapamil were 30 times and 45 times lower, respectively, for the CYP3A5-expressing microsome compared to the CYP3A5-nonexpressing microsome. These findings indicate that the presence of variable CYP3A5/CYP3A4 expression in the liver may contribute to the interindividual variability associated with verapamil mediated drug interactions.

Key words: CYP inhibition, CYP3A, cytochrome P450, cytochrome P450 isoforms, drug interactions, drug-drug interactions, enzyme inhibitors, genotype, inhibition, mechanism-based inhibition

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