Received for publication August 3, 2006.
Revised October 4, 2006.
Accepted for publication October 11, 2006.

CYP2B6, CYP2D6 and CYP3A4 catalyse the primary oxidative metabolism of perhexiline enantiomers by human liver microsomes

Abstract

The cytochrome P450 (CYP)-mediated 4-monohydroxylations of the individual enantiomers of the racemic anti-anginal agent perhexiline (PHX) were investigated in human liver microsomes (HLM) to identify stereoselective differences in metabolism and to determine the contribution of the polymorphic enzyme CYP2D6 and other CYPs to the intrinsic clearance of each enantiomer. The cis-, trans1- and trans2-4-monohydroxylation rates of (+)- and (-)-PHX by human liver microsomes from three extensive (EM), two intermediate (IM) and two poor (PM) CYP2D6 metabolisers were measured with a high-performance liquid chromatography (HPLC) assay. CYP isoform-specific inhibitors, monoclonal antibodies directed against CYP isoforms and recombinant expressed human CYP enzymes were used to define the CYP isoform profile of PHX 4-monohydroxylations. The total in vitro intrinsic clearance (mean ± SD) of (+)-and (-)-PHX was 1376 ± 330 and 2475 ± 321, 230 ± 225 and 482 ± 437, and 63.4 ± 1.6 and 54.6 ± 1.2 µl/min/mg for the EM, IM and PM HLM, respectively. CYP2D6 catalyses the formation of cis-OH-(+)-PHX and trans1-OH-(+)-PHX from (+)-PHX and cis-OH-(-)-PHX from (-)-PHX with high affinity. CYP2B6 and CYP3A4 each catalyse the trans1- and trans2-4-monohydroxylation of both (+)- and (-)-PHX with low affinity. Both enantiomers of PHX are subject to significant polymorphic metabolism by CYP2D6, although this enzyme exhibits distinct stereoselectivity with respect to the conformation of metabolites and the rate at which they are formed. CYP2B6 and CYP3A4 are minor contributors to the intrinsic CYP-mediated hepatic clearance of both enantiomers of PHX, except in CYP2D6 PM.

Key words: cytochrome P450 catalyzed oxidations, enzyme kinetics, genetic polymorphism, human CYP enzymes, liver microsomes, metabolite identification, pharmacogenetics, pharmacogenomics