Received for publication October 20, 2004.
Revised November 17, 2004.
Accepted for publication November 18, 2004.

Metabolism of alfentanil by cytochrome P4503A (CYP3A) enzymes

Abstract

The synthetic opioid alfentanil is an analgesic and an in vivo probe for hepatic and first-pass CYP3A activity. Alfentanil is a particularly useful CYP3A probe because pupil diameter change is a surrogate for plasma concentrations, thereby affording noninvasive assessment of CYP3A. Alfentanil undergoes extensive CYP3A4 metabolism via two major pathways, forming noralfentanil and N-phenylpropionamide. This investigation evaluated alfentanil metabolism in vitro to noralfentanil and N-phenylpropionamide, by expressed CYP3A5 and CYP3A7 in addition to CYP3A4, with and without co-expressed or exogenous cytochrome b₅. Effects of the CYP3A inhibitors troleandomycin and ketoconazole were also determined. Rates of noralfentanil and N-phenylpropionamide formation by CYPs 3A4 and 3A5 in the absence of b₅ were generally equivalent, although the metabolite formation ratio differed, while those by CYP3A7 were substantially less. CYPs 3A4 and 3A5 were equipotently inhibited by troleandomycin, whereas ketoconazole was an order of magnitude more potent towards CYP3A4. Cytochrome b5 qualitatively and quantitatively altered alfentanil metabolism, with b5 co-expression having a greater effect than exogenous addition. Addition or co-expression of b5 markedly stimulated the formation of both metabolites, and changed the formation of noralfentanil but not N-phenylpropionamide from apparent single-site to multi-site Michaelis-Menten kinetics. These results demonstrate that alfentanil is a substrate for CYP3A5 in addition to CYP3A4, and the effects of the CYP3A inhibitors troleandomycin and ketoconazole are CYP3A enzyme-selective. Alfentanil is one of the few CYP3A substrates which is metabolized in vitro as avidly by both CYPs 3A4 and 3A5. Polymorphic CYP3A5 expression may contribute to interindividual variability in alfentanil metabolism.

Key words: anesthetics, CYP3A, cytochrome b5, cytochrome P450 isoforms, human CYP enzymes, mass spectrometry, pharmacogenetics

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