Mechanism-based inactivation of CYP3A by HIV protease inhibitors

C. Steven Ernest, Stephen D. Hall, David R. Jones

Research output: Contribution to journalArticle

196 Scopus citations


Human immunodeficiency virus (HIV) protease inhibitors (PIs) are inhibitors of CYP3A enzymes, but the mechanism is poorly defined. In this study, time- and concentration-dependent decreases in activity as defined by maximum rate of inactivation (kinact) and inhibitor concentration that gives 50% maximal inactivation (Kl) of CYP3A by amprenavir, indinavir, lopinavir, nelfinavir, ritonavir, and saquinavir were quantified using testosterone 6β-hydroxylation as a marker for CYP3A activity with recombinant CYP3A4(+b5), recombinant CYP3A5, and pooled human liver microsomes (HLMs). All the PIs, except indinavir, displayed inactivation with CYP3A4(+b5) and HLMs. Ritonavir was the most potent (Kl = 0.10 and 0.17 μM) and demonstrated high kinact values (0.32 and 0.40 min-1) with both CYP3A4(+b5) and HLMs. Ritonavir was not significantly depleted by high-affinity binding with CYP3A4(+b5) and confirmed that estimation of reversible inhibition was confounded with irreversible inhibition. For CYP3A5, nelfinavir exhibited the highest k inact (0.47 min-1), but ritonavir was the most potent (Kl = 0.12 μM). Saquinavirand indinavir did not showtime- and concentration-dependent decreases in activity with CYP3A5. Spectrophototmetrically determined metabolic intermediate complex formation was observed for all of the PIs with CYP3A4(+b5), except for lopinavir and saquinavir. The addition of nucleophilic and free aldehyde trapping agents and free iron and reactive oxygen species scavengers did not prevent inactivation of CYP3A4(+b5) by ritonavir, amprenavir, or nelfinavir, but glutathione decreased the inactivation by saquinavir (17%) and catalase decreased the inactivation by lopinavir (39%). In conclusion, all the PIs exhibited mechanism-based inactivation, and predictions of the extent and time course of drug interactions with PIs could be underestimated if based solely on reversible inhibition.

Original languageEnglish (US)
Pages (from-to)583-591
Number of pages9
JournalJournal of Pharmacology and Experimental Therapeutics
Issue number2
StatePublished - Feb 1 2005

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology

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