title: Progress curve analysis of CYP1A2 inhibition: a more informative approach to the assessment of mechanism-based inactivation?
abstract: Mechanism-based cytochrome P450 inactivation is defined as a time- and NADPH-dependent inactivation that is not reversible upon extensive dialysis. Current methodologies use dilution approaches to estimate the rate of inactivation and offer limited mechanistic insight and are significantly influenced by experimental conditions. We investigated the potential of progress curve analysis because this experimental design allows investigation of both the reversible (K(iapp)) and irreversible (K(i), K(inact)) components of the reaction mechanism. The human liver microsomal CYP1A2 inactivation kinetics of resveratrol, oltipraz, furafylline, and dihydralazine (Fig. 2) were evaluated. The inactivation results for furafylline (K(i), 0.8 microM; K(inact), 0.16 min(-1)) are within 2-fold to published data (K(i), 1.6 microM; K(inact), 0.19 min(-1)). Resveratrol and dihydralazine results are within a 4.3-fold range of published data, which compares well with ranges of estimates of these parameters across publications (e.g., furafylline has estimates ranging of K(i) from 1.6 to 22.3 microM and K(inact) from 0.19 to 0.87 min(-1)). This range of estimates highlights the potential caveats surrounding the existing methodologies that have been previously discussed in depth. In addition to these inactivation parameters, we have been able to demonstrate a variation in balance of reversible versus irreversible inhibition within these inactivators. Oltipraz and resveratrol have K(iapp) values similar to their K(i), indicating a significant early onset reversible inhibition, whereas furafylline and dihydralazine are dominated by irreversible inactivation. This approach allows a more mechanistic investigation of an inactivator and in the future may improve the prediction of clinical drug-drug interactions.
authors.affiliation: Department of Pharmacokinetics, Dynamics, and Metabolism (IPC 664), Pfizer Global Research and Development, Sandwich, Kent, UK. david.fairman@pfizer.com
authors.forename: D A
authors.initials: DA
authors.lastname: Fairman
authors.forename: C
authors.initials: C
authors.lastname: Collins
authors.forename: S
authors.initials: S
authors.lastname: Chapple
mesh_terms: D019388:Cytochrome P-450 CYP1A2 / Q000378:metabolism; D065609:Cytochrome P-450 CYP1A2 Inhibitors; D004078:Dihydralazine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008862:Microsomes, Liver / Q000187:drug effects* / Q000201:enzymology; D008954:Models, Biological; D009249:NADP / Q000378:metabolism; D017374:Paroxetine / Q000494:pharmacology; D011719:Pyrazines / Q000494:pharmacology; D000077185:Resveratrol; D013267:Stilbenes / Q000494:pharmacology; D013806:Theophylline / Q000031:analogs & derivatives / Q000494:pharmacology; D013871:Thiones; D013876:Thiophenes