RT Journal Article SR Electronic T1 1-Aminobenzotriazole Coincubated with (S)-Warfarin Results in Potent Inactivation of CYP2C9 JF Drug Metabolism and Disposition JO Drug Metab Dispos FD American Society for Pharmacology and Experimental Therapeutics SP 813 OP 817 DO 10.1124/dmd.113.055913 VO 42 IS 5 A1 Jasleen K. Sodhi A1 Kevin A. Ford A1 Sophie Mukadam A1 Susan Wong A1 Cornelis E. C. A. Hop A1 S. Cyrus Khojasteh A1 Jason S. Halladay YR 2014 UL http://dmd.aspetjournals.org/content/42/5/813.abstract AB 1-Aminobenzotriazole (ABT) is a nonselective, mechanism-based inactivator of cytochrome P450 (P450) and a useful tool compound to discern P450- from non-P450-mediated metabolism. ABT effectively inactivates major human P450 isoforms, with the notable exception of CYP2C9. Here we propose that ABT preferentially binds to the warfarin-binding pocket in the CYP2C9 active-site cavity; thus, ABT bioactivation and subsequent inactivation is not favored. Therefore, coincubation with (S)-warfarin would result in displacement of ABT from the warfarin-binding pocket and subsequent binding to the active site, converting ABT into a potent inactivator of CYP2C9. To test this hypothesis, in vitro studies were conducted using various coincubation combinations of ABT and (S)-warfarin or diclofenac to modulate the effectiveness of CYP2C9 inactivation by ABT. Coincubation of ABT with (S)-warfarin (diclofenac probe substrate) resulted in potent inactivation, whereas weak inactivation was observed following coincubation of ABT with diclofenac [(S)-warfarin probe substrate]. The kinetic parameters of time-dependent inhibition of ABT for CYP2C9 in the absence and presence of (S)-warfarin (20 μM) were 0.0826 and 0.273 min−1 for kinact and 3.49 and 0.157 mM for KI, respectively. In addition, a 73.4-fold shift was observed in the in vitro potency (kinact/KI ratio), with an increase from 23.7 ml/min/mmol (ABT alone) to 1740 ml/min/mmol [ABT with (S)-warfarin (20 μM)]. These findings were supported by in silico structural modeling, which showed ABT preferentially binding to the warfarin-binding pocket and the displacement of ABT to the active site in the presence of (S)-warfarin.