Kinetics of Letrozole Metabolism by Cytochrome P450 3A4: Modulation and Fitting to Non‐Michaelis Menten Models

Abstract

Variable blood plasma concentration is an issue for many drugs, including letrozole, which can contribute to toxicity or sub‐effective dosing levels. There is evidence that letrozole is primarily metabolized by CYP2A6 however, CYP2A6 genetic variation only explains a portion (about 30%) of the variability in letrozole plasma concentrations. We hypothesized that CYP3A4 may play a larger role in letrozole metabolism than the current evidence suggests due to the influence of multiple ligand binding and the observation of non‐Michaelis‐Menten kinetic plots. The kinetics of letrozole metabolism by CYP3A4 were analyzed in the presence and absence of ketoconazole, a CYP3A4 modulator, and fitted to multiple ligand binding functions as well as the Michaelis‐Menten model. Further, the time‐dependent inhibition parameters of CYP2A6‐mediated letrozole metabolism were measured in the presence of the CYP2A6‐specific inactivator, trans‐cinnamic aldehyde (tCA). While the Eadie‐Hofstee plot for CYP3A4‐mediated letrozole metabolism was ambiguous, curve fitting showed the Michaelis‐Menten model appropriately characterized metabolism in the absence of ketoconazole (KM: 142+/−23.0 μM, Vmax: 13.0+/−1.44 pmol/nmol/min), suggesting a single ligand bound to the enzyme. A biphasic model most closely modeled letrozole metabolism in the presence of ketoconazole (KM1: 4.71+/−4.55 μM KM2: 513+/−164 μM, Vmax1:0.525+/−0.392 Vmax2: 30.9+/−7.60 pmol/nmol/min) and this was also reflected in the Eadie‐Hofstee plot, which was clearly biphasic; this supports a mechanism involving multiple‐ligand binding. TDI analysis of tCA revealed less effective inhibition of CYP2A6‐mediated letrozole metabolism by tCA (KI: 136 μM, kinact: 0.019 min−1) as compared to CYP2A6‐mediated coumarin metabolism. Together, these results suggest that CYP3A4 may contribute to variability in letrozole metabolism especially in the presence of an effector molecule (i.e., ketoconazole) since the value of KM1 is much closer to the letrozole plasma concentrations observed clinically than the KM value observed in the absence of an effector.Support or Funding InformationFunding from National Institute on Drug AbuseThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.