Time-dependent enzyme inactivation: Numerical analyses of in vitro data and prediction of drug-drug interactions

Abstract

Cytochrome P450 (CYP) enzyme kinetics often do not conform to Michaelis-Menten assumptions, and time-dependent inactivation (TDI) of CYPs displays complexities such as multiple substrate binding, partial inactivation, quasi-irreversible inactivation, and sequential metabolism. Additionally, in vitro experimental issues such as lipid partitioning, enzyme concentrations, and inactivator depletion can further complicate the parameterization of in vitro TDI. The traditional replot method used to analyze in vitro TDI datasets is unable to handle complexities in CYP kinetics, and numerical approaches using ordinary differential equations of the kinetic schemes offer several advantages. Improvement in the parameterization of CYP in vitro kinetics has the potential to improve prediction of clinical drug-drug interactions (DDIs). This manuscript discusses various complexities in TDI kinetics of CYPs, and numerical approaches to model these complexities. The extrapolation of CYP in vitro TDI parameters to predict in vivo DDIs with static and dynamic modeling is discussed, along with a discussion on current gaps in knowledge and future directions to improve the prediction of DDI with in vitro data for CYP catalyzed drug metabolism.

Static and Dynamic Projections of Drug-Drug Interactions Caused by Cytochrome P450 3A Time-Dependent Inhibitors Measured in Human Liver Microsomes and Hepatocytes

“…The catalytic cycle of P450 activity is complex and possesses steps wherein the process can be uncoupled resulting in generation of reactive oxygen species without catalyzing metabolism of the substrate, and alteration of this cycle in one system vs. the other could yield differences in enzyme inactivation. Furthermore, it has been proposed that there are multiple phases of inactivation in TDI experiments and that a more sophisticated method of deriving in vitro values from these experiments is needed as compared to the manner in which the present data were processed (Yadav et al, 2018;Yadav et al, 2020). Preliminary data has been generated that shows that not only are k inact values different in the two systems but that partition coefficients also differ (data on file).…”

Mechanisms of Herb-Drug Interactions Involving Cinnamon and CYP2A6: Focus on Time-Dependent Inhibition by Cinnamaldehyde and 2-Methoxycinnamaldehyde

“…As previously mentioned, curvature in the PRA plots indicates either MIC formation or partial inactivation (Nagar et al, 2014). Curved PRA plots are also observed in cases of inhibitor depletion (Yadav et al, 2020). Several models were developed to capture the inactivation kinetics of CA.…”

“…The concave upward curvature is indicative of either quasi-irreversible or partial inactivation as shown previously (Nagar et al, 2014) Using the numerical method (Korzekwa et al, 2014;Nagar et al, 2014), kinetic models were fit to the data, and kinetic parameters were estimated. The initial estimates of the rate constants were obtained from analyzing the data as detailed in previous publications (Korzekwa et al, 2014;Nagar et al, 2014;Barnaba et al, 2016;Yadav et al, 2018;Yadav et al, 2020). Association rate constants were fixed at 270 μM -1 min −1 (Barnaba et al, 2016).…”

“…Numerical analysis is a more recently developed approach for evaluating TDIs for DDI predictions (Yadav et al, 2020). It can account for diverse mechanistic complexities associated with P450 TDI and other factors (lipid partitioning).…”

Site-Directed Mutagenesis at the Molybdenum Pterin Cofactor Site of the Human Aldehyde Oxidase: Interrogating the Kinetic Differences Between Human and Cynomolgus Monkey

“…These intermediates could leave the reductive site before a complete nitro reduction into ADNTN occurs. While these compounds are reactive intermediates posing a risk of time-dependent inhibition (TDI) (Yadav et al, 2020), there is no correlation between the biphasic kinetics for O6BG in hAO and TDI since such nonlinearity does not exist This article has not been copyedited and formatted. The final version may differ from this version.…”