title: Determination of time-dependent inactivation of CYP3A4 in cryopreserved human hepatocytes and assessment of human drug-drug interactions.
abstract: Assessment of time-dependent inhibition (TDI), especially CYP3A4, is an important parameter for preclinical and clinical development. The use of human liver microsomes (HLM) is the most common in vitro matrix to assess TDI, but this often leads to an overprediction of an actual effect observed clinically. Recently, the use of human hepatocytes has been hypothesized as a more relevant and possibly predictive matrix for the assessment of CYP3A4 TDI. Our work evaluates and optimizes three different human hepatocyte assays for the assessment of CYP3A4 TDI using pooled cryopreserved human hepatocytes. Using two of the optimized methods, the time-dependent inhibition kinetic parameters (K(I) and k(inact)) for four known CYP3A4 TDI (diltiazem, erythromycin, verapamil, and troleandomycin) were determined. When comparing TDI in HLM, the K(I) values from hepatocytes were in general 4- to 13-fold higher than that in HLM, whereas the k(inact) values in human hepatocytes were similar or slightly higher or lower depending on the inhibitor. The inactivation potency (k(inact)/K(I)) for four tested CYP3A4 inactivators in human hepatocytes was generally lower than that in HLM due to either lower affinity (K(I)) or lower inactivation rate (k(inact)) or both. When drug interactions were simulated with Simcyp using either HLM or human hepatocyte data, the predictions using the kinetic parameters from human hepatocytes resulted in a much better simulated change in pharmacokinetics compared with observed clinical data.
authors.affiliation: Department of Drug Metabolism and Pharmacokinetics, Roche-Palo Alto, LLC, California, USA.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Chen
authors.forename: Liling
authors.initials: L
authors.lastname: Liu
authors.forename: Mario
authors.initials: M
authors.lastname: Monshouwer
authors.forename: Adrian J
authors.initials: AJ
authors.lastname: Fretland
mesh_terms: D015925:Cryopreservation; D051544:Cytochrome P-450 CYP3A / Q000378:metabolism*; D065692:Cytochrome P-450 CYP3A Inhibitors; D004110:Diltiazem / Q000494:pharmacology; D004347:Drug Interactions; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004917:Erythromycin / Q000494:pharmacology; D022781:Hepatocytes / Q000187:drug effects* / Q000201:enzymology*; D006801:Humans; D008862:Microsomes, Liver / Q000187:drug effects / Q000378:metabolism; D010599:Pharmacokinetics; D018570:Risk Assessment / Q000379:methods; D013997:Time Factors; D014217:Troleandomycin / Q000494:pharmacology; D014700:Verapamil / Q000494:pharmacology
chemical_list: D065692:Cytochrome P-450 CYP3A Inhibitors; D004791:Enzyme Inhibitors; D004917:Erythromycin; D014217:Troleandomycin; D014700:Verapamil; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D004110:Diltiazem