Abstract

Irreversible CYP3A inhibition by drugs constitutes one of the major causes of inhibition-based drug interactions. We evaluated time-dependent inactivation of CYP3A in cryopreserved human hepatocytes for six structurally diverse compounds known to exhibit this property. Inactivation kinetic parameters were also determined using human liver microsomes. Except for diclofenac, which did not cause CYP3A inactivation either in microsomes or in hepatocytes at concentrations up to 100 μM, time-dependent inactivation was observed in hepatocytes for amprenavir, diltiazem, erythromycin, raloxifene, and troleandomycin. The observed inactivation potency in hepatocytes (observed IC₅₀) was compared with the potency predicted using microsomal parameters (predicted IC₅₀). Despite satisfactory prediction for troleandomycin (1.35 and 2.14 μM for the predicted and observed IC₅₀, respectively), over-prediction of inactivation was observed for raloxifene, amprenavir, and erythromycin (observed IC₅₀ values 6.2-, 55-, and 7.8-fold higher, respectively, than the predicted IC₅₀). By contrast, the observed IC₅₀ for diltiazem in hepatocytes was approximately 4-fold lower than the IC₅₀ predicted from microsomal data (under-prediction). After correcting for factors including nonspecific binding and inactivator consumption, prediction was significantly improved for raloxifene (the observed IC₅₀ then became 2-fold higher than the predicted IC₅₀) and for amprenavir to a lesser extent. A specific P-glycoprotein inhibitor, 4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)-N-[2-(3.4-dimethoxyphenyl)ethyl]-6,7-dimethoxyquinazolin-2-amine (CP-100356), modulated the observed CYP3A inactivation potency by erythromycin and troleandomycin. In summary, these studies reveal three important factors that must be considered when microsomal inactivation parameters are used to predict inhibition-based drug interactions in intact cell systems.