The use of in vitro metabolic stability for rapid selection of compounds in early discovery based on their expected hepatic extraction ratios.

PURPOSE: The in vivo hepatic extraction ratio of cynomolgus monkeys was correlated with the corresponding in vitro extraction ratios that were determined in monkey microsomal incubations. METHOD: For compounds that are eliminated mainly through liver phase I metabolism, the extraction ratio calculated from liver microsomal stability studies should correlate with their in vivo hepatic extraction ratios and also with their oral bioavailability in monkey. We used both well-stirred and parallel tube models of intrinsic clearance for the correlation. We also calculated extraction ratios for compounds within a given therapeutic area from fraction absorbed values that were estimated from the Caco-2 absorption model. RESULT: The present data show that in vitro extraction ratios in monkey microsomes are predictive of the in vivo hepatic extraction ratios in monkeys. All compounds with high extraction ratio (>70%) in vivo were successfully classified as high-extraction-ratio compounds based on the in vitro monkey microsomal stability data. From the results of this study, it appears that the parallel tube model provided a slightly better classification than the well-stirred model. CONCULUSIONS: The present method appears to be a valuable tool to rapidly screen and prioritize compounds with respect to liver first-pass metabolism in monkeys at an early phase of drug discovery.

Development of a novel in vitro model to predict hepatic clearance using fresh, cryopreserved, and sandwich-cultured hepatocytes.

Sixty-four compounds with diverse structures were used in evaluation of intrinsic clearance by various hepatocyte preparations from rats, dogs, monkeys, and humans. Intrinsic clearance (CL(int)) was calculated from the ratio of the initial amount of the test compound minus the amount remaining after 2 h of incubation and the corresponding area under the concentration versus time curve. The predictive potential of this in vitro model was tested by comparing the intrinsic clearance with in vivo clearance using linear regression analysis. In addition, the intrinsic clearance values obtained from three different types of hepatocytes (cryopreserved, fresh, and sandwich-cultured) from the same species were compared to determine the influence of preservation and culture conditions. It seems that intrinsic clearance determined from human cryopreserved hepatocyte (R(2) = 0.87) was the best predictor for the corresponding human in vivo clearance. Dog and rat hepatocyte clearances were also demonstrated to be reasonable predictors (R(2) ranged 0.68-0.85 in dogs and 0.65-0.72 in rats) for their corresponding in vivo clearances. Monkey hepatocyte clearance seems to be the worst predictor for the corresponding in vivo clearance (R(2) = 0.35-0.67). Comparison of intrinsic clearance generated from cryopreserved and fresh hepatocytes demonstrated a very good correlation in dog (R(2) = 0.82) followed by rat (R(2) = 0.77), and then by monkey (R(2) = 0.70). A similar correlation profile was shown between cryopreserved hepatocytes and sandwich-cultured hepatocytes. These results demonstrated the predictive potential of intrinsic clearance for rat, dog, and human in vivo clearance, but also showed some limitation of the approach for monkey.