Utility of in vitro systems and preclinical data for the prediction of human intestinal first-pass metabolism during drug discovery and preclinical development.

A growing awareness of the risks associated with extensive intestinal metabolism has triggered an interest in developing robust methods for its quantitative assessment. This study explored the utility of intestinal S9 fractions, human liver microsomes, and recombinant cytochromes P450 to quantify CYP3A-mediated intestinal extraction in humans for a selection of marketed drugs that are predominantly metabolized by CYP3A4. A simple competing rates model is used to estimate the fraction of drug escaping gut wall metabolism (fg) from in vitro intrinsic clearance in humans. The fg values extrapolated from the three in vitro systems used in this study, together with literature-derived fg from human intestinal microsomes, were validated against fg extracted from human in vivo pharmacokinetic (PK) profiles using a generic whole-body physiologically-based pharmacokinetic (PBPK) model. The utility of the rat as a model for human CYP3A-mediated intestinal metabolism was also evaluated. Human fg from PBPK compares well with that from the grapefruit juice method, justifying its use for the evaluation of human in vitro systems. Predictive performance of all human in vitro systems was comparable [root mean square error (RMSE) = 0.22-0.27; n = 10]. Rat fg derived from in vivo PK profiles using PBPK has the lowest RMSE (0.19; n = 11) for the prediction of human fg for the selected compounds, most of which have a fraction absorbed close to 1. On the basis of these evaluations, the combined use of fg from human in vitro systems and rats is recommended for the estimation of CYP3A4-mediated intestinal metabolism in lead optimization and preclinical development phases.

The right compound in the right assay at the right time: an integrated discovery DMPK strategy.

The high rate of attrition during drug development and its associated high research and development (R&D) cost have put pressure on pharmaceutical companies to ensure that candidate drugs going to clinical testing have the appropriate quality such that the biological hypothesis could be evaluated. To help achieve this ambition, drug metabolism and pharmacokinetic (DMPK) science and increasing investment have been deployed earlier in the R&D process. To gain maximum return on investment, it is essential that DMPK concepts are both appropriately integrated into the compound design process and that compound selection is focused on accurate prediction of likely outcomes in patients. This article describes key principles that underpin the contribution of DMPK science for small-molecule research based on 15 years of discovery support in a major pharmaceutical company. It does not aim to describe the breadth and depth of DMPK science, but more the practical application for decision making in real-world situations.

In vitro and in silico identification and characterization of thiabendazole as a mechanism-based inhibitor of CYP1A2 and simulation of possible pharmacokinetic drug-drug interactions.

Thiabendazole (TBZ) and its major metabolite 5-hydroxythiabendazole (5OH-TBZ) were screened for potential time-dependent inhibition (TDI) against CYP1A2. Screen assays were carried out in the absence and presence of NADPH. TDI was observed with both compounds, with k(inact) and K(I) values of 0.08 and 0.02 min(-1) and 1.4 and 63.3 microM for TBZ and 5OH-TBZ, respectively. Enzyme inactivation was time-, concentration-, and NADPH-dependent. Inactivation by TBZ was irreversible by dialysis and oxidation by potassium ferricyanide, and there was no protection by glutathione. 5OH-TBZ was a weak TDI of CYP1A2, and enzyme activity was recovered by dialysis. IC(50) determination of TBZ and 5OH-TBZ showed both compounds to be potent inhibitors, with IC(50) values of 0.83 and 13.05 microM, respectively. IC(50) shift studies also demonstrated that TBZ was a TDI of CYP1A2. In silico methods identified the thiazole group as a TDI fragment and predicted it as the site of metabolism. The observation pointed to epoxidation of the thiazole and the benzyl rings of TBZ as possible routes of metabolism and mechanisms of TDI. Drug-drug interaction (DDI) simulation studies using SimCyp showed good predictions for competitive inhibition. However, predictions for mechanism-based inhibition (MBI)-based DDI were not in agreement with clinical observations. There was no TBZ accumulation upon chronic administration of the drug. The in vitro MBI findings might therefore not be capturing the in vivo situation in which the proposed bioactivation route is minor. This might be the case for TBZ in which, in vivo, UDP glucuronosyltransferases and sulfanotransferase metabolize and eliminate the 5OH-TBZ.