Hepatocyte Stability Assay
Creative Animodel provides hepatocyte stability studies to help our clients understand in vitro intrinsic clearance or identify metabolites formed from new drug candidates. We guarantee our clients consistent and high-quality data with cost-efficiency that comes from a highly automated approach.
Hepatocytes Serve as “Gold Standard”
Although drug metabolism generally occurs in a variety of tissues (such as lung, skin and kidney), the liver is regarded as the predominate site of drug metabolism in humans and animals. Intact hepatocytes contain all drug metabolising enzymes in both phase I and phase II including the cytochrome P450’s (CYPs), other non-P450 enzymes, sulfo- and glucuronosyltransferases. In addition, hepatocytes are also a living system, which closely mimic the in vivo situation. Thus, hepatocytes represent a prime model system for studying drug disposition, and have been referred to as the “gold standard” in vitro system for an accurate estimation of intrinsic clearance and to elucidate metabolic routes of new chemical entities.
Figure 1. Metabolism sites of drugs.
(1) Drugs delivered by oral administration are absorbed in GI tract and then delivered to liver. (2) Liver is the major site of drug metabolism, and 90% oral medicine is metabolized in liver before reaching the systemic circulation. (3) Through subcutaneous, transdermal or intravenous administration, drugs directly enter the systemic circulation and (4) reach the target organs before hepatic modificaiton.
Standard Conditions of Hepatocyte Stability Assay
Creative Animodel provides stability assay in hepatocytes from human and various animal species including but not restricted to primate, pig, dog, rabbit, rat and mouse. Tested compounds are incubated in hepatocyte suspensions at 37℃ in duplicate, and different concentrations are available. After incubation, the supernatant is collected by centrifugation, and the parent compound is investigated by LC-MS/MS analysis at five-time points over 120 minutes (0, 15, 30, 60 and 120 min). The percentage of parent compound remaining is calculated by comparing the peak area of the parent compound at each time point to time zero. Half-life is estimated from the slope of the initial linear range of the logarithmic curve of parent compound remaining vs. time, assuming first order kinetics. The apparent intrinsic clearance is further calculated from the half-life value.
Figure 2. Hepatocyte stability data from Creative Animodel and literature. Comparison of CLint, u in cryopreserved human hepatocytes with CLint, Obs obtained using the well stirred liver model. The solid line is the line of identity (±2-fold); the dashed line is the line of regression.
Creative Animodel provides fast and cost effective hepatocyte stability assay to determine the intrinsic clearance of new chemical entities, and all the assay conditions can be tailored to your requirements. We also have a wide variety of in vitro systems for metabolic stability studies to help our clients to optimize the bioavailability and duration of drug action of therapeutic candidates thereby increasing their success rate.
1. Lecluyse EL, Alexandre E. Isolation and culture of primary hepatocytes from resected human liver tissue. Methods Mol Biol. 2010, 640:57-82.
2. Ugo Zanelli, et al. Comparison of Cryopreserved HepaRG Cells with Cryopreserved Human Hepatocytes for Prediction of Clearance for 26 Drugs. Drug Metabolism and Disposition. 2012, 40:104–110.