Abstract
Human hepatocytes are a physiologically relevant tool useful in evaluating liver-related pharmacokinetics, including non-cytochrome P-450 (CYP) metabolism, due to their broad spectrum of metabolic enzyme activity. To verify the usefulness of human hepatocytes in evaluating non-CYP metabolism for drug discovery, we compared intrinsic clearance values (CLint) in freshly isolated and cryopreserved hepatocytes using 14 compounds primarily metabolized by non-CYP enzymes, including UDP-glucuronosyltransferase, carbonyl/aldo-keto reductase, aldehyde oxidase, flavin-containing monooxygenase, and monoamineoxidase. Cryopreservation resulted in a >20% reduction (maximum: 50%) in CLint in 7/14 compounds (statistically significant at 5 compounds) on comparing CLint values in freshly isolated and cryopreserved hepatocytes from the same donors (n = 4). However, the number of compounds with >20% CLint reduction decreased to 3 on comparing average of CLint values including un-matched donors (dolasetron: −27%, naltorexone: −32%, and phthalazine: −48%; statistically significant at phthalazine, n = 6-11). These findings suggest that fresh hepatocytes are useful in evaluating intact non-CYP enzyme activities. However, we must note that the reduction in CLint by cryopreservation could be rendered negligible if high-activity lots are selected for assay. We therefore recommend using cryopreserved hepatocytes for large-scale screening for non-CYP metabolism in drug discovery research considering the advantages in usability with cryopreserved hepatocytes.
