A Practical and Direct Comparison of Intrinsic Metabolic Clearance of Several Non-CYP Enzyme Substrates in Freshly Isolated and Cryopreserved Hepatocytes

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 dru...

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Published in:DRUG METABOLISM AND PHARMACOKINETICS 2012, Vol.27 (2), p.181-191
Main Authors: Akabane, Takafumi, Gerst, Nicolas, Naritomi, Yoichi, Masters, Jeffrey N., Tamura, Kouichi
Format: Article
Language:eng ; jpn
Subjects:
Alcohol Oxidoreductases - metabolism
Aldehyde Oxidase - metabolism
Aldehyde Reductase
Aldo-Keto Reductases
Cell Survival - physiology
Cryopreservation - methods
drug clearance
drug discovery
Drug Evaluation, Preclinical - methods
Enzymes - metabolism
Hepatocytes - enzymology
Humans
isolated hepatocytes
Metabolic Clearance Rate - physiology
metabolism
Pharmaceutical Preparations - chemistry
Pharmaceutical Preparations - metabolism
Substrate Specificity - physiology
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Summary: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 mono-oxygenase, and monoamineoxidase. Cryopreservation resulted in a >20% reduction (maximum: 50%) in CLint in 7/14 compounds (statistically significant for 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 for 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.
ISSN:1347-4367
1880-0920
DOI:10.2133/dmpk.DMPK-11-RG-097