Alterations of the catalytic activities of drug-metabolizing enzymes in cultures of human liver slices
VandenBranden, M.; Wrighton, S.A.; Ekins, S.; Gillespie, J.S.; Binkley, S.N.; Ring, B.J.; Gadberry, M.G.; Mullins, D.C.; Strom, S.C.; Jensen, C.B.
Drug Metabolism and Disposition the Biological Fate of Chemicals 26(11): 1063-1068
ISSN/ISBN: 0090-9556 PMID: 9806947 Accession: 010147801
Precision-cut human liver slices are an important tool for defining the metabolism and hepatotoxicity of drug candidates early in development. Because of the frequent use of this in vitro tool, a knowledge of the catalytic activities of the drug-metabolizing enzymes during human liver slice culture is necessary. Therefore, marker catalytic activities for various cytochrome P460 (P450 or CYP) forms, as well as phase II activities (glucuronidation and sulfation of 7-hydroxycoumarin), were measured in slices from three different human livers during 96 hr in culture. Standard viability measures were found to be stable from 8 to 24 hr and then declined to 96 hr. Catalytic activities measured for the P450s were ethoxyresorufin O-deethylase (CYP1A2), coumarin 7-hydroxylase (CYP2A6), (S)-mephenytoin N-demethylase (CYP2B6), diclofenac 4'-hydroxylase (CYP2C9), (S)-mephenytoin 4'-hydroxylase (CYP2C19), bufuralol 1'-hydroxylase (CYP2D6), chlorzoxazone 6-hydroxylase (CYP2E1), and midazolam 1'-hydroxylase (CYP3A). The P460 activities decreased by approximately 20% by 4 hr and by at least 66% by 24 hr and were not measurable by 96 hr. In contrast to the phase I activities, 7-hydroxycoumarin glucuronosyltransferase activity was increased at the 8-hr time point by approximately 100% and then decreased to approximately initial values by 96 hr. The 7-hydroxycoumarin sulfotransferase activity of the slices decreased significantly more slowly than did the P450 activities. In conclusion, using conventional methods of liver slice preparation and culture, most of the metabolic capabilities of human liver slices are rapidly lost with time. Therefore, the development of culture methods for human liver slices that can improve the preservation of the drug-metabolizing capabilities may be required.