Phase I and phase II metabolic activities are retained in liver slices from mouse, rat, dog, monkey and human after cryopreservation.
Precision-cut liver slices are described as a valuable tool for in vitro metabolism studies of potential drug candidates. Recently, some papers reported successful cryopreservation conditions for liver slices, facilitating a broader and more efficient use of the tissue (particularly of human origin). The aim of this study is to evaluate the effect of cryopreservation on both phase I and phase II metabolism in liver slices prepared from mouse, rat, dog, monkey and human, using rapid freezing in the presence of 18% DMSO. Glucuronidation and sulfation activities (phase II) in both freshly prepared and cryopreserved liver slices were determined by rapid LC-MS/MS analyses using 7-hydroxycoumarin as a marker substrate. Testosterone was used as a marker substrate for cytochrome P450 mediated drug metabolism (phase I). Although the metabolic patterns and rates varied among the different species, the phase I and phase II metabolic capacities of the liver slices were well maintained after cryopreservation. Despite the good biotransformation capacity of cryopreserved slices a decrease in viability, expressed as ATP content and LDH leakage, was observed. MTT reduction was well maintained after cryopreservation. The possibility to cryopreserve liver slices will allow a more efficient utilisation of tissue, in particular from human, but also from dog and monkey. Finally, cryopreserved liver slices from mouse, rat, dog, monkey and human with good phase I and II metabolism activities are a useful in vitro tool to compare metabolite profiles of new chemical entities between species.
Martignoni
Marcella
M
Global Drug Metabolism, Pharmacia Corporation, Viale Pasteur 10, 20014, Nerviano (MI), Italy. marcella.martignoni@pharmacia.com
Monshouwer
Mario
M
de Kanter
Ruben
R
Pezzetta
Daniele
D
Moscone
Anna
A
Grossi
Pietro
P
D000255:Adenosine Triphosphate / Q000737:chemistry; D000818:Animals; D001711:Biotransformation / Q000502:physiology*; D015925:Cryopreservation / Q000379:methods*; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000378:metabolism; D004285:Dogs; D004353:Drug Evaluation, Preclinical; D005562:Formazans / Q000378:metabolism; D014453:Glucuronosyltransferase / Q000096:biosynthesis / Q000737:chemistry; D006801:Humans; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D008252:Macaca fascicularis; D008297:Male; D051379:Mice; D051381:Rats; D017207:Rats, Sprague-Dawley; D013045:Species Specificity; D015238:Sulfotransferases / Q000096:biosynthesis / Q000737:chemistry; D013739:Testosterone / Q000378:metabolism / Q000494:pharmacology; D013778:Tetrazolium Salts / Q000378:metabolism; D014021:Tissue Preservation / Q000379:methods; D014468:Umbelliferones / Q000378:metabolism / Q000494:pharmacology
D005562:Formazans; D013778:Tetrazolium Salts; D014468:Umbelliferones; C400322:cytochrome P-450 CYP2C subfamily; C079782:MTT formazan; D013739:Testosterone; C031477:7-hydroxycoumarin; D000255:Adenosine Triphosphate; D003577:Cytochrome P-450 Enzyme System; D007770:L-Lactate Dehydrogenase; D014453:Glucuronosyltransferase; D015238:Sulfotransferases