High throughput liquid chromatography-mass spectrometry assessment of the metabolic activity of commercially available hepatocytes from 96-well plates.
We have assessed the metabolic activity of freshly isolated and commercially preserved rat, monkey, and human primary hepatocytes in a 96 well plate format utilizing eight beta-adrenolytic drugs as model compounds. Sample introduction from 96 well plates, HPLC solvent delivery, mass spectrometric (MS) detection, and/or UV detection were fully integrated and operated unattended. After drugs were incubated with hepatocytes for three or six hr, LC-MS analyses were carried out to determine the amount of drug which was not metabolized. Two LC-MS methods were used which had a sample throughput of 4 samples/hr and 12 samples/hr. Under optimal conditions, this hepatic assay could screen 300 samples/week or 1200 samples/month. Although freshly isolated hepatocytes were more active, commercially available rat, monkey, and human primary isolated hepatocytes metabolized drug substrates in similar relative rank orders. This drug-hepatocyte assay provides useful information for prioritizing pharmaceutical leads in relative rank orders or in a high/low manner according to their resistance toward liver metabolism.
R.W. Johnson Pharmaceutical Research Institute, Spring House, PA 19477-0776, USA. gcaldwel@prius.jnj.com
G W
GW
Caldwell
J A
JA
Masucci
E
E
Chacon
D000818:Animals; D002478:Cells, Cultured; D002853:Chromatography, Liquid / Q000379:methods*; D064420:Drug-Related Side Effects and Adverse Reactions; D000882:Haplorhini; D006801:Humans; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods*; D051381:Rats; D013056:Spectrophotometry, Ultraviolet