Use of cryopreserved transiently transfected cells in high-throughput pregnane X receptor transactivation assay.
Cryopreserved, transiently transfected HepG2 cells were compared to freshly transfected HepG2 cells for use in a pregnane X receptor (PXR) transactivation assay. Assay performance was similar for both cell preparations; however, cryopreserved cells demonstrated less interassay variation. Validation with drugs of different PXR activation potencies and efficacies demonstrated an excellent correlation (r(2) > 0.95) between cryopreserved and fresh cells. Cryopreservation did not change the effect of known CYP3A4 inducers that have poor cell permeability, indicating that cryopreservation had little effect on membrane permeability. In addition, cryopreserved HepG2 cells did not exhibit enhanced susceptibility to cytotoxic compounds compared to transiently transfected control cells. The use of cryopreserved cells enables this assay to run with enhanced efficiency.
Zhu
Zhengrong
Z
Bristol-Myers Squibb Company, Wallingford, CT 06492, USA. zhengrong.zhu@bms.com
Puglisi
Jaime
J
Connors
David
D
Stewart
Jeremy
J
Herbst
John
J
Marino
Anthony
A
Sinz
Michael
M
O'Connell
Jonathan
J
Banks
Martyn
M
Dickinson
Kenneth
K
Cacace
Angela
A
D001681:Biological Assay / Q000379:methods*; D018938:Caco-2 Cells; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D045744:Cell Line, Tumor; D002463:Cell Membrane Permeability / Q000187:drug effects; D015925:Cryopreservation / Q000379:methods*; D064420:Drug-Related Side Effects and Adverse Reactions; D006801:Humans; D008113:Liver Neoplasms / Q000473:pathology; D015735:Mifepristone / Q000378:metabolism / Q000494:pharmacology; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D000077297:Pregnane X Receptor; D011987:Receptors, Steroid / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015203:Reproducibility of Results; D012293:Rifampin / Q000378:metabolism / Q000494:pharmacology; D013442:Sulfinpyrazone / Q000378:metabolism / Q000494:pharmacology; D015533:Transcriptional Activation; D014162:Transfection
D004364:Pharmaceutical Preparations; D000077297:Pregnane X Receptor; D011987:Receptors, Steroid; D015735:Mifepristone; D013442:Sulfinpyrazone; D012293:Rifampin