title: A simultaneous assessment of CYP3A4 metabolism and induction in the DPX-2 cell line.
abstract: The DPX-2 cell line, a derivative of HepG2 cells, harbors human PXR and a luciferase-linked CYP3A4 promoter. These cells were used in a panel of cell-based assays for a parallel assessment of CYP3A4 induction, metabolism, and inhibition at the cellular level. CYP3A4 induction in the DPX-2 cell line by various agents was monitored in 96-well plates by a luciferase-based transcriptional activation assay. Of the prototypical CYP3A4 inducers examined, all exhibited elevated luciferase activity in DPX-2 cells. CYP3A4 enzyme activity in noninduced and rifampicin-induced DPX-2 cells was also assessed using Vivid fluorogenic substrates. Significantly elevated CYP3A4 activity levels (2.8-fold +/- 0.2-fold above DMSO-treated cells) were found in DPX-2 cells after 48 hours of exposure to rifampicin, but were undetectable in parental HepG2 cells. Rifampicin-induced activity levels were found to be suitable for assessing the inhibitory potential of new chemical entities in downstream CYP3A4 inhibition assays. The elevated CYP3A4 activity was inhibited 85% by 10 microM ketoconazole. In addition, a cytotoxicity assay to correct for possible toxic effects of compounds at the cellular level was applied. The comparative data obtained with a combination of the above assays suggests that the application of several independent in vitro technologies used in DPX-2 cells is the best possible strategy for the assessment of the complex phenomena of CYP3A4 induction and inhibition.
authors.affiliation: Invitrogen Corp, 501 Charmany Dr, Madison, WI 53719, USA.
authors.forename: Olga
authors.initials: O
authors.lastname: Trubetskoy
authors.forename: Bryan
authors.initials: B
authors.lastname: Marks
authors.forename: Thomas
authors.initials: T
authors.lastname: Zielinski
authors.forename: Mei-Fei
authors.initials: MF
authors.lastname: Yueh
authors.forename: Judy
authors.initials: J
authors.lastname: Raucy
mesh_terms: D006528:Carcinoma, Hepatocellular / Q000201:enzymology*; D045744:Cell Line, Tumor / Q000187:drug effects / Q000201:enzymology; D002839:Chromans / Q000494:pharmacology; D003022:Clotrimazole / Q000494:pharmacology; D019388:Cytochrome P-450 CYP1A2 / Q000096:biosynthesis / Q000235:genetics; D051544:Cytochrome P-450 CYP3A; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis*; D003907:Dexamethasone / Q000494:pharmacology; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004742:Enhancer Elements, Genetic; D004790:Enzyme Induction / Q000187:drug effects; D017930:Genes, Reporter; D005813:Genes, Synthetic; D006801:Humans; D007654:Ketoconazole / Q000494:pharmacology; D008113:Liver Neoplasms / Q000201:enzymology*; D008148:Lovastatin / Q000031:analogs & derivatives / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics; D015735:Mifepristone / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000235:genetics; D009543:Nifedipine / Q000494:pharmacology; D009853:Omeprazole / Q000494:pharmacology; D017239:Paclitaxel / Q000494:pharmacology; D010672:Phenytoin / Q000494:pharmacology; D000077297:Pregnane X Receptor; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics; D011987:Receptors, Steroid / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D012293:Rifampin / Q000494:pharmacology; D045162:Thiazolidinediones / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D000077288:Troglitazone; D014217:Troleandomycin / Q000494:pharmacology
chemical_list: D002839:Chromans; D065607:Cytochrome P-450 Enzyme Inhibitors; D009363:Neoplasm Proteins; D000077297:Pregnane X Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011987:Receptors, Steroid; D011993:Recombinant Fusion Proteins; D045162:Thiazolidinediones; C012258:mevastatin; D015735:Mifepristone; D010672:Phenytoin; D003907:Dexamethasone; D003577:Cytochrome P-450 Enzyme System; D008148:Lovastatin; D014217:Troleandomycin; D008156:Luciferases; C504573:CYP1A2 protein, human; C104464:CYP3A protein, human; D019388:Cytochrome P-450 CYP1A2; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D003022:Clotrimazole; D000077288:Troglitazone; D009543:Nifedipine; D009853:Omeprazole; D017239:Paclitaxel; D007654:Ketoconazole; D012293:Rifampin; D004121:Dimethyl Sulfoxide