Development of stably transfected human and rat hepatoma cell lines for the species-specific assessment of xenobiotic response enhancer module (XREM)-dependent induction of drug metabolism

Abstract Based on our current knowledge, PXR holds a key position in the induction of a selective battery of enzymes and transporters of drug metabolism. In order to prevent serious adverse drug effects or unpredicted drug–drug interactions (DDI), it is compulsory to investigate the possible inducing potency of drugs under development. Furthermore, analysis of the inducing potency of environmental pollutants and new or manufactured chemicals is part of toxicological risk assessment. In non-transfected human HepG2 and rat H4IIE hepatoma cells, we examined the characteristics of expression of 45 genes involved in drug metabolism. A few gene products such as CYP2B6 or CYP3A4 mRNA were prominent in HepG2 cells while their major rat counterparts were, e.g., CYP2B3 or CYP3A1/3A3. Furthermore, a number of xenobiotic receptors including PXR were expressed in both cell lines. A number of genes were regulated in a cell type and species-specific manner after incubation with the prototypical PXR agonists rifampicin or dexamethasone, respectively. Then, we established cell-based reporter gene assays for screening for PXR-dependent induction of drug metabolism. HepG2 and H4IIE cells were stably transfected with a reporter gene containing PXR responsive elements (XREMs) which mediate the induction of PXR target genes such as CYP3A enzymes. With both stable cell lines the CYP inducers clotrimazole, dexamethasone, omeprazole, phenobarbital, rifampicin, as well as the drug candidate EMD 392949 and the brominated flame retardants hexabromocylododecane (HBCD) and a pentabromodiphenyl ether (pentaBDE) mixture were screened. In the human HepG2-XREM3 and rat H4IIE-XREM3 cells, clotrimazole and HBCD were found as common activators of the human and rat PXR whereas pentaBDE was more effective with the human cell system. Omeprazole and phenobarbital did not induce the rat PXR-dependent reporter gene expression in H4IIE-XREM3 cells, while a moderate increase was found in HepG2-XREM3 cells. EMD 392949 also acted as inducer in human but not in rat cells confirming in vivo observations. In summary, the established PXR-dependent in vitro system allows the simultaneous, fast, and species-specific screening of chemicals, environmental contaminants, food ingredients and drugs for CYP3A induction in cells of human and rat origin.