PT  - JOURNAL ARTICLE
AU  - Haishan Li
AU  - Tao Chen
AU  - John Cottrell
AU  - Hongbing Wang
TI  - Nuclear Translocation of Adenoviral-Enhanced Yellow Fluorescent Protein-Tagged-Human Constitutive Androstane Receptor (hCAR): A Novel Tool for Screening hCAR Activators in Human Primary Hepatocytes
AID  - 10.1124/dmd.108.026005
DP  - 2009 May 01
TA  - Drug Metabolism and Disposition
PG  - 1098--1106
VI  - 37
IP  - 5
4099  - http://dmd.aspetjournals.org/content/37/5/1098.short
4100  - http://dmd.aspetjournals.org/content/37/5/1098.full
SO  - Drug Metab Dispos2009 May 01; 37
AB  - The constitutive androstane receptor [(CAR) NR1I3] is a hepatic transcription factor that controls the expression of numerous drug-metabolizing enzymes and transporters in response to xenobiotic exposures. In primary hepatocytes and intact liver, CAR resides in the cytoplasm under basal condition and translocates to the nucleus upon exposure to inducers. However, CAR spontaneously accumulates in the nucleus of immortalized cell lines and exhibits constitutive activation in the absence of activators, which makes the identification of CAR activators extremely challenging. Here, we have established an efficient screening method for determining the nuclear translocation of human (h) CAR in human primary hepatocytes (HPHs). Our results demonstrated that adenoviral-enhanced yellow fluorescent protein-tagged hCAR (Ad/EYFP-hCAR) infects HPHs with high efficiency, and the majority of Ad/EYFP-hCAR (>80%) is expressed in the cytoplasm of noninduced HPHs and is translocated to the nucleus in response to activators and antagonists of hCAR. Furthermore, 22 compounds including known hCAR activators, nonactivators, CYP2B inducers, and deactivators were evaluated in this system. Our results indicated that chemical-mediated Ad/EYFP-hCAR translocation in HPHs significantly correlated with hCAR activation and target gene induction. Compared with cell-based reporter assays in cell lines and in vitro ligand-binding assays, the established Ad/EYFP-hCAR translocation assay in HPHs exhibits apparent advantages such as sensitivity to chemical activators and responses to both direct and indirect hCAR activators. Thus, nuclear translocation of Ad/EYFP-hCAR in HPHs represents an efficient means for in vitro prediction of chemical-mediated hCAR nuclear accumulation. The American Society for Pharmacology and Experimental Therapeutics