Format

Send to

Choose Destination
Methods Mol Biol. 2016;1473:33-42. doi: 10.1007/978-1-4939-6346-1_4.

Quantitative High-Throughput Luciferase Screening in Identifying CAR Modulators.

Author information

1
National Center for Advancing Translational Sciences, National Institutes of Health, Building C, MSC: 3375, 9800 Medical Center Drive, Bethesda, MD, 20892, USA.
2
Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, Baltimore, MD, USA.
3
National Center for Advancing Translational Sciences, National Institutes of Health, Building C, MSC: 3375, 9800 Medical Center Drive, Bethesda, MD, 20892, USA. mxia@mail.nih.gov.

Abstract

The constitutive androstane receptor (CAR, NR1I3) is responsible for the transcription of multiple drug metabolizing enzymes and transporters. There are two possible methods of activation for CAR, direct ligand binding and a ligand-independent method, which makes this a unique nuclear receptor. Both of these mechanisms require translocation of CAR from the cytoplasm into the nucleus. Interestingly, CAR is constitutively active in immortalized cell lines due to the basal nuclear location of this receptor. This creates an important challenge in most in vitro assay models because immortalized cells cannot be used without inhibiting the high basal activity. In this book chapter, we go into detail of how to perform quantitative high-throughput screens to identify hCAR1 modulators through the employment of a double stable cell line. Using this line, we are able to identify activators, as well as deactivators, of the challenging nuclear receptor, CAR.

KEYWORDS:

Constitutive Androstane Receptor (CAR); Cytochrome P450 2B6 (CYP2B6); Luciferase; Quantitative high-throughput screening (qHTS)

PMID:
27518621
PMCID:
PMC5341601
DOI:
10.1007/978-1-4939-6346-1_4
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Springer Icon for PubMed Central
Loading ...
Support Center