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Fluorescent retinoid X receptor ligands for fluorescence polarization assay
Shoya Yamada1, Fuminori Ohsawa, Shuji Fujii
1Division of Pharmaceutical Sciences, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Tsushima-Naka, Okayama, Japan.
Abstract:
Retinoid X receptor (RXR) agonists are candidate agents for the treatment of metabolic syndrome and type 2 diabetes via activation of peroxisome proliferator-activated receptor (PPAR)/RXR or liver X receptor (LXR)/RXR-heterodimers, which control lipid and glucose metabolism. Reporter gene assays or binding assays with radiolabeled compounds are available for RXR ligand screening, but are unsuitable for high-throughput screening. Therefore, as a first step towards stabilizing a fluorescence polarization (FP) assay system for high-throughput RXR ligand screening, we synthesized fluorescent RXR ligands by modification of the lipophilic domain of RXR ligands with a carbostyril fluorophore, and selected the fluorescent RXR agonist 6-[ethyl(1-isobutyl-2-oxo-4-trifluoromethyl-1,2-dihydroquinolin-7-yl)amino]nicotinic acid 8d for further characterization. Compound 8d showed FP in the presence of RXR and the FP was decreased in the presence of the RXR agonist LGD1069 (2). This compound should be a lead compound for use in high-throughput assay systems for screening RXR ligands.
Insights
Researchers developed a novel fluorescent Retinoid X receptor (RXR) ligand for high-throughput screening. This compound enables a new fluorescence polarization assay to identify potential treatments for metabolic syndrome and type 2 diabetes.
Area of Science:
- Molecular Endocrinology
- Drug Discovery
- Assay Development
Background:
- Retinoid X receptor (RXR) agonists are investigated for treating metabolic syndrome and type 2 diabetes by modulating lipid and glucose metabolism through PPAR/RXR and LXR/RXR heterodimers.
- Current RXR ligand screening methods, such as reporter gene and binding assays, are not suitable for high-throughput screening (HTS).
Purpose of the Study:
- To develop a fluorescence polarization (FP) assay system for high-throughput screening of Retinoid X receptor (RXR) ligands.
- To synthesize novel fluorescent RXR ligands as tools for assay development.
Main Methods:
- Synthesis of fluorescent RXR ligands by conjugating a carbostyril fluorophore to the lipophilic domain of known RXR ligands.
- Selection and characterization of a lead fluorescent compound (8d).
- Validation of the fluorescent ligand in a fluorescence polarization assay using RXR and a known RXR agonist (LGD1069).
Main Results:
- A fluorescent RXR agonist, 6-[ethyl(1-isobutyl-2-oxo-4-trifluoromethyl-1,2-dihydroquinolin-7-yl)amino]nicotinic acid (8d), was synthesized and selected.
- Compound 8d demonstrated measurable fluorescence polarization (FP) in the presence of RXR.
- The FP signal of compound 8d was diminished by the addition of a known RXR agonist, LGD1069, confirming its utility in an RXR binding assay.
Conclusions:
- Compound 8d serves as a promising lead compound for developing a high-throughput fluorescence polarization assay for RXR ligand screening.
- This novel fluorescent probe facilitates the discovery of new therapeutic agents for metabolic disorders targeting RXR pathways.
