Multiplex GPCR Assay in Reverse Transfection Cell Microarrays
| Autor: | Chatanika Stoop-Myer, Robin Pedersen, Vic E. Myer, Sena Jong, Yuji Mishina, Brent L. Kreider, Susan K. Lyman, Chris Wilson, Jesse J. Smith, Linda Bruett, Lizabeth P. Amaral, Craig M. Thompson |
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| Rok vydání: | 2004 |
| Předmět: |
0301 basic medicine
G protein Drug Evaluation Preclinical Protein Array Analysis Computational biology Biology Ligands Transfection 01 natural sciences Biochemistry Receptors G-Protein-Coupled Analytical Chemistry 03 medical and health sciences Plasmid GTP-Binding Proteins Calcium flux Humans Multiplex Receptor G protein-coupled receptor Dose-Response Relationship Drug Molecular biology 0104 chemical sciences 010404 medicinal & biomolecular chemistry 030104 developmental biology Molecular Medicine DNA microarray Reverse transfection Biotechnology |
| Zdroj: | SLAS Discovery. 9:196-207 |
| ISSN: | 2472-5552 |
| DOI: | 10.1177/1087057103261880 |
| Popis: | G protein-coupled receptors (GPCRs) are a superfamily of proteins that include some of the most important drug targets in the pharmaceutical industry. Despite the success of this group of drugs, there remains a need to identify GPCR-targeted drugs with greater selectivity, to develop screening assays for validated targets, and to identify ligands for orphan receptors. To address these challenges, the authors have created a multiplexed GPCR assay that measures greater than 3000 receptor: ligand interactions in a single microplate. The multiplexed assay is generated by combining reverse transfection in a 96-well plate format with a calcium flux readout. This assay quantitatively measures receptor activation and inhibition and permits the determination of compound potency and selectivity for entire families of GPCRs in parallel. To expand the number of GPCR targets that may be screened in this system, receptors are cotransfected with plasmids encoding a promiscuous G protein, permitting the analysis of receptors that do not normally mobilize intracellular calcium upon activation. The authors demonstrate the utility of reverse transfection cell microarrays to GPCR-targeted drug discovery with examples of ligand selectivity screening against a panel of GPCRs as well as dose-dependent titrations of selected agonists and antagonists. |
| Databáze: | OpenAIRE |
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