title: Identification and characterization of mGlu3 ligands using a high throughput FLIPR assay for detection of agonists, antagonists, and allosteric modulators.
abstract: When targeting G-protein coupled receptors (GPCRs) in early stage drug discovery, or for novel targets, the type of ligand most likely to produce the desired therapeutic effect may be unknown. Therefore, it can be desirable to identify potential lead compounds from multiple categories: agonists, antagonists, and allosteric modulators. In this study, we developed a triple addition calcium flux assay using FLIPR Tetra to identify multiple ligand classes for the metabotropic glutamate receptor 3 (mGlu3), using a cell line stably co-expressing the human G-protein-coupled mGlu3 receptor, a promiscuous G-protein (G(α16)), and rat Glast, a glutamate transporter. Compounds were added to the cells followed by stimulation with EC(10) and then EC(80) concentration of glutamate, the physiological agonist for mGlu receptors. This format produced a robust assay, facilitating the identification of agonists, positive allosteric modulators and antagonists/negative allosteric modulators. Follow up experiments were conducted to exclude false positives. Using this approach, we screened a library of approximately 800,000 compounds using FLIPR Tetra and identified viable leads for all three ligand classes. Further characterization revealed the selectivity of individual ligands.
authors.affiliation: Advanced Technology, Global Pharmaceutical Research and Development, Abbott Laboratories, IL 60064, USA. steve.pratt@abbott.com
authors.forename: Steven D
authors.initials: SD
authors.lastname: Pratt
authors.forename: Mario
authors.initials: M
authors.lastname: Mezler
authors.forename: Hervé
authors.initials: H
authors.lastname: Geneste
authors.forename: Margot H M
authors.initials: MH
authors.lastname: Bakker
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Hajduk
authors.forename: Sujatha M
authors.initials: SM
authors.lastname: Gopalakrishnan
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects; D002478:Cells, Cultured; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D008024:Ligands; D018094:Receptors, Metabotropic Glutamate / Q000819:agonists* / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D008024:Ligands; D018094:Receptors, Metabotropic Glutamate; C102718:metabotropic glutamate receptor 3