title: Development of a high-throughput screening-compatible cell-based functional assay to identify small molecule probes of the galanin 3 receptor (GalR3).
abstract: The galanin 3 receptor (GalR3) belongs to the large G protein-coupled receptor (GPCR) family of proteins. GalR3 and two other closely related receptors, GalR1 and GalR2, together with their endogenous ligand galanin, are involved in a variety of physiological and pathophysiological processes. GalR3 in particular has been strongly implicated in addiction and mood-related disorders such as anxiety and depression. It has been the target of many drug discovery programs within the pharmaceutical industry, but despite the significant resources and effort devoted to discovery of galanin receptor subtype selective small molecule modulators, there have been very few reports for the discovery of such molecules. GalR3 has proven difficult to enable in cell-based functional assays due to its apparent poor cell surface expression in recombinant systems. Here, we describe the generation of a modified GalR3 that facilitates its cell surface expression while maintaining wild-type receptor pharmacology. The modified GalR3 has been used to develop a high-throughput screening-compatible, cell-based, cAMP biosensor assay to detect selective small molecule modulators of GalR3. The performance of the assay has been validated by challenging it against a test library of small molecules with known pharmacological activities (LOPAC; Sigma Aldrich). This approach will enable identification of GalR3 selective modulators (chemical probes) that will facilitate dissection of the biological role(s) that GalR3 plays in normal physiological processes as well as in disease states.
authors.affiliation: 1 Department of Molecular Therapeutics, The Scripps Research Institute , Jupiter, Florida.
authors.forename: James
authors.initials: J
authors.lastname: Robinson
authors.forename: Anthony
authors.initials: A
authors.lastname: Smith
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Sturchler
authors.forename: Sahba
authors.initials: S
authors.lastname: Tabrizifard
authors.forename: Theodore
authors.initials: T
authors.lastname: Kamenecka
authors.forename: Patricia
authors.initials: P
authors.lastname: McDonald
mesh_terms: D015374:Biosensing Techniques; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D000242:Cyclic AMP / Q000378:metabolism; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D019084:Fluorescent Antibody Technique, Indirect; D019004:Galanin / Q000378:metabolism; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D008024:Ligands; D044093:Receptor, Galanin, Type 3 / Q000187:drug effects* / Q000235:genetics; D054852:Small Molecule Libraries
chemical_list: D018076:DNA, Complementary; D008024:Ligands; D044093:Receptor, Galanin, Type 3; D054852:Small Molecule Libraries; D019004:Galanin; D000242:Cyclic AMP