Identification of small molecule ligands targeting GPR83, a G‐protein coupled receptor activated by the abundant neuropeptide PEN

Abstract

The G‐protein coupled receptor GPR83 is expressed in numerous brain regions, including striatum and hypothalamus, as well as in the immune system, including the thymus and spleen, of mice. Previously, GPR83 was classified as an orphan GPCR with no known endogenous ligand; we identified the neuropeptide PEN, derived from the precursor ProSAAS, as a selective, high‐affinity endogenous ligand for GPR83. Previous studies using ProSAAS or GPR83 knockout models have shown both the neuropeptide and receptor to be important in modulation of stress and anxiety, as well as feeding and body weight regulation. Additionally, the expression of GPR83 on immune cells indicates a potential role in immune function. Therefore, development of small molecule ligands for GPR83 is necessary in order to further our understanding of the receptor in these biological processes. In order to identify potential agonists and antagonists, a homology model for GPR83, based on the crystal structure of phylogenetically related receptors, was generated and used to screen in silico with the eMolecules library, which consists of 7 million compounds. From these, the top fifty predicted hits were screened in a cell based, high‐throughput calcium‐release assay. The compounds that gave a large signal to noise ratio only in cells expressing GPR83 were further characterized using GTPγS, adenylyl cyclase, phospholipase C, and receptor trafficking assays. This led to the identification of CPD1 and CPD27as selective agonists. Those compounds that did not give an agonistic response were re‐screened in an antagonist mode (i.e., in the presence of PEN, the endogenous agonist). The compounds that blocked PEN‐mediated calcium increase were further tested in GTPγS, adenylyl cyclase and receptor trafficking assays in the antagonist mode. From these assays, we identified CPD25 as a GPR83‐selective antagonist. Currently, we are characterizing these compounds using the native receptors in the brain and immune cells, with tissues from mice lacking GPR83 as controls. Characterization of these small molecule compounds provides much needed tools for furthering our understanding of the precise role of GPR83 in these biological processes. Also, since GPR83 has been highly implicated in the regulation of stress, mood and reward‐related behaviors, as well as immune function, identification of GPR83 selective ligands opens the door to development of novel therapeutics as regulators of this neuropeptide‐receptor system.Support or Funding InformationFunding: NS026880 and DA019521 to L.A.D and T32 grant DA007135 to L.M.L.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.