In vitro profiling of orphan G protein coupled receptor (GPCR) constitutive activity

Abstract

Background and PurposeMembers of the G protein coupled receptor (GPCR) family are targeted by a significant fraction of the available FDA-approved drugs. However, the physiological role and pharmacological properties of many GPCRs remain unknown, representing untapped potential in drug design. Of particular interest are ~100 less-studied GPCRs known as orphans because their endogenous ligands are unknown. Intriguingly, disease-causing mutations identified in patients, together with animal studies, have demonstrated that many orphan receptors play crucial physiological roles, and thus, represent attractive drug targets. Experimental ApproachThe majority of deorphanized GPCRs demonstrate coupling to Gi/o, however a limited number of techniques allow the detection of intrinsically small constitutive activity associated with Gi/o protein activation which represents a significant barrier in our ability to study orphan GPCR signaling. Using luciferase reporter assays, we effectively detected constitutive Gs, Gq, and G12/13 protein signaling by unliganded receptors, and introducing various G protein chimeras, we provide a novel, highly-sensitive tool capable of identifying Gi/o coupling in unliganded orphan GPCRs. Key ResultsUsing this approach, we measured the constitutive activity of the entire class C GPCR family that includes 8 orphan receptors, and a subset of 20 prototypical class A GPCR members, including 11 orphans. Excitingly, this approach illuminated the G protein coupling profile of 8 orphan GPCRs (GPR22, GPR137b, GPR88, GPR156, GPR158, GPR179, GPRC5D, and GPRC6A) previously linked to pathophysiological processes. Conclusion and ImplicationsWe provide a new platform that could be utilized in ongoing studies in orphan receptor signaling and deorphanization efforts. What is already knownO_LIA large group of understudied orphan GPCRs controls a variety of physiological process. C_LI What this study addsO_LIA new strategy to identify G protein signaling associated with orphan GPCRs. C_LIO_LIIdentification of Gi/o coupling for 8 orphan GPCRs. C_LI What is the clinical significanceO_LIMany orphan GPCRs are associated with pathological conditions and represent promising druggable targets. C_LI