The activation of the glucagon-like peptide-1 (GLP-1) receptor by peptide and non-peptide ligands

The glucagon-like peptide-1 receptor (GLP-1R) potentiates glucose-stimulated insulin release from pancreatic β cells and promotes correct β cell function, as such it is a validated target for the treatment of type 2 diabetes (T2D). GLP-1R is a Family B GPCR, activated by the cognate ligand GLP-1(7-36), a 30 residue peptide hormone secreted after eating, and Exendin4 (Ex4), a 39-residue synthetic peptide. Peptide ligands interact with both the large extracellular domain and core domain of GLP-1R. Core domain interaction is thought to activate the receptor. Whilst the interaction between the receptor extracellular domain and ligand is well characterised, the ligand-core domain interaction and subsequent activation is not fully understood. Herein, a combination of mutant peptides and non-peptide ligands based on a pyrimidine scaffold (Pm compounds) are used in HTR-FRET cAMP accumulation assays, using recombinant FlpIn-HEK293 cells expressing human GLP-1R, to characterise the activation profiles of these ligands to decipher the underlying activation mechanism at the GLP-1R core domain. Structure-function studies of Pm compounds showed a trifluoromethyl and sulphur dioxide group are essential for GLP-1R activation, and that they allosterically enhance GLP-1(9-36) and Ex4(9-39) cAMP signalling profiles independently from their own cAMP response. Insulin secretion assays showed Pm compounds potentiate insulin release from INS-1 832/13 cells in combination with truncated GLP-1(9-36), implicating the use of allosteric modulators as treatment for T2D. Truncated GLP-1(15-36) was capable of binding and activating GLP-1R with low affinity and low potency, yet analogously truncated Ex4(9-39) was an antagonist with high affinity. Previous studies had demonstrated GLP-1(15-36) was an antagonist, and peptide-mediated activity had been attributed to the amino-terminus. Furthermore, the Pm compound-mediated cAMP response at GLP-1R was potentiated by Ex4(9-39). Mutant peptide activation data suggest activating residues D15, V16 and S17 are situated more centrally within the peptide ligand, and an extension to the currently accepted GLP-1R activation model is proposed.