Novel Small Molecule Glucagon-Like Peptide-1 Receptor Agonist Stimulates Insulin Secretion in Rodents and From Human Islets

OBJECTIVE

The clinical effectiveness of parenterally-administered glucagon-like peptide-1 (GLP-1) mimetics to improve glucose control in patients suffering from type 2 diabetes strongly supports discovery pursuits aimed at identifying and developing orally active, small molecule GLP-1 receptor agonists. The purpose of these studies was to identify and characterize novel nonpeptide agonists of the GLP-1 receptor.

RESEARCH DESIGN AND METHODS

Screening using cells expressing the GLP-1 receptor and insulin secretion assays with rodent and human islets were used to identify novel molecules. The intravenous glucose tolerance test (IVGTT) and hyperglycemic clamp characterized the insulinotropic effects of compounds in vivo.

RESULTS

Novel low molecular weight pyrimidine-based compounds that activate the GLP-1 receptor and stimulate glucose-dependent insulin secretion are described. These molecules induce GLP-1 receptor-mediated cAMP signaling in HEK293 cells expressing the GLP-1 receptor and increase insulin secretion from rodent islets in a dose-dependent manner. The compounds activate GLP-1 receptor signaling, both alone or in an additive fashion when combined with the endogenous GLP-1 peptide; however, these agonists do not compete with radiolabeled GLP-1 in receptor-binding assays. In vivo studies using the IVGTT and the hyperglycemic clamp in Sprague Dawley rats demonstrate increased insulin secretion in compound-treated animals. Further, perifusion assays with human islets isolated from a donor with type 2 diabetes show near-normalization of insulin secretion upon compound treatment.

CONCLUSIONS

These studies characterize the insulinotropic effects of an early-stage, small molecule GLP-1 receptor agonist and provide compelling evidence to support pharmaceutical optimization.Hyperglucagonemia and dysregulation of insulin secretion impair postprandial glucose homeostasis and thus are central to the pathogenesis of type 2 diabetes. Impairment of the incretin effect, including reduced secretion of glucagon-like peptide-1 (GLP-1) (1,2), is implicated in the progression of pancreatic islet dysfunction in type 2 diabetic patients. Development and use of incretin-based therapeutics may, therefore, be an effective strategy to restore normal islet function by providing insulinotropic and glucagon-suppressive capabilities. In recent years, clinical studies have shown that replacement therapy with metabolically stable GLP-1 mimetics greatly improves management of hyperglycemia, nearly correcting blood glucose regulation for some patients. For example, treatment with either exenatide or liraglutide reduces fasting hyperglycemia and results in sustained lowering of glycosylated A1C (A1C) levels (3,4). In addition, these therapies often reduce body weight and improve several cardiovascular parameters (5,6). Unfortunately, both of these GLP-1 analogues are peptides requiring administration by subcutaneous injection.The GLP-1 receptor is a member of the class B/II family of seven transmembrane G protein-coupled receptors (GPCRs) that include receptors for peptide hormones such as secretin, GLP-1, glucose-dependent insulinotropic polypeptide (GIP), glucagon, vasoactive intestinal peptide (VIP), corticotropin-releasing factor (CRF), calcitonin, and parathyroid hormone (PTH) (7–9). Historically, the discovery of nonpeptide agonists of these receptors that could enable development of orally active pharmaceuticals has been generally unsuccessful. To a large extent, this difficulty has been attributed to the mechanisms used by class B GPCRs to recognize ligands and induce signaling. Interaction of endogenous peptide hormones with their receptors typically involves large receptor:ligand binding sites and is often initiated by receptor NH₂-terminal ectodomains (ECDs). This extracellular structure interacts with COOH-terminal residues of cognate ligands and positions the NH₂-terminus of the ligand to interact with critical determinants in receptor transmembrane regions, thereby activating heterotrimeric G-proteins and subsequently adenylyl cyclase (10–12).The recent reports describing GLP-1 receptor activation by a series of substituted quinoxalines (13–16) and a cyclobutane derivative (17) suggest that it may be possible to develop nonpeptide GLP-1 receptor agonists. Both scaffolds activate GLP-1 receptor signaling in heterologous GLP-1 receptor-expressing cellular systems, and the quinoxalines clearly induce glucose-stimulated insulin secretion in vitro from rodent islets and ex vivo via perfused pancreas (13,16). We now report novel, low molecular weight pyrimidines that activate the GLP-1 receptor to induce glucose-dependent insulin secretion both in vitro and in vivo. These molecules may offer therapeutic advantage because of the ability of the compounds to act alone or in combination with GLP-1. Further, these agonists nearly restore insulin secretion to normal in human diabetic islets.