Abstract
Hyperglucagonemia is implicated in the pathophysiology of hyperglycemia. Antagonism of the glucagon receptor (GCGR) thus represents a potential approach to diabetes treatment. Herein we report the characterization of GRA1, a novel small-molecule GCGR antagonist that blocks glucagon binding to the human GCGR (hGCGR) and antagonizes glucagon-induced intracellular accumulation of cAMP with nanomolar potency. GRA1 inhibited glycogenolysis dose-dependently in primary human hepatocytes and in perfused liver from hGCGR mice, a transgenic line of mouse that expresses the hGCGR instead of the murine GCGR. When administered orally to hGCGR mice and rhesus monkeys, GRA1 blocked hyperglycemic responses to exogenous glucagon. In several murine models of diabetes, acute and chronic dosing with GRA1 significantly reduced blood glucose concentrations and moderately increased plasma glucagon and glucagon-like peptide-1. Combination of GRA1 with a dipeptidyl peptidase-4 inhibitor had an additive antihyperglycemic effect in diabetic mice. Hepatic gene-expression profiling in monkeys treated with GRA1 revealed down-regulation of numerous genes involved in amino acid catabolism, an effect that was paralleled by increased amino acid levels in the circulation. In summary, GRA1 is a potent glucagon receptor antagonist with strong antihyperglycemic efficacy in preclinical models and prominent effects on hepatic gene-expression related to amino acid metabolism.
Highlights
Glucagon is a 29 amino acid polypeptide hormone that is secreted by pancreatic alpha cells primarily during the fasting state [1]
In Vitro Assays Transfected Chinese hamster ovary (CHO) cell lines were acquired and maintained as previously described [17,20]. These included separate cell lines stably expressing human glucagon receptor (GCGR), mouse GCGR, rhesus GGCR, glucose-dependent insulinotropic peptide receptor (GIPR), glucagon-like peptide-1 (GLP-1) receptor (GLP1R), pituitary adenylate cyclase-activating polypeptide receptor type 1 (PAC1R), and vasoactive adenylate cyclase-activating polypeptide receptor type 2 (VPAC2R)
Production of cAMP by glucagon-stimulated human GCGR (hGCGR)-expressing CHO cells was inhibited by GRA1 with an IC50 of 12 nM
Summary
Glucagon is a 29 amino acid polypeptide hormone that is secreted by pancreatic alpha cells primarily during the fasting state [1]. It plays a critical role in glucose homeostasis and the prevention of hypoglycemia, primarily by promoting glycogenolysis and gluconeogenesis in the liver and attenuating inhibition of these processes by insulin [2,3]. In the mouse, targeted disruption of the GCGR gene results in reduced plasma glucose concentrations [11,12] and treatment with GCGR antisense oligonucleotides has an antihyperglycemic effect in rodent models of diabetes [13,14]. The phenotype of GCGR knockout mice does, include some potentially troublesome features; GCGR mice have prominent a-cell hyperplasia and very high plasma concentrations of glucagon and both active and inactive GLP-1 [12,15]
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