Celia P. Briscoe

The orphan G protein-coupled receptor GPR40 is activated by medium and long-chain fatty acids

GPR40 is a member of a subfamily of homologous G protein-coupled receptors that include GPR41 and GPR43 and that have no current function or ligand ascribed. Ligand fishing experiments in HEK293 cells expressing human GPR40 revealed that a range of saturated and unsaturated carboxylic acids with carbon chain lengths greater than six were able to induce an elevation of [Ca2+] i , measured using a fluorometric imaging plate reader. 5,8,11-Eicosatriynoic acid was the most potent fatty acid tested, with a pEC50 of 5.7. G protein coupling of GPR40 was examined in Chinese hamster ovary cells expressing the Gαq/i-responsive Gal4-Elk1 reporter system. Expression of human GPR40 led to a constitutive induction of luciferase activity, which was further increased by exposure of the cells to eicosatriynoic acid. Neither the constitutive nor ligand-mediated luciferase induction was inhibited by pertussis toxin treatment, suggesting that GPR40 was coupled to Gαq/11. Expression analysis by quantitative reverse transcription-PCR showed that GPR40 was specifically expressed in brain and pancreas, with expression in rodent pancreas being localized to insulin-producing β-cells. These data suggest that some of the physiological effects of fatty acids in pancreatic islets and brain may be mediated through a cell-surface receptor.

Pharmacological regulation of insulin secretion in MIN6 cells through the fatty acid receptor GPR40: identification of agonist and antagonist small molecules

1 Long chain fatty acids have recently been identified as agonists for the G protein‐coupled receptors GPR40 and GPR120. Here, we present the first description of GW9508, a small‐molecule agonist of the fatty acid receptors GPR40 and GPR120. In addition, we also describe the pharmacology of GW1100, a selective GPR40 antagonist. These molecules were used to further investigate the role of GPR40 in glucose‐stimulated insulin secretion in the MIN6 mouse pancreatic β‐cell line. 2 GW9508 and linoleic acid both stimulated intracellular Ca2+ mobilization in human embryonic kidney (HEK)293 cells expressing GPR40 (pEC50 values of 7.32±0.03 and 5.65±0.06, respectively) or GPR120 (pEC50 values of 5.46±0.09 and 5.89±0.04, respectively), but not in the parent HEK‐293 cell line. 3 GW1100 dose dependently inhibited GPR40‐mediated Ca2+ elevations stimulated by GW9508 and linoleic acid (pIC50 values of 5.99±0.03 and 5.99±0.06, respectively). GW1100 had no effect on the GPR120‐mediated stimulation of intracellular Ca2+ release produced by either GW9508 or linoleic acid. 4 GW9508 dose dependently potentiated glucose‐stimulated insulin secretion in MIN6 cells, but not in primary rat or mouse islets. Furthermore, GW9508 was able to potentiate the KCl‐mediated increase in insulin secretion in MIN6 cells. The effects of GW9508 on insulin secretion were reversed by GW1100, while linoleic acid‐stimulated insulin secretion was partially attenuated by GW1100. 5 These results add further evidence to a link between GPR40 and the ability of fatty acids to acutely potentiate insulin secretion and demonstrate that small‐molecule GPR40 agonists are glucose‐sensitive insulin secretagogues.

Pharmacological characterization of a small-molecule inhibitor of c-Jun kinase

c-Jun NH(2)-terminal kinase (JNK) plays an important role in insulin resistance; however, identification of pharmacologically potent and selective small molecule JNK inhibitors has been limited. Compound A has a cell IC(50) of 102 nM and is at least 100-fold selective against related kinases and 27-fold selective against glycogen synthase kinase-3beta and cyclin-dependent kinase-2. In C57BL/6 mice, compound A reduced LPS-mediated increases in both plasma cytokine levels and phosphorylated c-Jun in adipose tissue. Treatment of mice fed a high-fat diet with compound A for 3 wk resulted in a 13.1 +/- 1% decrease in body weight and a 9.3 +/- 1.5% decrease in body fat, compared with a 6.6 +/- 2.1% increase in body weight and a 6.7 +/- 2.1% increase in body fat in vehicle-treated mice. Mice pair fed to those that received compound A exhibited a body weight decrease of 7 +/- 1% and a decrease in body fat of 1.6 +/- 1.3%, suggesting that reductions in food intake could not account solely for the reductions in adiposity observed. Compound A dosed at 30 mg/kg for 13 days in high-fat fed mice resulted in a significant decrease in phosphorylated c-Jun in adipose tissue accompanied by a decrease in weight and reductions in glucose and triglycerides and increases in insulin sensitivity to levels comparable with those in lean control mice. The ability of compound A to reduce the insulin-stimulated phosphorylation of insulin receptor substrate-1 (IRS-1) von Ser307 and partially reverse the free fatty acid inhibition of glucose uptake in 3T3L1 adipocytes, suggests that enhancement of insulin signaling in addition to weight loss may contribute to the effects of compound A on insulin sensitization in vivo. Pharmacological inhibition of JNK using compound A may therefore offer an effective therapy for type 2 diabetes mediated at least in part via weight reduction.

Discovery of 6,7-Dihydro-5H-pyrrolo[2,3-a]pyrimidines as Orally Available G Protein-Coupled Receptor 119 Agonists

GPR119 is a 7-transmembrane receptor that is expressed in the enteroendocrine cells in the intestine and in the islets of Langerhans in the pancreas. Indolines and 6,7-dihydro-5H-pyrrolo[2,3-a]pyrimidines were discovered as G protein-coupled receptor 119 (GPR119) agonists, and lead optimization efforts led to the identification of 1-methylethyl 4-({7-[2-fluoro-4-(methylsulfonyl)phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarboxylate (GSK1104252A) (3), a potent and selective GPR119 agonist. Compound 3 showed excellent pharmacokinetic properties and sufficient selectivity with in vivo studies supporting a role for GPR119 in glucose homeostasis in the rodent. Thus, 3 appeared to modulate the enteroinsular axis, improve glycemic control, and strengthen previous suggestions that GPR119 agonists may have utility in the treatment of type 2 diabetes.