Homology Modeling of Ffa2 Identifies Novel Agonists that Potentiate Insulin Secretion

Abstract

Critical aspects of maintaining glucose homeostasis in the face of chronic insulin resistance and type 2 diabetes (T2D) are increased insulin secretion and adaptive expansion of beta cell mass. Nutrient and hormone sensing G protein-coupled receptors are important mediators of these properties. A growing body of evidence now suggests that the G protein-coupled receptor, free fatty acid receptor 2 (FFA2), is capable of contributing to the maintenance of glucose homeostasis by acting at the pancreatic beta cell as well as at other metabolically active tissues. We have previously demonstrated that Gαq/11-biased agonism of FFA2 can potentiate glucose stimulated insulin secretion (GSIS) as well as promote beta cell proliferation. However, the currently available Gαq/11-biased agonists for FFA2 exhibit low potency, making them difficult to examine in vivo. This study sought to identify Gαq/11-biased FFA2-selective agonists with potent GSIS-stimulating effects. To do this, we generated an FFA2 homology model that was used to screen a library of 10 million drug-like compounds. Although FFA2 and the related short chain fatty acid receptor FFA3 share 52% sequence similarity, our virtual screen identified over 50 compounds with predicted selectivity and increased potency for FFA2 over FFA3. Subsequent in vitro calcium mobilization assays and GSIS assays resulted in the identification of a compound that can potentiate GSIS via activation of Gαq/11 with 100-fold increased potency compared with previously described Gαq/11-biased FFA2 agonists. These methods and findings provide a foundation for future discovery efforts to identify biased FFA2 agonists as potential T2D therapeutics.

Short chain fatty acids stimulate insulin secretion and reduce apoptosis in mouse and human islets in vitro: Role of free fatty acid receptor 2

“…Thus, while FFAR3 couples only to Gi to decrease intracellular cAMP levels, FFAR2 is able to recruit both Gi and Gq to decrease cAMP and increase diacylglycerol and inositol trisphosphate generation. We have previously demonstrated that cannabinoid receptor agonists that signal through both Gi and Gq increase insulin secretion, as the stimulatory input outweighs the reduction in cAMP production, and it has recently been reported that a Gq‐biased FFAR2‐selective agonist potentiates glucose‐induced insulin secretion . Consistent with this, we have reported that propionate reversibly stimulates insulin secretion from human islets through a Gq/protein kinase C (PKC)‐dependent pathway …”

“…We have previously demonstrated that cannabinoid receptor agonists that signal through both Gi and Gq increase insulin secretion, as the stimulatory input outweighs the reduction in cAMP production, 15,16 and it has recently been reported that a Gq-biased FFAR2-selective agonist potentiates glucose-induced insulin secretion. 12 Consistent with this, we have reported that propionate reversibly stimulates insulin secretion from human islets through a Gq/protein kinase C (PKC)-dependent pathway. 5 Observations of stimulatory signalling in β-cells via FFAR2 contrast with the report that transgenic mice with global FFAR2 deletion show enhanced insulin secretion, which implicates pre-eminence of the Gi signalling pathway in mouse β-cells.…”

“…In particular, we and others have identified that FFAR2 and FFAR3 are expressed by islets of Langerhans, opening up the possibility that SCFAs contribute to the regulation of insulin secretion through activation of these receptors. However, the data published so far do not provide a consensus role for SCFAs and their receptors in islets: the first paper published in this area reported that acetate inhibited insulin release in vitro and that deletion of FFAR2 and FFAR3 improved glucose tolerance, while there have been more recent reports of SCFAs stimulating insulin secretion and of deletion of FFAR2 being associated with insufficient β‐cell mass expansion in obesity and gestational diabetes . Acetate is the most abundant circulating SCFA, reaching plasma concentrations of ~200 μM, while propionate concentrations are ~10‐fold lower, and these are the two SCFAs that are used most commonly for experimental studies.…”

“…However, the data published so far do not provide a consensus role for SCFAs and their receptors in islets: the first paper published in this area reported that acetate inhibited insulin release in vitro and that deletion of FFAR2 and FFAR3 improved glucose tolerance, 7 while there have been more recent reports of SCFAs stimulating insulin secretion and of deletion of FFAR2 being associated with insufficient β-cell mass expansion in obesity and gestational diabetes. 5,6,[8][9][10][11][12] Acetate is the most abundant circulating SCFA, reaching plasma concentrations of~200 μM, 13 while propionate concentrations are~10-fold lower, 2 and these are the two SCFAs that are used most commonly for experimental studies. They can couple to both inhibitory and stimulatory signalling through activation of FFAR2 and/or FFAR3, which differ in their downstream signalling cascades.…”

Discovery of novel Mnk inhibitors using mutation‐based induced‐fit virtual high‐throughput screening

“…The 100 highest‐scoring ligands were then manually evaluated to eliminate those that did not appear to form hydrogen‐bonds with the kinase hinge region or those that did not have a Glide XP score ≤−7.0, which resulted in a refined hit set of 43 compounds. Finally, this set was filtered for druggability, synthetic tractability, and compound availability in a manner similar to our recent work (Mishra, Shum, Platanias, Miller, & Schiltz, ; Mishra et al., ; Villa et al., ; Zhu et al., ), and 15 compounds were purchased for in vitro screening. The docked pose of hit compound NUCC‐54139 relative to the original Mnk1 structure and ligands used in the M‐IFD is shown in Figure b.…”