Yoshiyuki Tsujihata

TAK-875, an orally available G protein-coupled receptor 40/free fatty acid receptor 1 agonist, enhances glucose-dependent insulin secretion and improves both postprandial and fasting hyperglycemia in type 2 diabetic rats.

G protein-coupled receptor 40/free fatty acid receptor 1 (GPR40/FFA1) is highly expressed in pancreatic β cells and mediates free fatty acid-induced insulin secretion. This study examined the pharmacological effects and potential for avoidance of lipotoxicity of [(3S)-6-({2′,6′-dimethyl-4′-[3-(methylsulfonyl)propoxy]biphenyl-3-yl}meth-oxy)-2,3-dihydro-1-benzofuran-3-yl]acetic acid hemi-hydrate) (TAK-875), a novel, orally available, selective GPR40 agonist. Insulinoma cell lines and primary rat islets were used to assess the effects of TAK-875 in vitro. The in vivo effects of TAK-875 on postprandial hyperglycemia, fasting hyperglycemia, and normoglycemia were examined in type 2 diabetic and normal rats. In rat insulinoma INS-1 833/15 cells, TAK-875 increased intracellular inositol monophosphate and calcium concentration, consistent with activation of the Gqα signaling pathway. The insulinotropic action of TAK-875 (10 μM) in INS-1 833/15 and primary rat islets was glucose-dependent. Prolonged exposure of cytokine-sensitive INS-1 832/13 to TAK-875 for 72 h at pharmacologically active concentrations did not alter glucose-stimulated insulin secretion, insulin content, or caspase 3/7 activity, whereas prolonged exposure to palmitic or oleic acid impaired β cell function and survival. In an oral glucose tolerance test in type 2 diabetic N-STZ-1.5 rats, TAK-875 (1–10 mg/kg p.o.) showed a clear improvement in glucose tolerance and augmented insulin secretion. In addition, TAK-875 (10 mg/kg, p.o.) significantly augmented plasma insulin levels and reduced fasting hyperglycemia in male Zucker diabetic fatty rats, whereas in fasted normal Sprague-Dawley rats, TAK-875 neither enhanced insulin secretion nor caused hypoglycemia even at 30 mg/kg. TAK-875 enhances glucose-dependent insulin secretion and improves both postprandial and fasting hyperglycemia with a low risk of hypoglycemia and no evidence of β cell toxicity.

The Effects of TAK-875, a Selective G Protein-Coupled Receptor 40/Free Fatty Acid 1 Agonist, on Insulin and Glucagon Secretion in Isolated Rat and Human Islets

G protein-coupled receptor 40 (GPR40)/free fatty acid 1 (FFA1) is a G protein-coupled receptor involved in free fatty acid-induced insulin secretion. To analyze the effect of our novel GPR40/FFA1-selective agonist, [(3S)-6-({2′,6′-dimethyl-4′-[3-(methylsulfonyl)propoxy]biphenyl-3-yl}methoxy)-2,3-dihydro-1-benzofuran-3-yl]acetic acid hemi-hydrate (TAK-875), on insulin and glucagon secretion, we performed hormone secretion assays and measured intracellular Ca2+ concentration ([Ca2+]i) in both human and rat islets. Insulin and glucagon secretion were measured in static and dynamic conditions by using groups of isolated rat and human pancreatic islets. [Ca2+]i was recorded by using confocal microscopy. GPR40/FFA1 expression was measured by quantitative polymerase chain reaction. In both human and rat islets, TAK-875 enhanced glucose-induced insulin secretion in a glucose-dependent manner. The stimulatory effect of TAK-875 was similar to that produced by glucagon-like peptide-1 and correlated with the elevation of β-cell [Ca2+]i. TAK-875 was without effect on glucagon secretion at both 1 and 16 mM glucose in human islets. These data indicate that GPR40/FFA1 influences mainly insulin secretion in a glucose-dependent manner. The β-cell-specific action of TAK-875 in human islets may represent a therapeutically useful feature that allows plasma glucose control without compromising counter-regulation of glucagon secretion, thus minimizing the risk of hypoglycemia.

Optimization of (2,3-dihydro-1-benzofuran-3-yl)acetic acids: discovery of a non-free fatty acid-like, highly bioavailable G protein-coupled receptor 40/free fatty acid receptor 1 agonist as a glucose-dependent insulinotropic agent.

G protein-coupled receptor 40 (GPR40)/free fatty acid receptor 1 (FFA1) is a free fatty acid (FFA) receptor that mediates FFA-amplified glucose-stimulated insulin secretion in pancreatic β-cells. We previously identified (2,3-dihydro-1-benzofuran-3-yl)acetic acid derivative 2 as a candidate, but it had relatively high lipophilicity. Adding a polar functional group on 2 yielded several compounds with lower lipophilicity and little effect on caspase-3/7 activity at 30 μM (a marker of toxicity in human HepG2 hepatocytes). Three optimized compounds showed promising pharmacokinetic profiles with good in vivo effects. Of these, compound 16 had the lowest lipophilicity. Metabolic analysis of 16 showed a long-acting PK profile due to high resistance to β-oxidation. Oral administration of 16 significantly reduced plasma glucose excursion and increased insulin secretion during an OGTT in type 2 diabetic rats. Compound 16 (TAK-875) is being evaluated in human clinical trials for the treatment of type 2 diabetes.

A Novel Antidiabetic Drug, Fasiglifam/TAK-875, Acts as an Ago-Allosteric Modulator of FFAR1

Selective free fatty acid receptor 1 (FFAR1)/GPR40 agonist fasiglifam (TAK-875), an antidiabetic drug under phase 3 development, potentiates insulin secretion in a glucose-dependent manner by activating FFAR1 expressed in pancreatic β cells. Although fasiglifam significantly improved glycemic control in type 2 diabetes patients with a minimum risk of hypoglycemia in a phase 2 study, the precise mechanisms of its potent pharmacological effects are not fully understood. Here we demonstrate that fasiglifam acts as an ago-allosteric modulator with a partial agonistic activity for FFAR1. In both Ca2+ influx and insulin secretion assays using cell lines and mouse islets, fasiglifam showed positive cooperativity with the FFAR1 ligand γ-linolenic acid (γ-LA). Augmentation of glucose-induced insulin secretion by fasiglifam, γ-LA, or their combination was completely abolished in pancreatic islets of FFAR1-knockout mice. In diabetic rats, the insulinotropic effect of fasiglifam was suppressed by pharmacological reduction of plasma free fatty acid (FFA) levels using a lipolysis inhibitor, suggesting that fasiglifam potentiates insulin release in conjunction with plasma FFAs in vivo. Point mutations of FFAR1 differentially affected Ca2+ influx activities of fasiglifam and γ-LA, further indicating that these agonists may bind to distinct binding sites. Our results strongly suggest that fasiglifam is an ago-allosteric modulator of FFAR1 that exerts its effects by acting cooperatively with endogenous plasma FFAs in human patients as well as diabetic animals. These findings contribute to our understanding of fasiglifam as an attractive antidiabetic drug with a novel mechanism of action.

TAK‐875, a GPR40/FFAR1 agonist, in combination with metformin prevents progression of diabetes and β‐cell dysfunction in Zucker diabetic fatty rats

TAK‐875, a selective GPCR40/free fatty acid receptor 1 agonist, improves glycaemic control by increasing glucose‐dependent insulin secretion. Metformin is a first‐line drug for treatment of type 2 diabetes that improves peripheral insulin resistance. Based on complementary mechanism of action, combining these agents is expected to enhance glycaemic control. Here, we evaluated the chronic effects of TAK‐875 monotherapy and combination therapy with metformin in diabetic rats.

Discovery of a novel series of indoline carbamate and indolinylpyrimidine derivatives as potent GPR119 agonists.

GPR119 has emerged as an attractive target for anti-diabetic agents. We identified a structurally novel GPR119 agonist 22c that carries a 5-(methylsulfonyl)indoline motif as an early lead compound. To generate more potent compounds of this series, structural modifications were performed mainly to the central alkylene spacer. Installation of a carbonyl group and a methyl group on this spacer significantly enhanced agonistic activity, resulting in the identification of 2-[1-(5-ethylpyrimidin-2-yl)piperidin-4-yl]propyl 7-fluoro-5-(methylsulfonyl)-2,3-dihydro-1H-indole-1-carboxylate (20). To further expand the chemical series of indoline-based GPR119 agonists, several heterocyclic core systems were introduced as surrogates of the carbamate spacer that mimic the presumed active conformation. This approach successfully produced an indolinylpyrimidine derivative 37, 5-(methylsulfonyl)-1-[6-({1-[3-(propan-2-yl)-1,2,4-oxadiazol-5-yl]piperidin-4-yl}oxy)pyrimidin-4-yl]-2,3-dihydro-1H-indole, which has potent GPR119 agonist activity. In rat oral glucose tolerance tests, these two indoline-based compounds effectively lowered plasma glucose excursion and glucose-dependent insulin secretion after oral administration.

Fasiglifam/TAK-875, a Selective GPR40 Agonist, Improves Hyperglycemia in Rats Unresponsive to Sulfonylureas and Acts Additively with Sulfonylureas.

Sulfonylureas (SUs) are widely used insulin secretagogues, but they have adverse effects including hypoglycemia and secondary failure. Fasiglifam/TAK-875, a selective GPR40 agonist, enhances glucose-stimulated insulin secretion and improves hyperglycemia. In the present study, we compared the in vivo glucose-lowering effects of fasiglifam with SUs. The risk of secondary failure of fasiglifam and the efficacy in rats desensitized to SUs were also evaluated. Moreover, we assessed whether fasiglifam was effective when combined with SUs. In diabetic neonatally streptozotocin-induced rats 1.5 days after birth (N-STZ-1.5), oral administrations of fasiglifam (3–30 mg/kg) dose dependently improved glucose tolerance; the effect was greater than that of glibenclamide at maximal effective doses (glucose AUC: fasiglifam, −37.6%; glibenclamide, −12.3%). Although the glucose-lowering effects of glibenclamide (10 mg/kg/day) were completely diminished in N-STZ-1.5 rats after 4 weeks of treatment, effects were maintained in rats receiving fasiglifam (10 mg/kg/day), even after 15 weeks. Fasiglifam (3–10 mg/kg) was still effective in two models desensitized to SUs: 15-week glibenclamide-treated N-STZ-1.5 rats and aged Zucker diabetic fatty (ZDF) rats. Acute administration of fasiglifam (3 mg/kg) and glimepiride (10 mg/kg) in combination additively decreased glucose AUC (fasiglifam, −25.3%; glimepiride, −20.0%; combination, −43.1%). Although glimepiride (10 mg/kg) decreased plasma glucose below normal in nonfasted control rats, fasiglifam (3 mg/kg) maintained normoglycemia, and no further exaggeration of hypoglycemia was observed with combination treatment. These results indicate that GPR40 agonists could be more effective and durable than SUs. Our results also provide new insights into GPR40 pharmacology and rationale for the use of GPR40 agonists in diabetic patients with SU failure.

Fasiglifam (TAK‐875) has dual potentiating mechanisms via Gαq‐GPR40/FFAR1 signaling branches on glucose‐dependent insulin secretion

Fasiglifam (TAK‐875) is a free fatty acid receptor 1 (FFAR1)/G‐protein–coupled receptor 40 (GPR40) agonist that improves glycemic control in type 2 diabetes with minimum risk of hypoglycemia. Fasiglifam potentiates glucose‐stimulated insulin secretion (GSIS) from pancreatic β‐cells glucose dependently, although the precise mechanism underlying the glucose dependency still remains unknown. Here, we investigated key cross‐talk between the GSIS pathway and FFAR1 signaling, and Ca2+ dynamics using mouse insulinoma MIN6 cells. We demonstrated that the glucose‐dependent insulinotropic effect of fasiglifam required membrane depolarization and that fasiglifam induced a glucose‐dependent increase in intracellular Ca2+ level and amplification of Ca2+ oscillations. This differed from the sulfonylurea glimepiride that induced changes in Ca2+ dynamics glucose independently. Stimulation with cell‐permeable analogs of IP3 or diacylglycerol (DAG), downstream second messengers of Gαq‐FFAR1, augmented GSIS similar to fasiglifam, indicating their individual roles in the potentiation of GSIS pathway. Intriguingly, the IP3 analog triggered similar Ca2+ dynamics to fasiglifam, whereas the DAG analog had no effect. Despite the lack of an effect on Ca2+ dynamics, the DAG analog elicited synergistic effects on insulin secretion with Ca2+ influx evoked by an L‐type voltage‐dependent calcium channel opener that mimics glucose‐dependent Ca2+ dynamics. These results indicate that the Gαq signaling activated by fasiglifam enhances GSIS pathway via dual potentiating mechanisms in which IP3 amplifies glucose‐induced Ca2+ oscillations and DAG/protein kinase C (PKC) augments downstream secretory mechanisms independent of Ca2+ oscillations.

Antihypertrophic Effects of Small Molecules that Maintain Mitochondrial ATP Levels Under Hypoxia

Since impaired mitochondrial ATP production in cardiomyocytes is thought to lead to heart failure, a drug that protects mitochondria and improves ATP production under disease conditions would be an attractive treatment option. In this study, we identified small-molecule drugs, including the anti-parasitic agent, ivermectin, that maintain mitochondrial ATP levels under hypoxia in cardiomyocytes. Mechanistically, transcriptomic analysis and gene silencing experiments revealed that ivermectin increased mitochondrial ATP production by inducing Cox6a2, a subunit of the mitochondrial respiratory chain. Furthermore, ivermectin inhibited the hypertrophic response of human induced pluripotent stem cell-derived cardiomyocytes. Pharmacological inhibition of importin β, one of the targets of ivermectin, exhibited protection against mitochondrial ATP decline and cardiomyocyte hypertrophy. These findings indicate that maintaining mitochondrial ATP under hypoxia may prevent hypertrophy and improve cardiac function, providing therapeutic options for mitochondrial dysfunction.