Fasiglifam (TAK875) is a potent, selective and orally bioavailable GPR40 agonist.

FFA1/GPR40:72 nM.(EC50)

Fasiglifam在CHO-hGPR40細胞中以濃度依賴的方式增加細胞內IP產量，其EC50值為0.072 μM。在CHO細胞中，Fasiglifam以濃度範圍0.1-10 μM呈濃度依賴性提高細胞內IP產量[1]。此外，Fasiglifam在3-30 μM的濃度範圍內，依賴於濃度地增加[Ca2+]i。在10 mM葡萄糖存在的環境下，Fasiglifam以0.001-10 μM的濃度範圍內，依賴於濃度地促進INS-1 833/15細胞中胰島素的分泌[2]。

Fasiglifam（10 mg/kg，p.o.）在ZDF大鼠中提高了血浆胰岛素水平。Fasiglifam（30 mg/kg，p.o.）改善了空腹高血糖，而不影响空腹正常血糖。与改善糖尿病大鼠葡萄糖耐量的剂量相比，Fasiglifam在30 mg/kg的剂量，即3至10倍较高剂量，不会改变具有正常葡萄糖稳态的SD大鼠的空腹血糖水平。同样，Fasiglifam在具有正常空腹血糖水平的SD大鼠中并未显著改变胰岛素分泌[1]。

INS-1 832/13 cells are suspended in RPMI medium containing 11 mM glucose and the supplements described above. These cells are seeded at a density of 2×104?cells/well in a 96-well black plate coated with poly-D-lysine, and 1% BSA and 0.1% DMSO alone (control), palmitic acid (62.5, 125, 250, 500, and 1000 μM), oleic acid (62.5, 125, 250, 500, and 1000 μM), or TAK-875 (6.25, 12.5, 25, 50, and 100 μM) is added to the plate with 1% BSA and 0.1% DMSO, followed by culture for 72 h. After the culture, caspase 3/7 activity is measured with the Apo-one homogeneous caspase 3/7 assay according to the manufacturer's instructions. Fluorescence intensity is measured at an excitation of 485 nm and an emission at 535 nm.

TAK-875 is dissolved in 1% BSA and 0.1% DMSO.? INS-1 832/13 cells are suspended in RPMI medium and seeded in a 96-well plate at a density of 2×104 cells/well; 1% BSA and 0.1% DMSO alone (control), palmitic acid (10, 100, and 1000 μM), oleic acid (10, 100, and 1000 μM), or TAK-875 (1, 10, and 100 μM) is added to the plate. After 72-h culture, medium is discarded, and cells are preincubated for 2 h with KRBH containing 1 mM glucose and 0.2% BSA at 37°C. After discarding of the preincubation buffer, KRBH containing 1 or 20 mM glucose and 0.2% BSA is added, and the plate is further incubated for 2 h. The insulin concentration in the supernatant is measured as described above. To measure intracellular insulin content, INS-1 832/13 cells are exposed to 1% BSA and 0.1% DMSO alone (control), palmitic acid (1000 μM), oleic acid (1000 μM), or TAK-875 (100 μM) with 1% BSA and 0.1% DMSO. After incubation, cells are washed once with phosphate-buffered saline, and acid-ethanol solution is added to each well, followed by sonication on ice. Intracellular insulin is extracted by overnight incubation at ?30°C, followed by separation of supernatant by centrifugation at 12,000 rpm×5 min at 4°C.