AS1517499 is a potent STAT6 inhibitor with IC50 of 21 nM

STAT6:21 nM

AS1517499具有强大的STAT6抑制作用，IC50值为21 nM。它还能抑制小鼠脾脏T细胞在IL-4诱导下向Th2分化的过程，其IC50值为2.3 nM，同时不影响IL-12诱导的辅助性T细胞1（Th1）分化。AS1517499选择性地抑制Th2分化，而不影响Th1分化[1]。在培养的人类BSM细胞中，IL-13（100 ng/mL）会导致STAT6的磷酸化和RhoA的上调，RhoA是负责平滑肌收缩的Ca2+敏感性的单核苷酸酶：这两个事件均可通过与AS1517499（100 nM）共孵育而被抑制[2]。

在对BALB/c小鼠进行主动致敏并反复用卵清蛋白抗原挑战后，观察到最后一次抗原挑战后小鼠支气管肺泡灌洗液中IL-13水平增加以及支气管组织中STAT6的磷酸化。这些小鼠对乙酰胆碱的平滑肌收缩能力增强，并伴随着支气管组织中RhoA的上调。每次接触卵清蛋白前1小时，腹腔注射AS1517499（10 mg/kg）几乎完全抑制了由抗原引起的RhoA上调和BSM过度反应性[2]。

Normal human BSM cells (hBSMCs) are maintained in SmBM medium supplemented with 5% fetal bovine serum, 0.5 ng/mL human epidermal growth factor (hEGF), 5 μg/mL insulin, 2 ng/mL human fibroblast growth factor-basic (hFGFb), 50 μg/mL gentamicin, and 50 ng/mL amphotericin B. Cells are maintained at 37°C in a humidified atmosphere (5% CO2), fed every 48 to 72 hours, and passaged when cells reached 90 to 95% confluence. Then the hBSMCs (passages 7-9) are seeded in 6-well plates and 8-well chamber slides at a density of 3,500 cells/cm2 and, when 80 to 85% confluence observed, cells are cultured without serum for 24 hours before addition of recombin is ant human IL-13. AS1517499 (100 nM) or its vehicle (0.3% DMSO) is treated 30 minutes before the addition of IL-13 (100 ng/mL). In some experiments, AS1517499 is treated 0 (co-incubation), 3, or 12 hours after the addition of IL-13. In another series of experiments, a selective Rho-kinase inhibitor Y-27632 (1 μM) or its vehicle (0.3% DMSO) is also applied 15 minutes before the IL-13 application. At the indicated time after the IL-13 treatment, cells are washed with PBS, immediately collected, and disrupted with 1× SDS sample buffer (250 μL/well), and used for Western blot analyses[2] .

Mice[2]