SB505124 是TGF-βI型受体 (ALK4，ALK5，ALK7) 的选择性抑制剂，能够抑制 ALK4 (IC50:129 nM)，ALK5 (IC50:47 nM)，对 ALK1，2，3 或 6 无影响。

SB505124 is a selective inhibitor of TGFβR for ALK4, ALK5.

在兔GFS模型中,SB505124降低了GFS手术部位的眼内压（IOP）水平并减少了结膜下细胞浸润和瘢痕形成.在他克莫司（TAC）治疗的小鼠和FK12EC KO小鼠中,SB505124抑制内皮细胞TGF-β受体的活化和肾小动脉玻璃样变性的诱导.

SB505124被鉴定为ALK4和ALK5的可逆ATP竞争性和选择性ALK抑制剂。SB-505124抑制紧密相关的ALK4,IC50为129 nM,选择性比作用于ALK5低2.5倍。SB-505124浓度高达10 μM时也不抑制ALK2。SB-505124抑制COS-1细胞内源性Smad2磷酸化。ALK4和ALK5激活Smad1和Smad2,加入SB-505124抑制激活。SB-505124作用于HepG2人类肝癌细胞,C2C12小鼠成肌细胞和Mv1Lu水貂肺细胞,浓度依赖性抑制转化生长因子-β诱导的Smad2磷酸化。SB-505124有效抑制转化生长因子-β和活化素诱导CAGA12-荧光素酶和ARE-荧光素酶受体结构的活力,这种作用存在浓度依赖性。在人脐静脉内皮细胞（HUVEC）中,SB505124（500 nM）阻断TGF-β1对F-actin组装的影响,并阻止TGF-β诱导的ROS产生。

In Vitro Protein Kinase Assay : Kinase assays are performed as described by Laping et al., 2002 using the kinase domain of ALK5 and full-length N-terminal fused GST-Smad3. Kinase assays are performed with 65 nM GST-ALK5 and 184 nM GST-Smad3 in 50 mM HEPES, 5 mM MgCl2, 1 mM CaCl2, 1 mM dithiothreitol, and 3 μM ATP. Reactions are incubated with 0.5 μCi of [33P]γATP for 3 hours at 30 °C. Phosphorylated protein is captured on P-81 paper , washed with 0.5% phosphoric acid, and counted by liquid scintillation. Alternatively, Smad3 or Smad1 protein is also coated onto FlashPlate Sterile Basic Microplates. Kinase assays are then performed in FlashPlates with same assay conditions using either the kinase domain of ALK5 with Smad3 as substrate or the kinase domain of ALK6 (BMP receptor) with Smad1 as substrate. Plates are washed three times with phosphate buffer and counted by TopCount.

Cell viability is measured as described by Laping et al., 2002 or by using the modified tetrazolium salt WST-1. XTT assay: The cells are serum-deprived for 24 hours and then treated with SB505124 for 48 hours to assess the cellular toxicity. Cell viability is determined by incubating cells for 4 hours with XTT labeling and electron coupling reagent according to the manufacturer's directions. Live cells with active mitochondria produce an orange-colored product, formazan, which is detected using a plate reader at between A450 nm and A500 nm with a reference wavelength greater than 600 nm. The absorbance values correlate with the number of viable cells. Modified tetrazolium salt WST-1: Approximately 2000 cells are seeded in 96-well dishes in 100 μL of 0.2% FBS phenol red-free media overnight. The cells are treated with 50 μL of SB505124 (to achieve the final concentrations indicated) for 30 minutes before being treated with or without TGF-β1 and TNF-α to a final volume of 200 μL. Cell growth is measured at the indicated time points by incubating each well with 10 μL of WST-1 for 3 hours at 37 °C. Metabolically active cells cleave WST-1 to water-soluble formazan, which is directly quantitated with an enzyme-linked immunosorbent assay plate reader. Each experiment is done at least twice, and treatment for each cell line is done in triplicate.(Only for Reference)

694433-59-5

C20H21N3O2

335.407

1eq. HCl:33.5 mg/mL (100 mM)
DMSO:33.5 mg/mL (100 mM)
Powder: -20°C for 3 years
In solvent: -80°C for 2 years