SB 415286 is a potent GSK3α inhibitor with IC50/Ki of 78 nM/31 nM with equally effective inhibition of GSK-3β.

GSK-3α:78 nM, GSK-3β:31 nM(ki)

SB 415286以Ki为31nM的方式与ATP竞争性抑制GSK3α，并对GSK3β显示出类似的效力。SB 415286对其他24种蛋白激酶几乎没有活性，其IC50 > 10 μM。SB 415286在Chang人肝细胞系中刺激糖原合成，EC50为2.9 μM，并在HEK293细胞中诱导β-连环蛋白-LEF/TCF受控的报告基因表达。[1] SB 415286通过与抑制GSK-3活性和调节GSK-3底物tau和β-连环蛋白相关的方式，浓度依赖性地保护培养中的中枢和周围神经系统神经元免遭因PI3激酶通路活性降低而引起的死亡。[2] 在L6肌管中，SB 415286诱导的GS激活水平(6.8倍)远大于胰岛素(4.2倍)或Li(4倍)引起的激活。[3] SB 415286 (10 μM)抑制雷帕霉素诱导的cyclin D1下调，并阻断了雷帕霉素和紫杉醇诱导的细胞凋亡，提示GSK3β在雷帕霉素介导的紫杉醇增敏中扮演关键角色。[4] SB 415286通过稳定β-连环蛋白的方式，呈剂量依赖性地防止柯萨奇病毒引起的细胞死亡。[5] SB 415286对B65大鼠神经母细胞瘤细胞和神经元中过氧化氢诱导的细胞死亡具有保护作用，而锂并不减轻过氧化氢的毒性作用。[7] SB 415286处理增强了TRAIL和CH-11在HepG2细胞中诱导的细胞凋亡。[8] 通过激活内源性途径，SB 415286的GSK-3抑制作用导致多发性骨髓瘤(MM)细胞生长停滞和凋亡。[9] SB 415286降低Neuro-2A细胞的存活率，并诱导细胞在细胞周期的G2/M期积累及随后的凋亡。[10]

SB 415286 (~10 mg/kg，每日两次)能够降低大鼠三硝基苯磺酸(TNBS)引起的结肠炎症的程度和范围，同时减轻体重下降，这一效果与NF-κB活动的下调有关，NF-κB参与了促炎介质的产生。[6] 在裸鼠体内，1 mg/kg的SB 415286处理显著延缓了Neuro-2A细胞的生长。[10]

GSK-3 activity assay: GSK-3 kinase activity is measured, in the presence of various concentrations of SB 415286, in a reaction mixture containing final concentrations of: 1 nM human GSK3α or rabbit GSK3α; 50 mM MOPS pH 7.0; 0.2 mM EDTA; 10 mM Mg-acetate; 7.5 mM L-mercaptoethanol; 5% (w/v) glycerol; 0.01% (w/v) Tween-20; 10% (v/v) DMSO; 28 μM GS-2 peptide substrate. The GS-2 peptide sequence corresponds to a region of glycogen synthase that is phosphorylated by GSK-3. The assay is initiated by the addition of 0.34 μCi [33P]γ-ATP (IC50 determinations) or 2.7 μCi [33P]γ-ATP (Ki determinations). The total ATP concentration is 10 μM (IC50 determinations) or ranged from 0 to 45 μM (Ki determinations). Following 30 minutes incubation at room temperature the assay is stopped by the addition of one third assay volume of 2.5% (v/v) H3PO4 containing 21 mM ATP. Samples are spotted onto P30 phosphocellulose mats and these are washed six times in 0.5% (v/v) H3PO4. The filter mats are sealed into sample bags containing Wallac betaplate scintillation fluid. 33P incorporation into the substrate peptide is determined by counting the mats in a Wallac microbeta scintillation counter.

Cells are exposed to different concentrations of SB 415286 for 48 or 72 hours in 96-flat well plates. After 48 or 72 hours, [3H]thymidine is added to the cultures (10 μCi/well) for the last 12 hours. The [3H]thymidine incorporation is evaluated by scintillation counting by using a top count β-counter. Apoptosis is assessed by annexin V/Propidium Iodide staining or by detection of mitochondrial membrane potential. Cell death is evaluated by the analysis of Forward/Side scatter fluorescence changes. Fluorescence Activated Cell Sorting (FACS) analysis is performed using a FACS-Calibur Cell Cytometer. (Only for Reference)