SP600125 (JNK Inhibitor II) is a JNK inhibitor that inhibits JNK1, JNK2, and JNK3 (IC50=40/40/90 nM) with oral potency, reversibility, and ATP-competitive properties. SP600125 inhibits autophagy and induces apoptosis.

JNK2:40 nM (cell free), JNK1:40 nM (cell free), JNK3:90 nM (cell free)

方法：小鼠肺癌细胞 LLC 和小鼠肿瘤细胞 4T1 用 SP600125 (3-10 μM) 处理 72 h，使用 MTT assay 检测细胞活力。
结果：SP600125 剂量依赖性抑制 LLC 和 4T1 细胞生长，IC50 为 8.14 μM 和 7.37 μM。[1]
方法：Jurkat T 细胞 用 SP600125 (1-50 μM) 预处理 10 min，随后用 PMA (50 ng/mL)、anti-CD3 (0.5 μg/mL) 和 anti-CD28 (2 μg/mL) 刺激 30 min，使用 Western Blot 方法检测靶点蛋白表达水平。
结果：SP600125 以 5-10 μM 的 IC50 阻断 c-Jun 的磷酸化。在 50 μM 的浓度下，SP600125 不阻断 ERK 的磷酸化，也不抑制 IκBα 的降解。在 50 μM 时观察到 磷酸-p38 和 ATF2 的部分抑制，但在 25 μM 时没有观察到。[2]

方法：为检测体内对 TNF-α 的抑制活性，将 SP600125 (7.5-30 mg/kg，30% PEG-400/20% polypropylene glycol/15% Cremophor EL/5% ethanol/30% saline) 静脉注射或口服给药给 CD-1 小鼠，15 min 后注射 LPS 诱导的 TNF-α 表达。
结果：静脉注射 15 或 30 mg/kg SP600125 可显著抑制 TNF-α 血清水平，而口服给药剂量依赖性阻断 TNF-α 表达，每次口服 30 mg/kg 可观察到显著抑制作用。[2]
方法：为检测体内抗肿瘤活性，将 SP600125 (5 mg/kg) 和 C-2 (10 mg/kg) 腹腔注射给携带膀胱癌肿瘤 BIU87 的 nude 小鼠，每天一次，持续二十一天。
结果：C-2 治疗抑制了肿瘤的生长，C-2/SP600125 组肿瘤显著低于用载体或 C-2 单独治疗的小鼠。[3]

Multiarray plate screening of mRNA was performed by High Throughput Genomics. In brief, cell lysates were prepared by using a single-step proprietary lysis buffer. Lysates were incubated with a 16-gene capture array manufactured into each well of a 96-well plate. Detection was by luminescence and was performed by HTG. SDs for triplicate samples were typically 3–8% for samples with high levels of gene expression and 15–25% for samples with very low (near-threshold) levels of cytokine gene expression [1].

Mouse LPS/TNF assay was performed as follows: Female CD-1 mice (8–10 weeks of age) were dosed i.v. or per os with SP600125 in PPCES vehicle (30% PEG-400/20% polypropylene glycol/15% Cremophor EL/5% ethanol/30% saline), final volume of 5 ml/kg, 15 min before i.v. injection with LPS in saline (0.5 mg/kg; Escherichia coli 055:B5). At 90 min, a terminal bleed was obtained from the abdominal vena cava, and the serum was recovered. Samples were analyzed for mouse TNF-α by using an ELISA. The in-life phase of the thymocyte apoptosis assay was performed in female C57BL/6 mice. SP600125 was administered at 0, 12, 24, and 36 h, 15 mg/kg s.c. in PPCES vehicle. Anti-CD3 (50 μg) i.p. (clone 145-2C11) was administered as a single dose immediately after SP600125 at time 0. After 48 h, mice were killed, and the thymus was dissected for thymocyte isolation. Treated and untreated mice thymuses were excised and immediately placed in complete medium (RPMI medium 1640 with 10% FBS, penicillin/streptomycin, and l-glutamine) on ice. Each thymus was then pressed between the frosted ends of 2 microscope slides to form a single cell suspension and collected through a 30 μm nylon mesh. Cells were stained for cell surface CD4 and CD8 and apoptosis and measured by flow cytometry [1].