p38 (IC50 = 50 nM); p38β2 (IC50 = 500 nM)

SB203580 的 IC50 为 3-5 μm，当 IL-2 存在时，可抑制小鼠 CT6 T 细胞、BAF F7 B 细胞或原代人 T 细胞的增殖。虽然所需浓度稍高且 IC50 高于 10 μm，但 SB203580 也能抑制 IL-2 诱导的 p70S6 激酶激活。 SB203580 的 IC50 在 3-10 μm 范围内，还以剂量依赖性方式抑制 PDK1 的活性。 [1] SB203580 阻断 p38-MAPK 对 MAPKAPK2 的刺激的 IC50 约为 0.07 μm，而阻断总 SAPK/JNK 活性的 IC50 为 3-10 μm。较高浓度的 SB203580 会导致 ERK 通路被激活，从而提高 NF-κB 的转录活性。[2] SB203580 诱导人肝细胞癌细胞 (HCC) 发生自噬。[3]

SB203580 在体内模型中保护猪心肌免受缺血性损伤。[4] SB203580 可有效预防和治疗 MRL/lpr 小鼠的系统性红斑狼疮 (SLE)。[5]

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SB203580治疗的MRL/lpr小鼠可预防蛋白尿。[5]
SB203580对MRL/lpr小鼠的ALT和AST没有影响。[5]
SB203580治疗的MRL/lpr小鼠的BUN降低，但Cr没有降低。 SB203580治疗的MRL/lpr小鼠的肾脏重量减少，但脾脏重量没有减少。[5]
SB203580治疗的MRL/lpr小鼠的肾脏病理变化减弱。[5]
SB203580治疗的MRL/lpr小鼠的肝脏病理变化得到缓解。[5]
SB203580治疗的MRL/lpr小鼠的脾脏病理变化得到缓解。[5]
SB203580治疗的MRL/lpr小鼠肾小球IgG、IgM、IgA和C3沉积减少。[5]

细胞受体激酶磷酸化测定：将 4μg 羊抗 PKBα 固定在 25 μL Protein G-Sepharose 上过夜（或 1.5 小时），并用 Buffer A（50 mm Tris，pH 7.5，1 mm EDTA，1 mm EGTA，0.5 mm Na3VO4、0.1% β-巯基乙醇、1% Triton X-100、50 mm 氟化钠、5 mm 焦磷酸钠、0.1 mm 苯甲基磺酰氟、1 μg/mL 抑肽酶、胃酶抑素、亮肽素和 1 μm 微囊藻毒素）。然后将固定化的抗 PKB 与 0.5 ml 裂解液（来自 5 × 106 个细胞）一起孵育 1.5 小时，在补充有 0.5 m NaCl 的 0.5 mL 缓冲液 A 中洗涤五次，在 0.5 mL 缓冲液 B（50 mm）中洗涤两次。 Tris-HCl，pH 7.5，0.03% (w/v) Brij-35，0.1 mm EGTA 和 0.1% β-巯基乙醇），并用 100 μl 测定稀释缓冲液两次； 5× 测定稀释缓冲液为 100 mm MOPS，pH 7.2，125 mm β-甘油磷酸盐，25 mm EGTA，5 mm 原钒酸钠，5 mm DTT。 PKB 酶免疫复合物补充有 10 μL 测定稀释缓冲液、40 μm 蛋白激酶 A 抑制肽、100 μm PKB 特异性底物肽和 10 μCi 的 [γ-32P]ATP。室温下振荡反应 20 分钟，然后脉冲旋转样品，将 40 μL 反应体积转移至另一管中，加入 20 μL 40% 三氯乙酸终止反应。混合并在室温下孵育 5 分钟后，将 40 μL 混合物转移到 P81 磷酸纤维素纸上并结合 30 秒。 P81片在0.75%磷酸中清洗3次后，在室温下用丙酮清洗。然后使用闪烁计数对 γ-32P 的掺入进行定量。

为了使 CT6 细胞和 BA/F3 F7 细胞静息，将它们在 RPMI 中洗涤 3 次，并在含有 5% 胎牛血清的 RPMI 中培养过夜，不添加生长因子、抗生素或 β-巯基乙醇。使用 SB203580 或载体对照在 2 mL RPMI、5% 胎牛血清和 2–5 × 106 个静息 CT6 细胞上进行预孵育，如图图例所示。然后，用 20 ng/ml 重组人 IL-2 在 37°C 下刺激细胞 5 分钟，在微型离心机中沉淀 30 秒，吸出培养基，并在适当的缓冲液中裂解沉淀。 BA/F3 细胞维持在含有谷氨酰胺的 RPMI 中，另外补充有 5% 胎牛血清和 0.2 μg/mL G418，并稳定表达 IL-2 受体 β 链的缺失突变体。然后彻底清洗细胞，静置过夜，然后再次清洗，然后用 IL-2 激活。此类细胞制剂含有 >90% T 细胞。在细胞增殖测定中测量[3H]胸苷的掺入。

Systemic lupus erythematosus (SLE) are established in female MRL/lpr mice and female C57BL/6 mice
0.1 M/day
Orally administered

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Female MRL/lpr mice were randomized into two groups (n = 10 per group) and were fed control diet (named as group 2 in the following) or diet with SB203580 (named as group 3 in the following) starting at the age of 14 weeks and continuing for up to 22 weeks. Adezmapimod (SB203580) was dissolved in drinking water (250 μmol/L), was orally administered (0.4 ml/day). Ten C57BL/6 female mice were used as negative controls (named as group 1 in the following). Two mice in MRL/lpr group 2 were dead at 16 weeks and 18 weeks of age respectively. Two mice in MRL/lpr group 3 were dead at 19 weeks of age. Significant increase of urine protein (300–2000 mg/dl) was found in each mouse before death, indicating a probable renal failure be the cause of death. Ultimately, 10 mice in group 1, 8 mice in group 2 and group 3 were included in statistical analysis.[5]

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Systemic lupus erythematosus (SLE) is an autoimmune disease accompanying excessive inflammatory responses in a wide range of organs. Abnormal activation of p38 MAPK has been postulated to contribute to the inflammation of SLE, leading to progressive tissue and organ damages to develop lupus nephritis and autoimmune hepatitis. In order to determine whether p38 MAPK inhibitor is effective in mouse model of SLE, a specific inhibitor of p38 MAPK Adezmapimod (SB203580) was orally administrated to MRL/lpr mice aged from 14 to 22 weeks. Renal and hepatic functions, as well as pathologic changes of important organs including kidney, liver and spleen of MRL/lpr mice were evaluated. As a result, we showed that SB203580 improved renal function by decreasing the levels of proteinuria and serum BUN, ameliorating the pathologic changes of kidney and reducing Ig and C(3) depositions in the kidney. Hepatocytes necrosis, recruitment and proliferation of leucocytes in liver and spleen were found to be inhibited by administration of SB203580. Therefore, p38 MAPK activation may be partially responsible for escalating autoimmune renal, hepatic and splenic destruction, and its inhibitor may lighten the autoimmune attack in these important organs and improve renal function. Our study reveals that the selective blockade of p38 MAPK is effective to prevent and treat the disease in this model of SLE.[5]

[1]. J Biol Chem . 2000 Mar 10;275(10):7395-402.

[2]. Br J Pharmacol . 2000 Sep;131(1):99-107.

[3]. ACS Med Chem Lett . 2017 Feb 8;8(3):316-320.

[4]. J Cardiovasc Pharmacol . 2000 Mar;35(3):474-83.

[5]. Int Immunopharmacol . 2011 Sep;11(9):1319-26.

SB 203580 is a member of the class of imidazoles carrying 4-methylsulfinylphenyl, 4-pyridyl and 4-fluorophenyl substituents at positions 2, 4 and 5 respectively. An inhibitor of mitogen-activated protein kinase. It has a role as an EC 2.7.11.24 (mitogen-activated protein kinase) inhibitor, a Hsp90 inhibitor, a neuroprotective agent, an EC 2.7.11.1 (non-specific serine/threonine protein kinase) inhibitor and a geroprotector. It is a member of imidazoles, a member of monofluorobenzenes, a member of pyridines and a sulfoxide.
4-(4-Fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole has been reported in Annulohypoxylon truncatum, Eleutherococcus divaricatus, and other organisms with data available.

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Pyridinyl imidazole inhibitors, particularly SB203580, have been widely used to elucidate the roles of p38 mitogen-activated protein (MAP) kinase (p38/HOG/SAPKII) in a wide array of biological systems. Studies by this group and others have shown that SB203580 can have antiproliferative activity on cytokine-activated lymphocytes. However, we recently reported that the antiproliferative effects of SB203580 were unrelated to p38 MAP kinase activity. This present study now shows that SB203580 can inhibit the key cell cycle event of retinoblastoma protein phosphorylation in interleukin-2-stimulated T cells. Studies on the proximal regulator of this event, the phosphatidylinositol 3-kinase/protein kinase B (PKB)(Akt/Rac) kinase pathway, showed that SB203580 blocked the phosphorylation and activation of PKB by inhibiting the PKB kinase, phosphoinositide-dependent protein kinase 1. The concentrations of SB203580 required to block PKB phosphorylation (IC(50) 3-5 microM) are only approximately 10-fold higher than those required to inhibit p38 MAP kinase (IC(50) 0.3-0.5 microM). These data define a new activity for this drug and would suggest that extreme caution should be taken when interpreting data where SB203580 has been used at concentrations above 1-2 microM.[1]

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In the present study we investigated a possible role for the p38 mitogen-activated protein (MAP) kinase pathway in mediating nuclear factor-kappa B (NF-kappaB) transcriptional activity in the erythroleukaemic cell line TF-1. TF-1 cells stimulated with the phosphatase inhibitor okadaic acid (OA) demonstrated enhanced NF-kappaB and GAL4p65-regulated transcriptional activity which was associated with elevated p38 phosphorylation. However, pretreatment with the p38 MAPK specific inhibitor SB203580 (1 microM) or overexpression of kinase-deficient mutants of MKK3 or MKK6 did not affect OA-enhanced NF-kappaB transcriptional potency, as determined in transient transfection assays. In fact, 5 and 10 microM SB203580 enhanced rather than inhibited NF-kappaB-mediated promoter activity by 2 fold, which was independent of phosphorylation of the p65 subunit. The SB203580-mediated increase in NF-kappaB transcriptional activity was associated with enhanced phosphorylation of extracellular signal-regulated kinase (ERK)1/2 and c-Jun N-terminal kinase (JNK), but not p38 kinase. Overexpression of kinase-deficient mutants belonging to the ERK1/2, JNK, and p38 pathways showed that only dominant-negative Raf-1 abrogated SB203580-enhanced NF-kappaB activity. This would implicate the involvement of the ERK1/2 pathway in the enhancing effects of SB203580 on NF-kappaB-mediated gene transcription. This study demonstrates that the p38 MAP kinase pathway is not involved in the OA-induced activation of NF-kappaB. SB203580 at higher concentrations activates the ERK pathway, which subsequently enhances NF-kappaB transcriptional activity.[2]

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SB203580 is a well-known inhibitor of p38 mitogen-activated protein kinase (MAPK). However, it can suppress cell proliferation in a p38 MAPK independent manner. The inhibitory mechanism remains unknown. Here, we showed that SB203580 induced autophagy in human hepatocellular carcinoma (HCC) cells. SB203580 increased GFP-LC3-positive cells with GFP-LC3 dots, induced accumulation of autophagosomes, and elevated the levels of microtubule-associated protein light chain 3 and Beclin 1. It stimulated the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) and p53, but inhibited the phosphorylation of death-associated protein kinase (DAPK). Inhibition of AMPK, p53, or DAPK attenuated SB203580-induced autophagy. AMPK activation appeared to predate the DAPK signal. The activation of both AMPK and DAPK prompted the phosphorylation of p53 and enhanced Beclin 1 expression. Neither the downregulation of p38 MAPK by its siRNA or chemical inhibitor nor the upregulation of p38 MAPK by p38 MAPK DNA transfection affected B203580-induced autophagy. Collectively, the findings demonstrate a novel function of SB203580 to induce autophagy via activating AMPK and DAPK but independent of p38 MAPK. The induction of autophagy can thus account for the antiproliferative effect of SB203580 in HCC cells.[3]

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We report that SB203580 (SB), a specific inhibitor of p38-MAPK, protects pig myocardium against ischemic injury in an in vivo model. SB was applied by local infusion into the subsequently ischemic myocardium for 60 min before a 60-min period of coronary occlusion followed by 60-min reperfusion (index ischemia). Infarct size was reduced from a control value of 69.3 +/- 2.7% to 36.8 +/- 3.7%. When SB was infused systemically for 10 min before index ischemia, infarct size was reduced to 36.1 +/- 5.6%. We measured the content of phosphorylated p38-MAPK after systemic infusion of SB and Krebs-Henseleit buffer (KHB; negative control) and during the subsequent ischemic period using an antibody that reacts specifically with dual-phosphorylated p38-MAPK (Thr180/ Tyr182). Ischemia with and without SB significantly increased phospho-p38-MAPK, with a maximum reached at 20 min but was less at 30 and 45 min under the influence of the inhibitor. The systemic infusion of SB for 10 min before index ischemia did not significantly change the p38-MAPK activities (compared with vehicle, studied by in-gel phosphorylation) < or =20 min of ischemia, but activities were reduced at 30 and 45 min. Measurements of p38-MAPK activities in situations in which SB was present during in-gel phosphorylation showed significant inhibition of p38-MAPK activities. The systemic infusion of SB significantly inhibited the ischemia-induced phosphorylation of nuclear activating transcription factor 2 (ATF-2). Using a specific ATF-2 antibody, we did not observe significant changes in ATF-2 abundance when nuclear fractions from untreated, KHB-, and SB-treated tissues were compared. We investigated also the effect of local and systemic infusion of SB on the cardioprotection induced by ischemic preconditioning (IP). The infusions (local or systemic) of SB before and during the IP protocol did not influence the infarct size reduction mediated by IP. The observed protection of the myocardium against ischemic damage by SB points to the negative role of the p38-MAPK pathway during ischemia.[4]

C21H16FN3OS

377.43

377.099

C, 66.83; H, 4.27; F, 5.03; N, 11.13; O, 4.24; S, 8.49

152121-47-6

Adezmapimod hydrochloride;869185-85-3

176155

White to light yellow solid powder

1.4±0.1 g/cm3

615.6±55.0 °C at 760 mmHg

249 - 250ºC

326.1±31.5 °C

0.0±1.7 mmHg at 25°C

1.715

4.1

77.85

1

5

4

27

500

0

S(C([H])([H])[H])(C1C([H])=C([H])C(=C([H])C=1[H])C1=NC(C2C([H])=C([H])C(=C([H])C=2[H])F)=C(C2C([H])=C([H])N=C([H])C=2[H])N1[H])=O

CDMGBJANTYXAIV-UHFFFAOYSA-N

InChI=1S/C21H16FN3OS/c1-27(26)18-8-4-16(5-9-18)21-24-19(14-2-6-17(22)7-3-14)20(25-21)15-10-12-23-13-11-15/h2-13H,1H3,(H,24,25)

4-[4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-1H-imidazol-5-yl]pyridine;hydrochloride

RWJ 64809; PB 203580; Adezmapimod; 4-(4-Fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole; 4-[4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-1H-imidazol-5-yl]pyridine; 4-(4-(4-fluorophenyl)-2-(4-(methylsulfinyl)phenyl)-1H-imidazol-5-yl)pyridine; RWJ64809; SB203580; SB203580; SB 203580; RWJ-64809; PB-203580; PB203580

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

配方 1 中的溶解度: ≥ 2.5 mg/mL (6.62 mM) (饱和度未知) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 2 中的溶解度: 2 mg/mL (5.30 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液; 超声助溶.
例如，若需制备1 mL的工作液，可将 100 μL 20.0 mg/mL 澄清的 DMSO 储备液加入到400 μL PEG300中，混匀；再向上述溶液中加入50 μL Tween-80，混匀；然后加入450 μL 生理盐水定容至1 mL。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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配方 3 中的溶解度: 2 mg/mL (5.30 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液; 超声助溶.
例如，若需制备1 mL的工作液，可将 100 μL 20.0mg/mL澄清的DMSO储备液加入到900μL 20％SBE-β-CD生理盐水中，混匀。
*20% SBE-β-CD 生理盐水溶液的制备（4°C，1 周）：将 2 g SBE-β-CD 溶解于 10 mL 生理盐水中，得到澄清溶液。

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配方 4 中的溶解度: 2 mg/mL (5.30 mM) in 10% DMSO + 90% Corn Oil (这些助溶剂从左到右依次添加，逐一添加), 澄清溶液; 超声助溶.
例如，若需制备1 mL的工作液，您可以将 100 μL 20.0 mg/mL 澄清 DMSO 储备液加入到 900 μL 玉米油中并混合均匀。

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配方 5 中的溶解度: 4% DMSO+30% PEG 300+5% Tween 80+ddH2O: 5mg/mL

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配方 6 中的溶解度: 16.67 mg/mL (44.17 mM) in 0.5% CMC-Na/saline water (这些助溶剂从左到右依次添加，逐一添加), 悬浊液; 超声助溶。
*生理盐水的制备：将 0.9 g 氯化钠溶解在 100 mL ddH₂O中，得到澄清溶液。

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请根据您的实验动物和给药方式选择适当的溶解配方/方案：
1、请先配制澄清的储备液(如：用DMSO配置50 或 100 mg/mL母液(储备液));
2、取适量母液，按从左到右的顺序依次添加助溶剂，澄清后再加入下一助溶剂。以 下列配方为例说明 (注意此配方只用于说明，并不一定代表此产品 的实际溶解配方):
10% DMSO → 40% PEG300 → 5% Tween-80 → 45% ddH2O (或 saline);
假设最终工作液的体积为 1 mL, 浓度为5 mg/mL: 取 100 μL 50 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀/澄清；向上述体系中加入50 μL Tween-80，混合均匀/澄清；然后继续加入450 μL ddH2O (或 saline)定容至 1 mL；
3、溶剂前显示的百分比是指该溶剂在最终溶液/工作液中的体积所占比例;
4、 如产品在配制过程中出现沉淀/析出，可通过加热(≤50℃)或超声的方式助溶;
5、为保证最佳实验结果，工作液请现配现用！
6、如不确定怎么将母液配置成体内动物实验的工作液，请查看说明书或联系我们;
7、 以上所有助溶剂都可在 Invivochem.cn网站购买。