RIP3K (IC50 = 1.3 nM)

当浓度为 1 μM 时，GSK'872 无法抑制所测试的 300 种人类蛋白激酶中的大部分。直接测试表明它不能抑制RIP1激酶。在 HT-29 细胞中，GSK'872 以浓度依赖性方式抑制 TNF 诱导的坏死性凋亡。与无细胞生化测定相比，基于细胞的测定的 IC50 高 100-1000 倍。在从全血中分离的原代人中性粒细胞中，GSK'872 也能抑制坏死性凋亡。 GSK'872 阻断两种与 RIP1 无关的坏死性凋亡、TLR3 或 DAI 诱导的死亡途径。它会导致 caspase 激活，随后导致细胞凋亡[1]。

与体内缺血损伤后未治疗相比，GSK'872 治疗显着降低了 HIF-1 表达[3]。< br > GSK ' 872可改善脑水肿患者的神经功能[3]
与假手术大鼠相比，SAH大鼠神经功能受损（P < 0.01，图3A），脑含水量明显增加（P < 0.01，图3B）。与对照大鼠相比，GSK 872给药显著改善了神经功能（P < 0.05，图3A），降低了脑含水量（P < 0.01，图3B）。

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注射GSK ' 872可减少SAH后72 h的坏死神经细胞数量[3]
与实验1一致，在72 h时，SAH大鼠的坏死细胞广泛分布（P < 0.001，图3C， D）。注射GSK ' 872后，与对照大鼠相比，坏死细胞数量明显减少（P < 0.01，图3C， D）。

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GSK ' 872降低RIPK3和MLKL蛋白表达，减少坏死神经细胞数量[3]
RIPK3表达显著增加在SAH +车辆在SAH后72 h组与虚假的组(P < 0.001,图4 a, B),葛兰素史克公司872年政府显著降低RIPK3表达相比SAH +车辆组(P < 0.01,图4 a, B)。符合RIPK3表达式,MLKL水平也显著增加在SAH +车辆组(P < 0.001,图4 a, C),但减少了葛兰素史克公司的872年治疗SAH后72 h (P < 0.05,图4 a, C)。

GSK872（也称为 GSK2399872A、GSK872 或 GSK-872）是一种有效的选择性 RIPK3（受体相互作用蛋白激酶 3）抑制剂。它对 RIP3 激酶结构域具有高结合亲和力，IC50 值为 1.8 nM，并且抑制激酶活性，IC50 为 1.3 nM。

RIP3 激酶抑制剂 GSK'843 或 GSK'872 用于在 TNF 处理后，在载体对照 (DMSO) 或其他处理中存在 Z-VAD-fmk 的情况下，以指定浓度处理 3T3-SA 细胞 18 小时。

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细胞活力测定[3]
采用台盼蓝排除法和3-（4,5 -二甲基噻唑-2-基）- 2,5 -二苯基溴化四唑（MTT）法测定细胞活力。在DLM治疗前4小时给予抑制剂[n-乙酰半胱氨酸（NAC）、丁基羟基异素（BHA）、IM54、Bay11-7082、Z-VAD-FMK、caspase-8抑制剂、GSK-872和necrostatin-1 (Nec-1)]治疗。

Experiment II: To study the role of RIPK3 in the pathological process of EBI following SAH, Rats were randomly assigned to the following groups: (1) sham group (n = 24); (2) SAH + vehicle group (n = 24); (3) SAH + GSK’872 (n = 24). Neurological function (n = 24) was evaluated at 24 h and 72 h after operation. Brain edema (n = 6), western blot (n = 6), PI staining (n = 6) and HMGB1 immunofluorescence (n = 6) were evaluated at 72 h after SAH.[3]
In Experiment II, GSK’872 was diluted with 1% DMSO to a concentration of 25 mM, and 6 μL of GSK’872 or diluted DMSO was administrated by a syringe pump at 30 min after SAH as previously described. The coordinates for left lateral ventricle is 1.5 mm right, 0.8 mm anterior to bregma and 3.8 mm deep. Equal volumes of vehicle were given for sham and SAH + vehicle rats at the same time point.[3]

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Dissolved in DMSO (<0.1%) and diluted in saline; 1.9 mmol/kg; i.p.

C57BL/6 mice

[1]. RIP3 induces apoptosis independent of pronecrotic kinase activity. Mol Cell. 2014 Nov 20;56(4):481-95.

[2]. Deltamethrin induced RIPK3-mediated caspase-independent non-apoptotic cell death in rat primary hepatocytes. Biochem Biophys Res Commun. 2016 Oct 14;479(2):217-223.

[3]. Inhibiting of RIPK3 attenuates early brain injury following subarachnoid hemorrhage: Possibly through alleviating necroptosis. Biomed Pharmacother. 2018;107:563-570.

Receptor-interacting protein kinase 3 (RIP3 or RIPK3) has emerged as a central player in necroptosis and a potential target to control inflammatory disease. Here, three selective small-molecule compounds are shown to inhibit RIP3 kinase-dependent necroptosis, although their therapeutic value is undermined by a surprising, concentration-dependent induction of apoptosis. These compounds interact with RIP3 to activate caspase 8 (Casp8) via RHIM-driven recruitment of RIP1 (RIPK1) to assemble a Casp8-FADD-cFLIP complex completely independent of pronecrotic kinase activities and MLKL. RIP3 kinase-dead D161N mutant induces spontaneous apoptosis independent of compound, whereas D161G, D143N, and K51A mutants, like wild-type, only trigger apoptosis when compound is present. Accordingly, RIP3-K51A mutant mice (Rip3(K51A/K51A)) are viable and fertile, in stark contrast to the perinatal lethality of Rip3(D161N/D161N) mice. RIP3 therefore holds both necroptosis and apoptosis in balance through a Ripoptosome-like platform. This work highlights a common mechanism unveiling RHIM-driven apoptosis by therapeutic or genetic perturbation of RIP3.[1]

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Deltamethrin (DLM), a synthetic pyrethroid insecticide, is used all over the world for indoor and field pest management. In the present study, we investigated the elicited pathogenesis of DLM-induced hepatotoxicity in rat primary hepatocytes. DLM-induced cell death was accompanied with increased ROS generation, decreased mitochondrial membrane potential and G2/M arrest. Pre-treatment with N-acetyl cysteine/butylated hydroxyanisole/IM54 could partly rescue hepatocytes suggesting that ROS might play a role in DLM-induced toxicity. Interestingly, DLM treatment resulted in a caspase-independent but non-apoptotic cell death. Pre-treatment with pan-caspase inhibitor (ZVAD-FMK) could not rescue hepatocytes. Unaltered caspase-3 activity and absence of cleaved caspase-3 also corroborated our findings. Further, LDH release and Transmission electron microscopy (TEM) analysis demonstrated that DLM incites membrane disintegrity and necrotic damage. Immunochemical staining revealed an increased expression of inflammatory markers (TNFα, NFκB, iNOS, COX-2) following DLM treatment. Moreover, the enhanced RIPK3 expression in DLM treated groups and prominent rescue from cell death by GSK-872 indicated that DLM exposure could induce programmed necrosis in hepatocytes. The present study demonstrates that DLM could induce hepatotoxicity via non-apoptotic mode of cell death.[2]

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Necroptosis is an inflammatory form of cell death that depends on receptor-interacting serine-threonine kinase 3 (RIPK3) and mixed lineage kinase domain-like (MLKL) and displays the morphological characteristics of necrosis. To date, it is unclear to what extent necroptosis contributes to subarachnoid hemorrhage (SAH) induced brain injury. The present study aimed to investigate the RIPK3-mediated necroptosis and the effects of the RIPK3 selective inhibitor GSK'872 in early brain injury following SAH. After SAH, RIPK3 expression increased as early as 6 h and peaked at 72 h. Double immunofluorescence staining revealed that RIPK3 was mainly located in neurons. Most necrotic cells were neurons, which were further confirmed by TEM. Intracerebroventricular injection of GSK'872 (25 mM) could attenuate brain edema and improve neurological function following SAH and reduce the number of necrotic cells. In addition, GSK'872 could also decrease the protein levels of RIPK3 and MLKL, and cytoplasmic translocation and expression of HMGB1, an important pro-inflammatory protein. Taken together, the current study provides the new evidence that RIPK3-mediated necroptosis is involved in early brain injury and GSK'872 decreases the RIPK3-mediated necroptosis and subsequent cytoplasmic translocation and expression of HMGB1, as well as ameliorates brain edema and neurological deficits.[3]

C19H17N3O2S2

383.49

383.076

C, 59.51; H, 4.47; N, 10.96; O, 8.34; S, 16.72

1346546-69-7

GSK-872 hydrochloride;2703752-81-0

54674134

White to yellow solid powder

1.4±0.1 g/cm3

625.7±55.0 °C at 760 mmHg

332.2±31.5 °C

0.0±1.8 mmHg at 25°C

1.704

3.1

109

1

6

4

26

592

0

S(C1C=CC2C(=C(C=CN=2)NC2C=CC3=C(C=2)N=CS3)C=1)(C(C)C)(=O)=O

ZCDBTQNFAPKACC-UHFFFAOYSA-N

InChI=1S/C19H17N3O2S2/c1-12(2)26(23,24)14-4-5-16-15(10-14)17(7-8-20-16)22-13-3-6-19-18(9-13)21-11-25-19/h3-12H,1-2H3,(H,20,22)

N-(6-propan-2-ylsulfonylquinolin-4-yl)-1,3-benzothiazol-5-amine

GSK2399872A; GSK872; GSK-872; GSK 872; GSK2399872-A; GSK2399872 A; GSK-2399872A; 1346546-69-7; GSK'872; N-(6-(isopropylsulfonyl)quinolin-4-yl)benzo[d]thiazol-5-amine; N-5-benzothiazolyl-6-[(1-methylethyl)sulfonyl]-4-Quinolinamine; C19H17N3O2S2; GSK 872; GSK-2399872 A

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 (6.52 mM) (饱和度未知) 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。
例如，若需制备1 mL的工作液，可将100 μL 25.0 mg/mL澄清DMSO储备液加入900 μL 20% SBE-β-CD生理盐水溶液中，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 2 中的溶解度: ≥ 2.5 mg/mL (6.52 mM) (饱和度未知) 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。
例如，若需制备1 mL的工作液，可将 100 μL 25.0 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 3 中的溶解度: ≥ 2.08 mg/mL (5.42 mM) (饱和度未知) 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。
例如，若需制备1 mL的工作液，可将 100 μL 20.8 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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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。