Li J-X, Feng J-M, Wang Y, Li X-H, Chen X-X, Su Y, Shen Y-Y, Chen Y, Xiong B, Yang C-H, Ding J, Miao Z-H
Division of Antitumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Zhang Jiang Hi-Tech Park, Shanghai 201203, China.
Department of Medicinal Chemistry, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Zhang Jiang Hi-Tech Park, Shanghai 201203, China.
Cell Death Dis. 2014 Jun 5;5(6):e1278. doi: 10.1038/cddis.2014.241.
Receptor-interacting protein (RIP)3 is a critical regulator of necroptosis and has been demonstrated to be associated with various diseases, suggesting that its inhibitors are promising in the clinic. However, there have been few RIP3 inhibitors reported as yet. B-Raf(V600E) inhibitors are an important anticancer drug class for metastatic melanoma therapy. In this study, we found that 6 B-Raf inhibitors could inhibit RIP3 enzymatic activity in vitro. Among them, dabrafenib showed the most potent inhibition on RIP3, which was achieved by its ATP-competitive binding to the enzyme. Dabrafenib displayed highly selective inhibition on RIP3 over RIP1, RIP2 and RIP5. Moreover, only dabrafenib rescued cells from RIP3-mediated necroptosis induced by the necroptosis-induced combinations, that is, tumor necrosis factor (TNF)α, TNF-related apoptosis-inducing ligand or Fas ligand plus Smac mimetic and the caspase inhibitor z-VAD. Dabrafenib decreased the RIP3-mediated Ser358 phosphorylation of mixed lineage kinase domain-like protein (MLKL) and disrupted the interaction between RIP3 and MLKL. Notably, RIP3 inhibition of dabrafenib appeared to be independent of its B-Raf inhibition. Dabrafenib was further revealed to prevent acetaminophen-induced necrosis in normal human hepatocytes, which is considered to be mediated by RIP3. In acetaminophen-overdosed mouse models, dabrafenib was found to apparently ease the acetaminophen-caused liver damage. The results indicate that the anticancer B-Raf(V600E) inhibitor dabrafenib is a RIP3 inhibitor, which could serve as a sharp tool for probing the RIP3 biology and as a potential preventive or therapeutic agent for RIP3-involved necroptosis-related diseases such as acetaminophen-induced liver damage.
受体相互作用蛋白（RIP）3是坏死性凋亡的关键调节因子，已被证明与多种疾病相关，这表明其抑制剂在临床上具有应用前景。然而，目前报道的RIP3抑制剂很少。B-Raf（V600E）抑制剂是转移性黑色素瘤治疗的一类重要抗癌药物。在本研究中，我们发现6种B-Raf抑制剂可在体外抑制RIP3的酶活性。其中，达拉非尼对RIP3的抑制作用最强，这是通过其与该酶的ATP竞争性结合实现的。达拉非尼对RIP3的抑制作用相对于RIP1、RIP2和RIP5具有高度选择性。此外，只有达拉非尼能使细胞从由坏死性凋亡诱导组合（即肿瘤坏死因子（TNF）α、TNF相关凋亡诱导配体或Fas配体加Smac模拟物和半胱天冬酶抑制剂z-VAD）诱导的RIP3介导的坏死性凋亡中获救。达拉非尼降低了RIP3介导的混合谱系激酶结构域样蛋白（MLKL）的Ser358磷酸化，并破坏了RIP3与MLKL之间的相互作用。值得注意的是，达拉非尼对RIP3 的抑制作用似乎与其对B-Raf的抑制作用无关。进一步研究发现，达拉非尼可预防对乙酰氨基酚诱导的正常人肝细胞坏死，这种坏死被认为是由RIP3介导的。在对乙酰氨基酚过量的小鼠模型中，发现达拉非尼可明显减轻对乙酰氨基酚引起的肝损伤。结果表明，抗癌药物B-Raf（V600E）抑制剂达拉非尼是一种RIP3抑制剂，可作为研究RIP3生物学的有力工具，以及作为RIP3相关坏死性凋亡疾病（如对乙酰氨基酚诱导的肝损伤）的潜在预防或治疗药物。
