Background: Stress-associated kinases are considered major pathological mediators in several incurable neurological disorders. Importantly, among these stress kinases, the c-Jun NH2-terminal kinase (JNK) has been linked to numerous neuropathological conditions, including oxidative stress, neuroinflammation, and brain degeneration associated with brain injuries such as ischemia/reperfusion injury. In this study, we adopted a drug repurposing/reprofiling approach to explore novel JNK3 inhibitors from FDA-approved medications to supplement existing therapeutic strategies.
Materials and Methods: We performed in silico docking analysis and molecular dynamics simulation to screen potential candidates from the FDA approved drug library using the standard JNK inhibitor SP600125 as a reference. After the virtual screening, dabigatran, estazolam, leucovorin, and pitavastatin were further examined in ischemic stroke using an animal rodent model of focal cerebral ischemia using transient middle cerebral artery occlusion (t-MCAO). The selected drugs were probed for neuroprotective effectiveness by measuring the infarct area (%) and neurological deficits using a 28-point composite score. Biochemical assays including ELISA and immunohistochemical experiments were performed.
Results: We obtained structural insights for dabigatran, estazolam, and pitavastatin binding to JNK3, revealing a significant contribution of the hydrophobic regions and significant residues of active site regions. To validate the docking results, the pharmacological effects of dabigatran, estazolam, leucovorin, and pitavastatin on MCAO were tested in parallel with the JNK inhibitor SP600125. After MCAO surgery, severe neurological deficits were detected in the MCAO group compared with the sham controls, which were significantly reversed by dabigatran, estazolam, and pitavastatin treatment. Aberrant morphological features and brain damage were observed in the ipsilateral cortex and striatum of the MCAO groups. The drugs restored the anti-oxidant enzyme activity and reduced the levels of oxidative stress-induced p-JNK and neuroinflammatory mediators such as NF-kB and TNF-ɑ in rats subjected to MCAO.
Conclusion: Our results demonstrated that the novel FDA-approved medications attenuate ischemic stroke-induced neuronal degeneration, possibly by inhibiting JNK3. Being FDA-approved safe medications, the use of these drugs can be clinically translated for ischemic stroke-associated brain degeneration and other neurodegenerative diseases associated with oxidative stress and neuroinflammation.



中文翻译：

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将 FDA 批准的药物重新用作 JNK3 抑制剂，以预防 MCAO 诱导的大鼠神经炎症

背景：应激相关激酶被认为是几种无法治愈的神经系统疾病的主要病理介质。重要的是，在这些应激激酶中，c-Jun NH2 末端激酶 (JNK) 与许多神经病理学状况有关，包括氧化应激、神经炎症和与脑损伤（如缺血/再灌注损伤）相关的脑退化。在这项研究中，我们采用药物再利用/再分析方法从 FDA 批准的药物中探索新型 JNK3 抑制剂，以补充现有的治疗策略。
材料和方法：我们使用标准 JNK 抑制剂 SP600125 作为参考，进行了计算机对接分析和分子动力学模拟，以从 FDA 批准的药物库中筛选潜在候选药物。虚拟筛选后，使用暂时性大脑中动脉闭塞 (t-MCAO) 的局灶性脑缺血动物啮齿动物模型，进一步检查达比加群、艾司唑仑、亚叶酸钙和匹伐他汀在缺血性中风中的作用。通过使用 28 分综合评分测量梗塞面积 (%) 和神经功能缺损，探讨所选药物的神经保护效果。进行了生化测定，包括 ELISA 和免疫组织化学实验。
结果：我们获得了达比加群、艾司唑仑和匹伐他汀与 JNK3 结合的结构见解，揭示了疏水区域和活性位点区域的重要残基的重要贡献。为了验证对接结果，与 JNK 抑制剂 SP600125 平行测试了达比加群、艾司唑仑、亚叶酸钙和匹伐他汀对 MCAO 的药理作用。MCAO 手术后，与假手术对照组相比，MCAO 组检测到严重的神经功能缺损，达比加群、艾司唑仑和匹伐他汀治疗显着逆转。在 MCAO 组的同侧皮质和纹状体中观察到异常的形态特征和脑损伤。
结论：我们的结果表明，FDA 批准的新型药物可能通过抑制 JNK3 来减轻缺血性中风引起的神经元变性。作为 FDA 批准的安全药物，这些药物的使用可以临床转化为缺血性中风相关的脑退化和其他与氧化应激和神经炎症相关的神经退行性疾病。