Targeting CaM Kinase II for Neuroprotection in Ischemic Stroke

描述（由申请人提供）：中风是一个重大的公共卫生问题，在美国的死亡率排名第三，仅次于心血管系统和癌症等其他疾病。我们建议通过靶向Ca2+稳态的关键调节因子，Ca2+/ cam依赖性蛋白激酶II （CaMKII）来追求缺血性卒中的神经保护。CaMKII的抑制代表了缺血性卒中神经保护的一种新模式，基于阻断CaMKII对Ca2+过载的影响，以及其对神经毒性介质的更直接影响。两项进展为我们在缺血性卒中中测试有效的小分子CaMKII抑制剂提供了理论依据和机会。首先，CaMKII的过度活跃已被证明在谷氨酸兴奋毒性中促进细胞死亡，而激酶抑制剂在原位和大脑中动脉闭塞（MCAO）模型中具有神经保护作用。其次，我们利用这个机会重新启动了CaMKII抑制剂项目，该项目最初是在一家制药公司推进的，现在我们正在基于人类酶的第一个晶体结构的见解加速该项目。我们的建议相对简单，增加我们开发的有效抑制剂的中枢神经系统渗透，并在永久性闭塞MCAO模型中测试优化的先导化合物。我们设计了一些修饰，以增加化合物的亲脂性，目的是增加它们的中枢神经系统渗透。这些化合物将在体外分析激酶抑制和选择性，以及在神经元培养中对谷氨酸兴奋毒性的抑制和保护。我们将根据MDR----MDCK单分子层的双向通透性来选择具有最大中枢神经系统穿透潜力的抑制剂，并量化其在体内的脑/血浆分布。我们将在大鼠永久性闭塞MCAO模型中测试导联CNS穿透抑制剂。我们将使用激酶活性的生物标志物来监测体内CaMKII抑制的程度，并量化抑制剂对梗死面积的影响。在缺血性卒中中----概念的成功证明----将使我们能够进行II期SBIR提案，以优化药物----样特性并对先导化合物进行IND研究。

DESCRIPTION (provided by applicant): Stroke is a major public health problem, with a US mortality rate that ranks third behind other diseases involving the cardiovascular system and cancer. We propose to pursue neuroprotection in ischemic stroke by targeting a key regulator of Ca2+ homeostasis, Ca2+/CaM-dependent protein kinase II (CaMKII). Inhibition of CaMKII represents a novel paradigm in ischemic stroke-neuroprotection based on blocking the effects of CaMKII on Ca2+ overload as well as its more direct effect on mediators of neurotoxicity. Two developments provide us with the rationale and opportunity to test potent small molecule CaMKII inhibitors in ischemic stroke. First, hyperactivity of CaMKII has been shown to promote cell death in glutamate excitotoxicity while inhibitors of the kinase are neuroprotective in situ and in the middle cerebral artery occlusion (MCAO) model. Second we have taken the opportunity to restart a CaMKII inhibitor program initially advanced at a pharm and which we are now accelerating based on insights from the first crystal structures of the human enzyme. Our proposal is relatively straightforward, to increase CNS penetration of the potent inhibitors we developed and test the optimized lead compound in a permanent occlusion MCAO model. We have designed a number of modifications that increase lipophilicity of our compounds with the aim of increasing their CNS penetration. The compounds will be analyzed for kinase inhibition and selectivity in vitro, and for inhibition and protection from glutamate excitotoxicity in neuronal cultures. The inhibitor with the greatest potential for CNS penetration will be selected based on bidirectional permeability through MDR----MDCK monolayers and its brain/plasma distribution in vivo will be quantified. The lead CNS penetrating inhibitor will be tested in the rat permanent occlusion MCAO model. We will monitor the degree of CaMKII inhibition achieved in vivo using biomarkers of kinase activity and quantify the effect of the inhibitor on infarct size. A successfully proof----of----concept in ischemic stroke will position us for a Phase II SBIR proposal to optimize the drug----like properties and conduct IND enabling studies of the lead compound. PUBLIC HEALTH RELEVANCE: Stroke is a major public health problem with a US mortality rate that ranks third behind other diseases involving the cardiovascular system and cancer. Recent data implicate hyperactivity of Ca2+/CaM- dependent protein kinase II (CaMKII), a major mediator of Ca2+ signaling, in ischemic damage. We aim to increase the brain penetration of our potent CaMKII inhibitors and test the optimized lead compound for efficacy in an animal model of human ischemic stroke.ischemic stroke.