The Role of Cdk5 in Stroke

描述(申请人提供)：中风是美国成人残疾的主要原因，也是世界范围内第三大死亡原因。最常见的形式是缺血性中风，是由于血液凝块阻塞动脉供应，导致永久性神经损伤或死亡。对于少数患者，可以通过清除或溶解血栓来预防或限制中风损伤。然而，目前还没有能提高存活率和恢复能力的神经保护剂。动脉闭塞后，神经元死亡从最初的脑梗塞向外扩散， 神经元去极化导致大量谷氨酸释放和NMDA受体(NMDAR)过度激活。结果，兴奋性毒性的钙离子流入细胞，在那里它激活了蛋白水解酶钙蛋白酶。这种兴奋性毒性级联反应是卒中病理生理学的主要组成部分。然而，由于不想要的影响，用NMDAR拮抗剂阻止级联反应的努力在临床试验中并未被证明有效。我们发现，蛋白激酶CDK5在物理上将Calain与NMDARs的NR2B亚单位联系在一起。CDK5也与其激活子p35结合。通过这个NR2B-calain-CDK5/p35信号复合体，NMDARs的兴奋毒性激活导致Calain将p35转化为p25。与p25相关的CDK5可导致神经元死亡，并被认为参与了缺血性损伤。然而，关于CDK5或p35/p25如何在卒中病理生理学中起作用，或者它如何被有效地靶向以改善预后，目前还知之甚少。在这里，我们建议描述大脑中动脉闭塞(MCAO)后CDK5的异常调节，并确定失去CDK5或p35的神经保护作用。作为一种实现神经保护的新策略，我们将通过识别功能性蛋白质-蛋白质相互作用基序和开发小干扰肽(Sip)来靶向nr2b-calain-cdk5/p35级联。 阻止Calain-CDK5和NR2B-CDK5的相互作用。这些sip将被优化为有效和特定地解离NR2B-calain-cdk5/p35的分子，并阻止NMDAR过度激活产生cdk5/p25，而不会以其他方式影响受体calain或cdk5的功能。然后，我们将评估它们对大脑中动脉阻塞小鼠的神经保护和改善行为和组织学结果的能力。这些转化性研究将促进我们对卒中损伤机制的了解，并衍生出新的神经保护剂，可能导致促进康复和减少与中风相关的残疾的治疗方法的开发。

DESCRIPTION (provided by applicant): Stroke is the leading cause of adult disability in the U.S. and the third leading cause of mortality world-wide. The most common form, ischemic stoke, is caused by blood clots occluding arterial supply, resulting in permanent neurological damage or death. For a minority of patients, blood clots can be removed or lysed to prevent or limit stroke injury. However, neuroprotectants that improve survival and recovery are not available. Following arterial occlusion, neuronal death spreads outward from the initial infarct as neurons depolarize causing massive glutamate release and hyperactivation of NMDA receptors (NMDAR). As a result, excitotoxic Ca2+ flows into cells where it activates the protease calpain. This excitotoxic cascade is a major component of stroke pathophysiology. However, efforts to block the cascade with NMDAR antagonists have not proven effective in clinical trials due to unwanted effects. We discovered that the protein kinase Cdk5 physically links calpain to the NR2B subunit of NMDARs. Cdk5 is also bound to its activator p35. Through this NR2B-calpain- Cdk5/p35 signaling complex, excitotoxic activation of NMDARs causes calpain to convert p35 to p25. Cdk5 associated with p25 causes neuronal death and has been suggested to mediate to ischemic injury. However little is yet known of how Cdk5 or p35/p25 contributes to stroke pathophysiology or how it may be effectively targeted to improve outcome. Here we propose to characterize the dysregulation of Cdk5 following middle cerebral artery occlusion (MCAO) and determine the neuroprotective effects of loss of Cdk5 or p35 in mice. As a novel strategy to achieve neuroprotection we will target the NR2B-calpain-Cdk5/p35 cascade by identifying functional protein-protein interaction motifs and developing small interfering peptides (SIPs) that prevent calpain-Cdk5 and NR2B-Cdk5 interactions. These SIPs will be optimized as molecules that potently and specifically dissociate the NR2B-calpain-Cdk5/p35 and prevent Cdk5/p25 generation by NMDAR hyperactivation without otherwise affecting the functions of the receptors, calpain, or Cdk5. We will then assess their ability to neuroprotect and improve behavioral and histological outcome in mice subjected to MCAO. These translational studies will advance our knowledge of the mechanisms that mediate stroke injury and derive novel neuroprotectants that will potentially lead to the development of therapies that improving recovery and reduce stroke-related disability.