The Role of Cdk5 in Stroke

描述（由申请人提供）：中风是美国成人残疾的主要原因，也是世界范围内死亡的第三大原因。最常见的形式是缺血性斯托克，是由血液凝块阻塞动脉供应引起的，导致永久性神经损伤或死亡。对于少数患者，可以去除或溶解血凝块以预防或限制中风损伤。然而，改善生存和恢复的神经保护剂是不可用的。动脉闭塞后，神经元死亡从最初的梗死向外扩散， 神经元去极化引起大量谷氨酸释放和NMDA受体（NMDAR）超活化。结果，兴奋性毒性Ca 2+流入细胞，在那里它激活蛋白酶钙蛋白酶。这种兴奋性毒性级联反应是中风病理生理学的主要组成部分。然而，用NMDAR拮抗剂阻断级联反应的努力由于不希望的作用而在临床试验中未被证明是有效的。我们发现蛋白激酶Cdk 5将钙蛋白酶与NMDAR的NR 2B亚基物理连接。cdk 5也与其激活剂p35结合。通过这种NR 2B-钙蛋白酶-Cdk 5/p35信号传导复合物，NMDAR的兴奋毒性激活导致钙蛋白酶将p35转化为p25。与p25相关的cdk 5导致神经元死亡，并被认为介导缺血性损伤。然而，关于Cdk 5或p35/p25如何促进卒中病理生理学或如何有效靶向其以改善结局，目前还知之甚少。在这里，我们提出了描述大脑中动脉闭塞（MCAO）后Cdk 5的失调，并确定在小鼠中Cdk 5或p35丢失的神经保护作用。作为实现神经保护的新策略，我们将通过鉴定功能性蛋白质-蛋白质相互作用基序和开发小干扰肽（SIP）来靶向NR 2B-calpain-Cdk 5/p35级联， 阻止钙蛋白酶-Cdk 5和NR 2B-Cdk 5相互作用。这些SIP将被优化为有效且特异性地解离NR 2B-钙蛋白酶-Cdk 5/p35并防止通过NMDAR超活化产生Cdk 5/p25而不影响受体、钙蛋白酶或Cdk 5的功能的分子。然后，我们将评估他们的能力，神经保护和改善行为和组织学结果的小鼠受到MCAO。这些转化研究将促进我们对介导中风损伤的机制的了解，并获得新的神经保护剂，这将可能导致开发改善恢复和减少中风相关残疾的疗法。

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.