Neuroprotection and ERK activation by HSV-2 gene ICP10PK

描述(申请人提供)：神经元死亡(细胞凋亡)发生在急性和慢性神经退行性疾病中，如中风、阿尔茨海默氏症、帕金森病和亨廷顿病，主要由激活的半胱氨酸蛋白酶(Caspase)介导。神经元死亡的级联逐渐出现，为治疗干预提供了一段时间。然而，神经元损伤的广泛性对治疗策略的发展提出了相当大的挑战。设计了几种策略来阻断凋亡级联，但它们受到与毒性或未能保留神经元功能有关的问题的限制，这可能是由于靶向在凋亡级联中功能较晚的效应器。我们的初步数据表明，HSV-2基因(ICP10PK)可以防止体外培养的中枢神经系统神经元在各种刺激下发生凋亡。我们构建了一个生长受损的HSV-2突变体(ICP10 RR)，它保留了ICP10PK和抗凋亡活性，在纹状体注射后无毒，并在鼻腔给药时扩散到中枢神经系统(包括海马区)的连接部位。病毒感染细胞的神经保护潜力是由于激活了Raf/MEK/ERK生存通路。我们建议评估ICP10 PK在体内急性兴奋性毒性损伤中的治疗潜力，并确定其抗凋亡活性的机制。这些研究的具体目的是：(I)检测以去除营养生长支持或氧化应激为代表的范例中ICP10 PK的抗凋亡活性的机制；(Ii)设计针对上游(ICP10 PK)和下游(XIAP或p35)凋亡效应的载体，并在奎尼酸处理的器型培养中检测它们(相对于ICP10 RR)的抗凋亡活性(相对于ICP10 RR)；(Iii)确定ICP10 RR和XlAP/p35突变体在体内防止兴奋性死亡的能力；以及(Iv)确定ICP10PK在海马区的表达是否维持突触传递和功能可塑性。这些研究将为开发基于ICP10PK的治疗与细胞凋亡相关的急慢性神经退行性疾病提供所需的重要信息。

DESCRIPTION (provided by applicant): Death of neurons (apoptosis) occurs in both acute and chronic neurodegenerative diseases such as stroke, Alzheimer's, Parkinson's, and Huntington's diseases and is primarily mediated by activated cysteine proteases (caspases). Cascades of neuronal death emerge gradually, providing a period available for therapeutic intervention. However, the widespread nature of the neuronal injury presents a considerable challenge to the development of therapeutic strategies. Several strategies were designed to interrupt the apoptotic cascade but they are limited by problems related to toxicity or the failure to retain neuronal function, likely due to targeting of effectors that function late in the apoptotic cascade. Our preliminary data indicate that a HSV-2 gene (ICP10PK) prevents apoptosis of CNS neurons in vitro produced by various stimuli. We constructed a growth-compromised HSV-2 mutant (ICP10 RR) that retains ICP10PK and anti-apoptotic activity, is not toxic following intrastriatal injection and disseminates to connected sites in the CNS (including hippocampus) upon intranasal delivery. Neuroprotective potential in virus-infected cells is due to activation of the Raf/MEK/ERK survival pathway. We propose to evaluate the therapeutic potential of ICP10 PK in acute excitotoxic injury in vivo and define the mechanism of anti-apoptotic activity. The Specific Aims are: (i) To examine the mechanism of ICP10 PK anti-apoptotic activity in paradigms represented by removal of trophic growth support or oxidative stress, (ii) To engineer vectors that target both upstream (ICP10 PK) and downstream (XIAP or p35) apoptotic effectors and examine their anti-apoptotic activity (relative to ICP10 RR) in organotypic cultures treated with kianic acid (excitotoxic model), (iii) To determine the ability of ICP10 RR and the XlAP/p35 mutants to prevent excitotoxic death in vivo, and (iv) To determine whether ICP10PK expression in the hippocampus maintains synaptic transmission and functional plasticity. The studies will provide significant information required for the development of ICP10PK based therapies for the treatment of acute and chronic neurodegenerative diseases that are associated with apoptosis.