GD3 synthase gene therapy to improve memory and prevent neurodegeneration

描述(申请人提供)：阿尔茨海默氏病的特征是β-淀粉样蛋白(Abeta)在大脑中聚集，广泛的神经变性和认知能力下降。我们的工作集中在通过减少或消除GD3合酶(GD3S)来改善阿尔茨海默病的病理，GD3S是一种负责合成四种主要脑神经节苷脂中的两种的酶。我们的初步数据表明，敲除编码GD3S(St8sia1)的基因显著减少A2聚集和氧化应激，并防止携带突变的人类淀粉样前体蛋白(App)和早老素1(PSEN1)转基因小鼠的记忆缺陷。然而，这些小鼠从出生起就缺乏GD3S，而且它们缺乏许多与正常发育有关的神经节苷脂。我们的目标是了解神经节苷脂在阿尔茨海默病中的作用，以及它们与记忆障碍的关系，记忆障碍是该疾病最早的认知症状。这项应用的目的是确定在5xFAD小鼠模型中，使用基因沉默(ShRNA)敲除GD3S是否与敲除GD3S一样有效地缓解阿尔茨海默病的特征。这项研究的一般假设是，GD3S的shRNA敲除将成功地减少斑块的形成，并防止成年突变小鼠的神经退化和记忆障碍。我们已经制作了三种病毒载体介导的GD3S-shRNA结构，可以将GD3S的表达减少约80%，并在大脑中持续存在。在拟议的实验中，shRNA构建物将被注射给5xFAD和对照组小鼠。将评估空间记忆，以及焦虑和感觉运动功能的控制测试。尸检分析将评估与阿尔茨海默氏症相关的神经病理和细胞死亡。在转基因小鼠中成功减少淀粉样蛋白负担、细胞死亡和记忆损伤，可能为阿尔茨海默病的新治疗策略提供洞察--治疗可以减少或预防阿尔茨海默病患者的痴呆症。 公共卫生相关性：拟议的实验将测试一种新型基因疗法rAAV.GD3S.shRNA对记忆和阿尔茨海默病相关神经病理的影响。该载体的使用不仅代表了一种新的治疗策略，也代表了阿尔茨海默病的新治疗靶点。这些实验与国家老龄研究所和国家神经疾病和中风研究所宣布的目标一致。

DESCRIPTION (provided by applicant): Alzheimer's disease is characterized by the aggregation of beta-amyloid (Abeta) protein in the brain, widespread neurodegeneration, and cognitive decline. Our work focuses on amelioration of Alzheimer's pathology by reducing or eliminating GD3 synthase (GD3S), an enzyme responsible for synthesis of two of the four major brain gangliosides. Our preliminary data demonstrate that knocking out the gene that codes for GD3S (St8sia1) drastically reduces A2 aggregation and oxidative stress, and prevents memory deficits in mice carrying mutant human transgenes for amyloid precursor protein (App) and presenilin 1 (Psen1). However, GD3S is absent from birth in these mice, and they lack many of the gangliosides involved in normal development. Our goal is to understand the role of gangliosides in Alzheimer's disease and how they relate to memory impairment, the earliest cognitive symptom of the disease. The objective of this application is to determine whether knocking down GD3S using gene silencing (shRNA) is as effective as knocking out GD3S in alleviating features of Alzheimer's disease in the 5xFAD mouse model. The general hypothesis of the proposed research is that shRNA knock-down of GD3S will successfully reduce plaque formation and prevent neurodegeneration and memory impairments in adult mutant mice. We have already made three viral- vector-mediated GD3S-shRNA constructs that reduce GD3S expression ~80 percent and persist in the brain. In the proposed experiments, shRNA constructs will be administered to 5xFAD and control mice. Spatial memory will be assessed, as well as control tests for anxiety and sensorimotor function. Post-mortem analyses will assess Alzheimer- related neuropathology and cell death. Successfully reducing amyloid burden, cell death, and memory impairment in the transgenic mice may provide insight into new treatment strategies for Alzheimer's disease--treatments that could reduce or prevent dementia in Alzheimer patients. PUBLIC HEALTH RELEVANCE: The proposed experiments will test the effects of a novel genetic therapy, rAAV.GD3S.shRNA, on memory and Alzheimer-related neuropathology. The use of this vector represents not only a novel treatment strategy, but a novel therapeutic target for Alzheimer's disease. These experiments are consistent with the stated goals of National Institute on Aging and National Institute of Neurological Disorders and Stroke.