Pathogenesis and possible therapies modelled in ovine Batten disease

描述(申请人提供)：神经性蜡样脂褐素增多症(NCLS，巴顿病)是一种毁灭性的遗传性儿童致死性神经退行性疾病。目前还没有有效的治疗方法，姑息治疗也很困难。对新西兰绵羊的CLN6形式的研究已被证明对理解生物化学和临床前病理有非常重要的价值。这些绵羊具有与人类相似的大脑解剖、生理和遗传组织，并表现出类似的以皮质萎缩为主导的神经病理，以及类似的临床疾病发展。这个项目的总体目标是完全理解到可以开始病毒载体基因疗法的体内试验的程度。分子生物学、生物化学和免疫组织学实验将确定矫正所需的基因产物及其亚细胞居留位置。神经元培养将用Lenti和腺相关病毒衍生载体进行测试，以确定是否适合将该基因转导到绵羊神经元和神经胶质细胞。产前和临床前的神经病理学研究表明，神经胶质细胞的激活在本病的发病机制中起主要作用。进一步的研究将确定胶质细胞激活和炎症的级联反应，以确定关键步骤，从而确定最有效的干预点。抗炎药物慢性治疗对绵羊疾病发展的有效性将在体内进行测试。病理学研究还表明，绵羊脑室下区的神经发生和细胞迁移到患区的时间延长。这一程度将通过体内新细胞的BrdU标记和抗体检测来确定。最后，通过将携带报告基因的首选载体定向注射到受影响的绵羊大脑中，并检测基因表达的扩散，将确定这种将病毒载体中的基因转移到所需区域的有用性。这些研究的目的是为在绵羊身上直接测试基因疗法以及抑制炎症提供所有要求。靶细胞将与与它们相关的CLN6基因产物一起被鉴定。治疗的时间窗口将与抑制炎症在减缓疾病进程中的作用一起确定。最有效的病毒载体将被确定，注射到SVZ的技术将被建立。Revelance：这个项目旨在获得知识，以确定一种大型动物形式的巴顿病的现实治疗方法，该方法可以测试为可能的人类使用。获得的大部分知识将与其他疾病和其他治疗策略相关，如阿尔茨海默病和干细胞移植。

DESCRIPTION (provided by applicant): The neuronal ceroid lipofuscinoses (NCLs, Batten disease) are devastating inherited fatal neurodegenerative diseases of children. At present there are no effective therapies and palliative care is difficult. Studies on a CLN6 form in New Zealand sheep have proved invaluable in understanding the biochemistry and preclinical pathology. These sheep have a human-like brain anatomy, physiology and genetic organization, and display similar neuropathology dominated by cortical atrophy, and a similar development of clinical disease. The overall aim of this project is to complete understanding to a point where in vivo trials of viral vector gene therapies can begin. Molecular biology, biochemistry and immunohistological experiments will define the gene product required for correction and its subcellular site of residence. Neuron cultures will be be tested with lenti- and adeno-associated virally derived vectors to determine suitability for transduction of the gene into sheep neurons and glial cells. Prenatal and preclinical neuropathology studies indicated that glial activation has a primary role in the pathogenesis of this disease. Further studies will define the cascade of glial cell activation and inflammation, to determine the critical steps and thus determine the most effective point for intervention. The effectiveness of chronic treatment with anti-inflammatory drugs on the development of the disease in sheep will be tested in vivo. Prolonged neurogenesis and migration of cells from the subventricular zone to the affected areas in the brains of affected sheep was also indicated in the pathology studies. The extent of this will be established by BrdU labeling of new cells in vivo and antibody detection. Finally the usefulness of this migration for carrying a gene in viral vectors to the areas where it is required will be determined by targeted injection of the preferred vector carrying a reporter gene into affected sheep brains and detection of the dispersal of gene expression. These studies are aimed at providing all the requirements for directly testing gene therapy on the sheep, in combination with suppression of inflammation. The target cells will have been identified along with the CLN6 gene product relevant to them. The time window for therapy will have been defined along with the role of suppression of inflammation in slowing the course of the disease. The most effective viral vector will have been identified and the techniques for injection into the SVZ established. REVELANCE: This project intends to gain knowledge to define realistic therapies in a large animal form of Batten disease that can be tested for possible human use. Much of the knowledge gained will be relevant to other diseases and other treatment strategies, such as Alzheimer's disease and stem cell transplantation.