STEREOTAXIC GENE THERAPY TO THE RHESUS CNS

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。所列机构为中心，不一定是研究者所在机构。中枢神经系统是基因治疗的主要目标，因为影响大脑的遗传和获得性疾病的范围很广。神经退行性疾病如帕金森病、亨廷顿病、阿尔茨海默病和肌萎缩侧索硬化症（ALS）是影响老年人的最常见的疾病之一，因此是基因治疗发展的主要目标。虽然罕见，遗传性溶酶体贮积症和脑白质营养不良，如泰-萨二氏病、粘多糖样变性和克拉伯病，也是开发新型基因治疗干预措施的主要目标。这些和其他CNS疾病的治疗选择主要限于支持性治疗，因此提供的治疗选择很少。直接CNS基因转移可能为消除与这些疾病相关的病理提供最佳机会。本提案的目标是比较重组慢病毒（LV）和腺相关病毒（腺相关病毒）载体系统的转导，包括非人灵长类动物（特别是恒河猴）中枢神经系统中的基因转移水平、（细胞和结构）模式以及基因表达的持续性。该提案的最终目的是确定哪种载体或载体组合在与人类非常密切相关的动物模型中向最广泛的细胞类型提供高水平的基因转移。该提案的主要优点是，将使用一致的生产方案制备两种载体系统的制剂，并直接施用于CNS。我们已经进行了一系列基因转移和生物识别研究，使用这些重组病毒制剂的直接立体定位注射靶向多个区域的大脑，如海马，皮质，尾壳核/纹状体，丘脑和小脑。在给予载体制剂后，我们目前正在通过确定基因转移到大脑选定区域的水平并通过免疫组织化学和共聚焦显微镜描绘转导的特定细胞类型来绘制各种载体的转导模式。我们目前的数据表明，可以检测到由各种血清型的AAV（AAV 1、2、7、9和10）载体提供的基因转移模式的显著差异。VSV-G假型慢病毒载体的生物分布证明了对许多细胞类型的广泛基因转移，但效率低于AAV载体。.拟议的翻译研究的结果将作为设计和执行CNS指导的基因治疗方案在人类，特别是儿童的宝贵信息。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The central nervous system is a primary target for gene therapy because the broad spectrum of inherited and acquired disorders that affect the brain. Neurodegenerative diseases such as Parkinson¿s disease, Huntington¿s disease, Alzheimer¿s disease, and amyotrophic lateral sclerosis (ALS) are among the most common disorders that affect older humans and as such are prime targets for the development of gene therapy. Although rare, inherited lysosomal storage disorders and leukodystrophys, such as Tay-Sach¿s, mucopolysaccharidosis and Krabbe¿s disease, are also primary targets for the development of novel gene therapy interventions. Treatment options for these and other CNS disorders are primarily limited to supportive care, thus providing few therapeutic options. Direct CNS gene transfer may provide the best opportunity for eliminating the pathology associated with these conditions. The goal of this proposal is the comparison of the transduction of recombinant lentivirus (LV) and adeno-associated virus (AAV) vector systems with regard to levels of gene transfer, pattern of (cells and structures), and persistence of gene expression in the CNS of nonhuman primates, specifically rhesus macaques. The ultimate aim of this proposal is to determine which vector or combination of vectors provides high levels of gene transfer to the broadest range of cell types in an animal model that is very closely related to humans. The primary advantage of this proposal is that preparations of both vector systems will be prepared using consistent production protocols and administered directly to the CNS. We have performed a series of gene transfer and biodstribution studies using direct stereotaxic injection of these recombinant virus preparations to target multiple regions of the brain, such as the hippocampus, cortex, caudate putamen/striatum, thalamus and cerebellum. Following administration of the vector preparations, we are currently mapping the transduction patterns of the various vectors by determining the levels of gene transfer to selected regions of the brain and delineating the specific cell types transduced by immunohistochemistry and confocal microscopy. Our current data suggests that marked differences in the patterns of gene transfer provided by various serotypes of AAV (AAV 1, 2, 7, 9 and 10) vectors can be detected. The biodistribution of VSV-G pseudotyped lentivirus vectors demonstrated broad gene transfer to many cell types, but at lower efficiency than AAV vectors. . The results of the proposed translational studies will serve as invaluable information on the design and execution of CNS-directed gene therapy protocols in humans, especially children.