Gene delivery methods for global therapy in the CNS

描述（由申请方提供）：溶酶体贮积病（LSD）是最常见的儿童遗传性疾病类型，估计在一般人群中的发生率为1/7700。据估计，60%的LSD有一定程度的神经系统受累，目前尚无治疗方法。因此，研究治疗神经病性LSD的新方法至关重要。这项工作将集中在GM 1-神经节苷脂沉积症，这是一种无法治愈的儿童疾病，伴有中度至重度神经功能障碍，由溶酶体酸性β-半乳糖苷酶（b-gal）缺乏引起，伴有GM 1-神经节苷脂在中枢神经系统中的蓄积。我们将在患有GM 1-神经节苷脂沉积症的年轻成年小鼠中进行这些研究，因为大多数LSD病例在儿童早期或之后被诊断，此时大脑发育相当于年轻成年或成年小鼠。我们实验室和其他实验室的实验表明，AAV 8载体对成年小鼠大脑的基因转移非常有效。此外，我们的研究已经表明，AAV 8载体可以通过尾静脉血管内输注后覆盖脑微毛细血管内皮。因此，将在这些实验中使用AAV 8载体。本申请的目的是检验以下两个假设：1）在成年GM 1-神经节苷脂沉积症小鼠的丘脑中小鼠b-gal的从头表达足以实现整个CNS的病理性生物化学异常的完全校正和神经运动功能测试中的正常表现; 2）b-gal在GM 1-神经节苷脂沉积症小鼠脑微毛细血管内皮中的表达将导致GM 1-和GA 1-的野生型水平。神经节苷脂在整个中枢神经系统和正常的表现，在测试中的神经运动功能。在此，我们将编码b-gal或GFP的AAV 8载体双侧注射到2个月大的GM 1-神经节苷脂沉积症小鼠的丘脑中（Aim 1），或在内皮或肝脏特异性启动子的控制下通过尾静脉递送编码b-gal或GFP的AAV 8载体（Aim 2）。将在1年内的不同时间点评价给药小鼠和对照小鼠的神经病理学、生物化学、免疫学和行为参数。我们预计这些实验的结果将适用于其他LSD模型，并且在不久的将来，在进入临床试验之前，可以在GM 1-神经节苷脂沉积症的大型动物模型中对其进行评估。

DESCRIPTION (provided by applicant): Lysosomal storage diseases (LSDs) comprise the most common type of childhood genetic disorder, with an estimated frequency of 1 in 7700 in the general population. It is estimated that 60% of all LSDs have some degree of neurological involvement for which no treatment is available. Thus it is of paramount importance to investigate new approaches to treat neuronopathic LSDs. This work will focus on GM1-gangliosidosis, which is an untreatable childhood disease with moderate to severe neurological impairment caused by a deficiency of lysosomal acid beta-galactosidase (b-gal) with accumulation of GM1-ganglioside in the central nervous system. We will conduct these studies in young adult mice with GM1-gangliosidosis because most LSD cases are diagnosed in early childhood or later when the brain is developmentally equivalent to young adult or adult mice. Experiments in our laboratory and others have shown that AAV8 vectors are exceptionally efficient for gene transfer to the adult mouse brain. Moreover, our studies have shown that AAV8 vectors can transduce the brain microcapillary endothelium after intravascular infusion via the tail vein. Thus AAV8 vectors will be used throughout these experiments. The aims in this application were designed to test the following two hypotheses: 1) De novo expression of mouse b-gal in the thalamus of adult GM1- gangliosidosis mice is sufficient to achieve complete correction of patho-biochemical abnormalities throughout the CNS and normal performance in tests of neuro-motor function; 2) Expression of b-gal in the brain microcapillary endothelium of GM1-gangliosidosis mice will result in wild type levels of GM1- and GA1- ganglioside throughout the CNS and normal performance in tests of neuro-motor function. Here we will inject AAV8 vectors encoding for b-gal or GFP bilaterally into the thalamus of 2 month old GM1- gangliosidosis mice (Aim 1) or deliver AAV8 vectors encoding b-gal or GFP under control of endothelial- or liver-specific promoters via the tail vein (Aim 2). Treated and control mice will be evaluated for neuropathological, biochemical, immunological, and behavioral parameters at different time points over 1 year. We anticipate the findings from these experiments to be applicable to other models of LSDs, and that in the near future they can be evaluated in a large animal model of GM1-gangliosidosis before moving to clinical trials.