Gene delivery methods for global therapy in the CNS

描述(由申请人提供)：溶酶体储存疾病(LSD)是最常见的儿童遗传性疾病类型，在普通人群中估计频率为每7700人中就有一人。据估计，所有LSD的60%都有不同程度的神经损害，没有治疗方法。因此，研究治疗神经病理性腰椎间盘突出症的新方法至关重要。这项工作将集中在GM1-神经节苷脂沉积症，这是一种无法治疗的儿童疾病，由于溶酶体酸性β-半乳糖苷酶(b-GAL)缺乏和GM1-神经节苷脂在中枢神经系统积聚而导致中到严重的神经功能损害。我们将在患有神经节苷脂GM1的年轻成年小鼠中进行这些研究，因为大多数LSD病例是在儿童早期或以后大脑发育与年轻成年或成年小鼠相同的时候被诊断出来的。我们实验室和其他实验室的实验表明，AAV8载体在将基因转移到成年小鼠大脑方面非常有效。此外，我们的研究表明，AAV8载体经尾静脉血管内注射后可以转导脑微血管内皮细胞。因此，在这些实验中将使用AAV8载体。本应用程序的目的是验证以下两个假设：1)小鼠b-Gal在成年GM1-神经节苷脂中毒小鼠丘脑中的从头表达足以完全纠正整个中枢神经系统的病理生化异常，并在神经运动功能测试中正常表现；2)b-Gal在GM1-神经节苷脂中毒小鼠脑内微血管内皮细胞的表达将导致GM1-和GA1-神经节苷脂在整个中枢神经系统内野生型水平，以及在神经运动功能测试中正常表现。在这里，我们将把编码b-Gal或GFP的AAV8载体注射到2月龄GM1神经节苷脂中毒小鼠的丘脑(目标1)，或者在内皮或肝脏特异性启动子的控制下通过尾静脉输送编码b-Gal或GFP的AAV8载体(目标2)。治疗和对照小鼠将在一年多的不同时间点进行神经病理、生化、免疫学和行为参数的评估。我们预计这些实验的发现将适用于LSD的其他模型，在不久的将来，它们可以在GM1-神经节苷脂沉着症的大型动物模型中进行评估，然后进入临床试验。

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.