Global AAV gene therapy of Tay-Sachs disease in sheep.

项目总结 溶酶体储存疾病(LSD)由40种不同的疾病组成，累积患病率为1/4。 7,700名活产婴儿，与囊性纤维化和血友病的发生率相似。每个LSD都有一个潜在的缺陷 溶酶体的功能，导致在所有组织中广泛储存未降解的底物。许多溶酶体 贮藏性疾病有一定程度的神经系统受累，有几个以前认为的周围组织 只有现在被归类为神经病理性的亚型。尽管已经开发出有效的治疗方法 一些溶酶体疾病的外周表现，那些具有神经成分的疾病已经虚拟 无法治愈。泰-萨克氏病(TSD)和桑德霍夫病(SD)都是由细胞内的一个亚单位缺乏引起的 酶和己糖胺酶(Hex)，临床上几乎无法区分。脑内腺样体注射 相关病毒(AAV)载体使人的寿命增加了4倍，并极大地提高了患者的生活质量 患有桑德霍夫病的老鼠和猫。新发现的泰萨克斯羊是唯一相关的动物 Tay-Sachs病(TSD)模型，并可能介绍与TSD相关的挑战，这些挑战在 桑德霍夫猫捉老鼠。此外，在一个真实的TSD模型中测试治疗与大脑和身体的大小 将提供有关媒介安全、生物分布和 功效。患有Tay-Sachs病的绵羊表现出不同的临床症状，这些症状具有高度的代表性 人类青少年患者的发病年龄和临床体征的变化。这项提议的第一个目的是 鉴定、鉴定和验证绵羊Tay-Sachs的生物标志物，并了解其与 疾病的发展。该项目将利用超高场(7特斯拉)磁共振成像， 结合功能磁共振-脑电、磁共振波谱、扩散张量成像以及 代谢组学和电诊断测试；这些技术将用于评估治疗成功 目标2.初步研究表明，同时传递两个十六进制亚单位对正常 Hex同工酶的比例，但酶水平低于正常。因此，我们计划测试一种新型的双顺反管 一种在单个构建中同时表达两个十六进制亚单位的载体，使用一种新型的酶促表达 AAV衣壳，比行业标准AAV9(目标2)更高效地传导大脑。至 绕过脑实质注射的侵袭性，我们将通过静脉注射这种新的AAV 并与丘脑-脑脊液注射的“黄金标准”途径进行对比。结论 该项目最终将为未来GM2神经节苷脂沉积症和其他AAV的人类临床试验提供信息 LSD的试验。

PROJECT SUMMARY Lysosomal storage diseases (LSDs) consist of >40 distinct disorders, with a cumulative prevalence of 1 in 7,700 live births, similar in frequency to cystic fibrosis and hemophilia. Each LSD has an underlying defect in lysosomal function that leads to widespread storage of undegraded substrates in all tissues. Many lysosomal storage diseases have some level of neurologic involvement, with several previously thought peripheral tissue only subtypes now categorized as neuropathic. Though effective treatments have been developed for peripheral manifestations of some lysosomal diseases, those with neurological components have been virtually untreatable. Tay-Sachs (TSD) and Sandhoff disease (SD) are each caused by a subunit deficiency in the enzyme Hexosaminidase (Hex), and are almost clinically indistinguishable. Intracranial injection of adeno associated viral (AAV) vectors has led to > 4-fold increases in life span and vastly improved quality of life in mice and cats with Sandhoff disease. The newly discovered Tay-Sachs sheep is the only relevant animal model of Tay-Sachs disease (TSD) and may inform on challenges associated with TSD not experienced in the Sandhoff cat and mouse. Additionally, testing therapy in an authentic model of TSD with brain and body size on the same order of magnitude as children, will provide invaluable data on vector safety, biodistribution and efficacy. Sheep with Tay-Sachs experience heterogeneous clinical signs which are representative of the highly variable age of onset and clinical signs experienced by human juvenile patients. The first aim of this proposal is to identify, characterize and validate biomarkers of sheep Tay-Sachs and understand their relationship to disease progression. This project will make use of ultra-high field (7 Tesla) magnetic resonance imaging, combined functional MRI–electroencephalography, MR spectroscopy, diffusion tensor imaging as well as metabolomics and electrodiagnostic testing; and these techniques will be used to evaluate therapeutic success in Aim 2. Preliminary studies have shown that simultaneous delivery of both Hex subunits is ideal for normal ratios of Hex isozymes, but enzymatic levels were below normal. Therefore, we plan to test a novel bicistronic vector that expresses both Hex subunits in a single construct with superior enzymatic expression using a novel AAV capsid that transduces the brain with greater efficiency than the industry standard AAV9 (Aim 2). To bypass the invasiveness of parenchymal brain injections, we will administer this new AAV by intravenous injection and compare side by side with the “the gold standard” routes of Thalamic+CSF delivery. Conclusions from this project will ultimately inform future human clinical trials for GM2 gangliosidosis as well as other AAV trials for LSDs.