Generation of a Large Animal Model of Sialidosis to Enable Future Translation of Novel Therapeutics

项目摘要 唾液酸沉积症是一种罕见的、致命的神经系统疾病，由NEU 1基因突变引起，导致视力丧失， 癫痫不自主肌阵挛和共济失调我们的团队在将基因疗法 该诊所并计划开发一种基因疗法来治疗唾液酸中毒。我们以往的经验表明， 在人类遗传疾病的大型动物模型中进行测试， 并且这些模型的使用增加了功效的可能性。我们已经开发了一个创始人羊与 突变像1型唾液酸中毒患者，并会繁殖他产生一个动物群体。自上次 我们已经使用CRISPR/spCas 9编辑创建了几个严重的突变，因此在这个目标中，我们将 使用胚胎的Prime基因组编辑，重新创建两种人类突变（1型和2型），最终目标是 一种重现人类状况的突变（目标1）。我们将评估每个模型的能力， 可靠地模拟唾液酸中毒，然后选择最佳模型（目的2）。这种表型分析包括活体临床指标，如 MRI、EEG、EMG、神经和认知测试以及深入的尸检分析，以确定 再现疾病的生物化学和组织病理学方面。活体临床试验的外部评价 将由我们的临床合作者蒂夫特医生进行另外，蒂夫特博士会提供人体样本 与新绵羊模型进行比较。疾病的生物化学方面将由外部验证， d 'Azzo博士（唾液酸中毒领域的领导者）和兽医Koehler博士的病理特征 神经病理学家在完成这些研究后，这个经过充分验证的模型将用于了解更多关于 唾液酸中毒作为一种疾病，也用于开发腺相关病毒基因疗法或其他 唾液酸中毒的未来治疗策略。

Project Summary Sialidosis is a rare, fatal, neurological disorder caused by a mutation in the NEU1 gene resulting in vision loss, seizures, involuntary myoclonus, and ataxia. Our team has a strong track record in brining gene therapies to the clinic and has plans to develop a gene therapy to treat sialidosis. Our previous experience has shown that testing in large animal models of human genetic diseases better approximates what will happen and patients and use of these models increases the likelihood of efficacy. We have developed a founder sheep with a mutation like type 1 sialidosis patients and will breed him to generate a colony of animals. Since the last submission we have created several severe mutations using CRISPR/spCas9 editing, therefore in this aim we will use Prime genome editing of embryos, to recreate two human mutations (Type 1 and Type 2) with the end goal of a mutation that recapitulates the human condition (Aim 1). We will evaluate each model for its ability to reliably mimic sialidosis then select the best model (Aim 2). This phenotyping includes in-life clinical metrics like MRI, EEG, EMG, neurological and cognitive testing as well as in depth post-mortem assays to determine if it reproduces biochemical and histopathological aspects of disease. External evaluation of in-life clinical testing will be performed by our clinical collaborator Dr. Tifft. Additionally, Dr. Tifft will make human samples available for comparison with the new sheep model. The biochemical aspects of disease will be externally validated by by Dr. d’Azzo (the leader in field of sialidosis) and pathological features characterized by Dr. Koehler a veterinary neuropathologist. After completion of these studies, this fully validated model will be used to learn more about sialidosis as a disorder, and also used in the development of an adeno associated viral gene therapy or other future treatment strategies for sialidosis.