The development of gene therapy for Niemann-Pick type C disease

尼曼-皮克C型疾病(NPC)是一种毁灭性的、最终致命的遗传疾病，严重影响大脑和身体的其他器官。诊断通常是在婴儿期/儿童期做出的，大约一半的患者有肝脏异常。这些患者中有10%将在六个月前死于肝功能衰竭。鼻咽癌患者在婴儿时期存活下来的症状主要是大脑的进行性退化，导致虚弱的后果和随后的死亡，通常是在生命的第二个十年。在绝大多数病例中，导致鼻咽癌的基因缺陷是由一种名为NPC1的基因突变引起的。这会导致体内细胞中大量复杂物质的积累，如胆固醇，从而导致大脑和内脏器官出现的症状。目前，还没有针对鼻咽癌患者的主要特效疾病治疗方法，开发有效的挽救生命的疗法的需求是压倒性的。基因治疗包括将基因的功能性拷贝送入细胞，以弥补缺陷版本。在大多数情况下，基因是通过病毒载体传递到细胞中的。这些载体是以去除有害成分的病毒为基础的，使其变得安全，并被用作一种载体，将治疗性基因有效地输送到人体细胞。使用这种方法，基因治疗在临床试验中证明了挽救生命和改变生命的结果。本项目的目标是开发治疗鼻咽癌的基因疗法。我们已经制造了一种病毒载体，可以有效地将NPC1基因的治疗性版本输送到细胞中。我们建议在临床前的概念验证基因治疗研究中使用该载体来挽救鼻咽癌的小鼠模型，该模型概括了人类患者的症状。本研究项目的目的是：(1)将携带治疗性NPC1基因的病毒载体直接导入新生的NPC小鼠的大脑，通过一系列分析，评估这是否改善了NPC1基因的治疗效果，并防止了脑内的症状。我们的初步数据表明，这具有显著的治疗作用，但还需要进一步的研究；(2)将病毒载体静脉注射到新生的NPC小鼠的血液中，并评估治疗效果。众所周知，这种病毒载体可以将基因输送到内脏器官，也可以在静脉注射后进入大脑。这种方法可用于治疗大脑和内脏器官的症状，如肝脏症状；(3)可同时将病毒载体直接和静脉输送至大脑，以有效增加全身用药剂量并评估疗效；(4)以治疗效率最高的途径(目标1、2和3中确定)将病毒载体递送给渐进性老龄小鼠并评估治疗效果。这样做的目的是为了评估基因疗法在更有症状的小鼠身上的表现。这将模拟鼻咽癌的潜在临床情况，即鼻咽癌的诊断可能会推迟，患者可能会出现症状。我们希望这项研究能为基因治疗作为鼻咽癌患者有效治疗方法的临床应用提供概念验证数据。

Niemann-Pick type C disease (NPC) is a devastating and ultimately fatal genetic disorder that profoundly affects the brain and other organs of the body. A diagnosis is usually made in infancy/childhood where approximately half of the patients have liver abnormalities. Ten per cent of these patients will die from liver failure before six months of age. The symptoms of NPC patients that survive infancy are dominated by progressive degeneration of the brain with debilitating consequences and subsequent death, usually in the second decade of life. The genetic defect causing NPC in the vast majority of cases is caused by mutations in a gene called NPC1. This causes an accumulation of a number of complex substances such as cholesterol in cells of the body resulting in the symptoms seen the brain and visceral organs. Currently, there are no major specific disease modifying treatments for NPC patients and the need to develop effective life-saving therapies is overwhelming. Gene therapy involves the delivery of a functional copy of a gene into a cell in order to compensate for a defective version. In most cases, the genes are delivered into cells using viral vectors. These vectors are based on viruses that have been stripped of their harmful components, rendered safe and used as a vehicle to efficiently deliver therapeutic genes to cells of the body. Using this approach, gene therapy has demonstrated life-saving and life-changing results in clinical trials. The objective of this project is to develop gene therapy for treating NPC. We have made a viral vector that can efficiently deliver a therapeutic version of the NPC1 gene into cells. We propose using this vector in a pre-clinical proof-of-concept gene therapy study to rescue a mouse model of NPC that recapitulates the symptoms seen in human patients. The aims of this research project are: (1) To deliver the viral vector carrying the therapeutic NPC1 gene directly into the brain of new-born NPC mice to assess, using a range of analyses, whether this ameliorates of prevents the symptoms in the brain. Our preliminary data shows that this has significant therapeutic affect but requires further investigation; (2) To deliver the viral vector intravenously into the bloodstream of new-born NPC mice and assess the therapeutic affect. This viral vector is known to deliver genes to visceral organs and also cross into the brain following intravenous injection. This approach offers potential treatment for both the brain and visceral organs symptoms such as in the liver; (3) To simultaneously deliver the viral vector both directly to the brain and also intravenously to effectively increase the systemic dose and assess the therapeutic efficacy; (4) To deliver the viral vector using the most therapeutically efficacious route of administration (identified in aims 1, 2 and 3) to progressively older mice and assess the therapeutic effect. The purpose of this is to gauge how the gene therapy may perform in more symptomatic mice. This would mimic the potential clinical situation where diagnosis of NPC may be delayed and patients have developed symptoms. It is our hope that this study will provide proof-of-concept data supporting the clinical application of gene therapy as an effective treatment for NPC patients.