Creation and correction of Spinal Muscular Atrophy in the pig

猪脊髓性肌萎缩症的产生和矫正

基本信息

批准号：
8702801
负责人：
ARTHUR H. M. BURGHES
金额：
$ 37.95万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-04-01 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8702801
关键词：
Address Affect Age Animal Model Animals Atrophic Axon Birth Blood - brain barrier anatomy Blood Vessels Brain Cause of Death Cells Cerebrospinal Fluid Cessation of life Child Clinical Clinical Trials Dependovirus Detection Development Disease Ensure Family suidae Functional disorder Future Gait Gene Transduction Agent Genes Genetic Human Immunosuppression Infant Injection of therapeutic agent Intervention Intrathecal Injections Limb structure Longevity Messenger RNA Modeling Monkeys Motor Motor Neurons Mus Mutation Neoadjuvant Therapy Neurologic Neurons Pathology Patients Phenotype Production Ramp Route SMN protein (spinal muscular atrophy)SMN2 gene Series Severities Spinal Cord Spinal Ganglia Spinal Muscular Atrophy Staging Symptoms Testing Therapeutic Therapeutic Agents Therapeutic Intervention Time Transcript Treatment Efficacy Treatment Protocols Vascular System Ventilator Virus Walking cell type clinically relevant design gene therapy infant death knock-down mouse model neuron loss programs public health relevance research study restoration small hairpin RNA success transduction efficiency vector

项目摘要

DESCRIPTION (provided by applicant): Spinal muscular atrophy (SMA) is an autosomal recessive disorder which is the most common genetic cause of infant death. SMA is caused by loss or mutation of survival motor neuron 1 gene (SMN1) and retention of the SMN2 gene, which leads to insufficient levels of SMN for motor neurons. Introduction of scAAV9-SMN either through the vascular system or via intrathecal (CSF) delivery in SMA mice results in a marked correction of the phenotype. The advantage of intrathecal delivery is the lower titer of virus that can be used for correction, thus simplifying the production of scAAV9-SMN. While the efficiency of transduction of motor neurons has been investigated in pig and in monkeys, the ability to correct a SMA phenotype in a large animal model has not been investigated. This is due to fact that there is no large animal model of SMA currently available. A large animal model of SMA can be used to address the importance of high SMN levels in motor neurons, the efficiency of SMN transduction required to obtain a clinical benefit, and when SMN needs to be induced to obtain clinical benefit. We therefore propose to optimize scAAV9-SMN delivery to motor neurons in a clinically relevant large animal porcine model of SMA and develop gene therapy protocols for the treatment of SMA in this model. In particular, we will address transduction efficiency including repeat administration and when SMN must be introduced to correct a pig model of SMA. We will create the pig SMA model by reducing SMN in the required neurons using scAAV9- shRNA: SMN knockdown. The shRNA is specific for pig SMN and does not affect the levels of human SMN. The shRNA is delivered via intrathecal delivery of a scAAV9shRNA construct that expresses SMN and GFP. Injection is performed in 5-day-old piglets and EMG, clinical presentation, and a ramp test will be used to evaluate the affected pig. The GFP can be used to follow the efficiency of motor neuron transduction and EMG to follow the progression of functional motor neuron loss. We will then introduce scAAV9-SMN at various stages of the disease including pre-symptomatic, at the first signs of phenotype, and later stages when the hind limbs are severely weak. Furthermore, we will determine if restoration of SMN levels must occur prior to CMAP and MUNE reduction, which will in turn indicate when SMN needs to be restored in SMA patients to obtain the greatest clinical benefit. These experiments will set the parameters for all SMN induction therapies in SMA and allow determination of when to introduce a therapeutic agent.

描述（由申请人提供）：脊髓性肌萎缩症（SMA）是一种常染色体隐性遗传疾病，是婴儿死亡最常见的遗传原因。 SMA 是由运动神经元存活基因 1 (SMN1) 缺失或突变以及 SMN2 基因保留引起的，导致运动神经元 SMN 水平不足。通过血管系统或鞘内 (CSF) 递送将 scAAV9-SMN 引入 SMA 小鼠中，可显着纠正表型。鞘内递送的优点是病毒滴度较低，可用于校正，从而简化scAAV9-SMN的生产。虽然已经在猪和猴子中研究了运动神经元转导的效率，但尚未研究在大型动物模型中纠正 SMA 表型的能力。这是因为目前还没有大型 SMA 动物模型。 SMA 的大型动物模型可用于解决运动神经元中高 SMN 水平的重要性、获得临床益处所需的 SMN 转导效率，以及何时需要诱导 SMN 以获得临床益处。因此，我们建议在临床相关的大型动物SMA猪模型中优化scAAV9-SMN向运动神经元的递送，并开发用于治疗该模型中的SMA的基因治疗方案。特别是，我们将解决转导效率问题，包括重复给药以及何时必须引入 SMN 来纠正 SMA 猪模型。我们将通过使用 scAAV9-shRNA：SMN 敲低来减少所需神经元中的 SMN，从而创建猪 SMA 模型。 shRNA 对猪 SMN 具有特异性，不会影响人 SMN 的水平。 shRNA 通过表达 SMN 和 GFP 的 scAAV9shRNA 构建体进行鞘内递送。对 5 日龄的仔猪进行注射，并通过肌电图、临床表现和斜坡测试来评估受影响的猪。 GFP 可用于跟踪运动神经元转导的效率，而 EMG 可用于跟踪功能性运动神经元损失的进展。然后，我们将在疾病的各个阶段引入 scAAV9-SMN，包括症状前、表型的最初迹象以及后肢严重无力时的后期阶段。此外，我们将确定 SMN 水平的恢复是否必须在 CMAP 和 MUNE 降低之前进行，这反过来将表明 SMA 患者何时需要恢复 SMN 以获得最大的临床益处。这些实验将为 SMA 中的所有 SMN 诱导疗法设定参数，并确定何时引入治疗剂。