Internal and External Validation of a Porcine Dystrophinopathy Model

将基本的生物医学知识转化为有效的治疗方法取决于经过验证的准确反映人类疾病的动物模型以及临床上相关和有效的结果衡量标准。虽然人们普遍认识到营养不良症(BDMD)患者骨骼肌功能的进行性丧失，但疾病的病因正在演变为包括严重的心肌病和心源性猝死，主要是由于有效的呼吸支持治疗。虽然现有的模型一直是并将继续成为机械研究和初步发现调查的有用工具，但固有的局限性要求继续开发用于翻译研究的大型动物模型。因此，有一种尚未得到满足的需求，即通过临床相关和验证的方法来评估明确和可预测地概括由营养不良引起的进行性功能障碍的BDMD的新模型。我们的长期目标是实现对BDMD的详细了解，以便开发和测试有效的干预策略。为了促进这些翻译研究，我们最近启动了一种新的、自发发生的猪BDMD模型的特征(外显子41点突变)。这项应用的目标是进行专注于进行性骨骼肌和心肌功能障碍的内部和外部验证研究，以便将这种自发发生的模型插入药物发现流水线。这一目标得到了强有力的生物学和翻译理论的支持，证明了这种疾病的遗传原因与人类疾病一致。此外，我们以前的工作和初步数据显示，由此导致的进行性骨骼肌损伤和功能障碍，以及进行性心肌病，也与人类营养不良症一致。在R21 Aim 1期间这项建议的内部验证阶段，我们将在内部验证疾病进展期间的运动和呼吸功能的客观测量。在R21 Aim 2中，随着疾病的进展，我们还将应用经过充分验证的超声心动图和电生理学方法在该模型中使用。我们将与我们的临床合作伙伴一起确定我们的措施是否经过了内部验证。如果是这样的话，我们将进行外部验证研究。在R33阶段，为了证明预测的有效性，我们将用BDMD患者广泛使用的药理药物治疗受影响的猪。具体地说，为了改善骨骼肌功能，将用泼尼松治疗猪，为了改善心脏功能，将用赖诺普利治疗，因为这两种药物都被反复证明分别改善骨骼肌和心肌功能。本申请中提出的研究具有创新性，因为它的目标是在一种新的微型猪肌营养不良模型中对进行性骨骼肌和心肌功能障碍进行内部和外部验证。这项研究对药物发现具有重要意义，因为它将首次在人体大小的模型中测试一系列治疗方法(传统和基因治疗)，该模型密切反映人类的病理生理进程，特别是在心血管系统。

The translation of basic biomedical knowledge to the development of effective therapeutic approaches depends on validated animal models that accurately reflect human diseases and clinically relevant and valid outcome measures. While it is widely appreciated that dystrophinopathy (BDMD) patients suffer progressive loss of skeletal muscle function, disease etiology is evolving to include significant cardiomyopathy and sudden cardiac death due largely to effective respiratory support therapy. While existing models have been and continue to be useful tools for mechanistic studies and initial discovery investigations, inherent limitations necessitate the continued development of large animal models for translational research. Hence, there is an unmet need to internally and externally validate novel models of BDMD that clearly and predictively recapitulate progressive dysfunction caused by dystrophinopathy as assessed by clinically relevant and validated measures. Our long-term goal is to achieve a detailed understanding of BDMD to enable development and testing of effective interventional strategies. To facilitate these translational studies, we have recently initiated the characterization of a novel, spontaneously occurring porcine BDMD model (exon 41 point mutation). The objective of this application is to perform internal and external validation studies focused on progressive skeletal and cardiac muscle dysfunction in order to insert this spontaneously occurring model into the drug discovery pipeline. This objective is buttressed by strong biological and translational rationale demonstrating a genetic cause of the disease consistent with human disease. Further, our previous work and preliminary data show resultant, progressive skeletal muscle injury and dysfunction, and progressive cardiomyopathy, which are also consistent with human dystrophinopathy. In the internal validation phase of this proposal during R21 Aim 1, we will internally validate objective measures of locomotion and respiratory function during disease progression. In R21 Aim 2 we will also apply well-validated echocardiography and electrophysiology approaches for use in this model as the disease advances. Together with our clinical partner we will determine if our measures have been internally validated. Should that be the case we will proceed to external validation studies. In the R33 phase, to demonstrate predictive validity, we will treat affected pigs with pharmacological agents used widely by BDMD patients. Specifically, to improve skeletal muscle function pigs will be treated with prednisone, and to improve cardiac function pigs will be treated with Lisinopril as both have repeatedly been shown to improve skeletal and cardiac muscle function, respectively. The research proposed in this application is innovative because it aims to internally and externally validate progressive skeletal and cardiac muscle dysfunction in a novel, miniature porcine muscular dystrophy model. This research is significant for drug discovery because it would enable, for the first time, testing of an array of treatment approaches (conventional and gene therapy) in a human-sized model that closely reflects human pathophysiological progression, particularly in the cardiovascular system.