Electroporation-mediated Pulmonary Gene Transfer for Acute Lung Injury

项目总结/摘要 急性肺损伤（ALI）、急性呼吸窘迫综合征（ARDS）和新生儿 呼吸窘迫综合征（NRDS）是一种常见的、破坏性的临床综合征， 大量成人和新生儿患者（美国每年20万例）， 按照目前的治疗标准死亡率是40%我们的最终目标是开发非病毒基因 治疗ALI的方法。我们已经证明，电穿孔可以用于转移 将基因转移到肺中，以治疗已确定的肺损伤和肺水肿， 减轻伤害的严重性。该方法简便、快速、安全。为了移动这个 为了临床应用，我们必须在大型动物模型中证明其有效性和安全性 它代表了人类的疾病，并开发出最简单和安全的方法， 让它被临床医生接受。动物模型的最高推荐是 好的证据 该研究的结果将在临床试验中相似，为此，我们 我最近将这种方法应用于一组猪（35-40 kg），使用的能量低于0.1 J/kg 并实现了基因转移到肺部，没有死亡或损伤。我们的R21阶段特定 目的是（1）优化电穿孔参数，以安全地将基因转移到肺中， 猪，（2）评估DNA递送方法，（3）开发和优化基于支气管镜的 用于肺基因递送的电极，以及（4）确定ENaC以及 Na，K-ATP酶亚单位基因增加基因治疗肺损伤的疗效 小鼠模型在R21阶段结束时，我们将建立最佳参数， 基因传递到猪肺，并确定了最佳的治疗基因组合， 在两种互补的猪ALI模型中治疗肺损伤。R33阶段的具体目标是（1） 电穿孔介导Na，K-ATPase/ENaC基因转移的效果评价 亚单位，以防止肺损伤的盐水灌洗模型的轻中度ALI，（2）确定 这些基因的转移是否可以治疗该模型中预先存在的肺损伤，（3）确定 这些基因的转移是否可以保护严重脓毒症诱导的盲肠炎患者免受肺损伤， 结扎和穿刺模型的ARDS，和（4）评估这种方法是否可以用于 治疗先前建立的脓毒症诱导的肺损伤。这些研究将提供 我们与经胸肺电穿孔的原理证明，并建立 在两个已建立的临床前模型中的安全性和有效性。项目叙述 急性肺损伤、急性呼吸窘迫综合征（ARDS）和新生儿 呼吸窘迫综合征是一种常见的、破坏性的临床综合征， 据估计，美国每年有20万成人和新生儿患者， 按照目前的治疗标准死亡率是40%我们开发了一种非病毒基因 使用电场的治疗方法可以预防和治疗现有的ARDS， 两个小动物模型。这项研究中提出的工作将把这些发现扩展到 一个大型动物模型，以评估这些发现是否可以转化为更大的 实验对象最终是人类在研究结束时，我们将证明 在已建立的临床前模型中证实了这种新方法治疗肺损伤的功效。

PROJECT SUMMARY/ABSTRACT Acute lung injury (ALI), Acute Respiratory Distress Syndrome (ARDS), and Neonatal Respiratory Distress Syndrome (NRDS) are common, devastating clinical syndromes that affect large numbers of adult and neonatal patients (200,000 cases in the US per year) and have up to 40% mortality with the current standard of care. Our ultimate goal is to develop nonviral gene therapy approaches to treat ALI. We have shown that electroporation can be used to transfer genes to lungs with established lung injury and pulmonary edema to treat the disease and lessen the severity of the injury. The method is simple, fast, and safe. In order to move this toward clinical application, we must demonstrate its efficacy and safety in a large animal model that represents the human disease and develop the most simple and safe method possible in order for it to be accepted by clinicians. The highest recommendation for an animal model is good evidence that the results of the study would be similar in a clinical trial To this end, we have recently applied this approach to a group pigs (35-40 kg) using energies less than 0.1 J/kg and have achieved gene transfer to the lungs with no mortality or injury. Our R21 Phase specific aims are to (1) optimize electroporation parameters for safety and gene transfer to the lungs of pigs, (2) evaluate methods of DNA delivery, (3) develop and optimize a bronchoscope-based electrode for lung gene delivery, and (4) determine whether gene transfer of ENaC as well as Na,K-ATPase subunit genes increases efficacy of gene therapy for treatment of lung injury in a mouse model. At the end of the R21 phase, we will have established optimal parameters for gene delivery to the pig lung and have determined the best therapeutic gene combination to treat lung injury in two complementary pig models of ALI. The R33 phase specific aims are (1) evaluate efficacy of electroporation-mediated gene transfer of Na,K-ATPase/ENaC gene subunits to prevent lung injury in a saline lavage model of mild-moderate ALI, (2) determine whethere transfer of these genes can treat pre-existing lung injury in this model, (3) determine whether transfer of these genes can protect from lung injury in a severe sepsis-induced cecal ligation and pucture model of ARDS, and (4) evaluate whether this approach can be used to treat previously established sepsis-induced lung injury in this model. These studies will provide us with the proof-of-principle for transthoracic pulmonary electroporation and establish the safety and efficacy in an two established pre-clinical models. PROJECT NARRATIVE Acute lung injury, Acute Respiratory Distress Syndrome (ARDS), and Neonatal Respiratory Distress Syndrome are common, devastating clinical syndromes that affect an estimated 200,000 adult and neonatal patients each year in the US and have up to 40% mortality with the current standard of care. We have developed a nonviral gene therapy approach using electric fields that can prevent as well as treat existing ARDS in two small animal models. The work proposed in this study will extend these findings to a large animal model to evaluate whether these findings can be translated into larger subjects and ultimately humans. At the end of the study, we will have demonstrated efficacy of this novel approach to treat lung injury in an established pre-clinical model.