Optimized Gene Replacement for AAT deficiency and Modeling of Clinical Outcomes in small and large animal models

针对 AAT 缺陷的优化基因替换以及小型和大型动物模型中的临床结果建模

• 批准号: 10463807
• 负责人: Terence R. Flotte
• 金额: $ 43.22万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-09 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10463807
• 关键词: Age; Albumins; Alleles; Animal Model; Animals; Biological Markers; CRISPR/Cas technology; Capsid; China; Chronic Obstructive Pulmonary Disease; Clinical; Clinical Trials; Clustered Regularly Interspaced Short Palindromic Repeats; Collaborations; Consumption; Densitometry; Development; Disease model; Dose; Elastin; Enterobacteria phage P1 Cre recombinase; Environment; Estrogen Receptors; European; Exposure to; Ferrets; Future; Gene Transduction Agent; Genes; Genetic; Genetic Diseases; Genetic study; Glycoproteins; Histology; Homozygote; Human; Human Genetics; Impairment; Inflammatory; Infusion procedures; Intravenous infusion procedures; Iowa; Knock-in; Knock-out; Knockout Mice; Laboratories; Leukocyte Elastase; Life; Liver; LoxP-flanked allele; Lung; Lung CAT Scan; Lung diseases; Mediating; Mendelian disorder; Modeling; Modification; Mus; Mutation; Outcome; Outcome Measure; Patients; Phenotype; Physiology; Plasma; Population; Problem Solving; Process; Prospective Studies; Protease Inhibitor; Proteins; Pulmonary Emphysema; Pulmonary Inflammation; Radiology Specialty; Randomized Clinical Trials; Serum; Signal Transduction; Stimulus; Structure of parenchyma of lung; Tamoxifen; Testing; Time; Tissue Preservation; Transgenes; Transgenic Animals; Transgenic Organisms; Universities; airway obstruction; alpha 1-Antitrypsin Deficiency; clinical efficacy; comparative efficacy; density; design; effectiveness evaluation; gene replacement; gene therapy; improved; innovation; interstitial; lung histology; lung preservation; mutant; neutrophil; nonhuman primate; novel; novel strategies; novel therapeutics; product development; promoter; prospective; pulmonary function; recombinant virus; reconstitution; response; targeted treatment; tool; vector

Project Summary (Project 1) Alpha-1 antitrypsin deficiency (AATD) is a common single gene disorder caused by mutations in the SERPINA1 gene, which normally encodes a very abundant serum antiprotease, whose primary function is to protect the interstitial elastin matrix of the lung from degradation by neutrophil elastase (NE). The E242K (PI*Z) mutant allele is very common among those of European ancestry, and E342K homozygotes encode a protein with impaired secretion, resulting in deficient serum levels, leading to unrestrained NE activity, degrading pulmonary elastin. This process triggers lung inflammation and leads to loss of lung elastic recoil and airways obstruction, creating the clinical picture of emphysema, which is the life-limiting effect of AATD in most patients. Current AATD therapy consists of weekly IV infusion of AAT, but this therapy has never been proven to benefit lung function in patients in prospective studies, leading to questions about whether the previously identified serum target level (11 µM) is sufficient, or whether gene therapy to restore such a level would be efficacious. Our laboratory has previously developed rAAV gene therapy vectors and used them in clinical trials. Those trials have failed to achieve the target, achieving levels just over 0.5 µM but being sustained over 5 years in patients. We have also use CRISPR-based technology to produce the first SERPINA1-knockout mice and have begun a collaboration with the University of Iowa group, who have created both SERPINA1-knockout and SERPINA1-E342K ferret models. In the current proposal, improved and optimized vectors will be tested in both mice and ferret models. In the ferrets, they will be tested alongside sophisticated conditional transgenic reconstitution to determine whether the optimal target for therapy is 11 µM vs. 25 µM. These studies will pave the way for better rAAV-AAT vectors appropriate for future clinical product development. In this project, we propose to study genetic emphysema due to alpha-1 antitrypsin deficiency (AATD). AATD is both fairly common as a genetic disease and is a model of much more common causes of chronic obstructive pulmonary disease (COPD). In the proposal, we will use advanced gene editing tools to create genetically defined animal models of AATD (known as transgenic animals), both in mice and in ferrets, which are a good model to study lung diseases. In the course of the study, we will use both additional transgenic approaches and modified recombinant viruses (rAAV) to genetically treat AATD in these models and examine how we might design evaluate the effectiveness of future gene therapies for genetic emphysema.

项目摘要（项目1） α-1抗胰蛋白酶缺乏症（AATD）是由突变引起的常见单基因疾病 Serpina1基因通常编码非常丰富的血清抗蛋白酶 主要功能是保护肺的间质弹性蛋白基质免受降解 中性粒细胞弹性酶（NE）。 E242K（PI*Z）突变等位基因在 欧洲血统和E342K纯合子编码一种蛋白质，分泌物受损， 导致血清水平不足，导致NE活性不受限制，降解肺 弹性。该过程触发肺注射并导致肺弹性后坐力丧失和 气道目标，创建肺气肿的临床图片，这是生命的影响 大多数患者的AATD。当前的AATD疗法包括每周的IV AAT输注，但这 在前瞻性研究中，从未证明治疗在患者中受益于肺功能 导致有关先前确定的血清靶标水平（11 µm）的问题 足够，或者是否有效地恢复了这种水平的基因疗法。我们的实验室 先前已经开发了RAAV基因疗法载体，并将其用于临床试验。那些 试验未能达到目标，达到超过0.5 µm的水平，但持续 患者超过5年。我们还使用基于CRISPR的技术来生产第一个 Serpina1-Knockout小鼠，并已与爱荷华大学集团合作， 他们同时创建了Serpina1-Knockout和Serpina1-E342K雪貂模型。在 当前的建议，改进和优化的向量将在小鼠和雪貂模型中进行测试。 在雪貂中，它们将与复杂的有条件转基因重构一起进行测试 确定治疗的最佳目标是否为11 µm，而25 µm。这些研究将铺路 更好的RAAV-AAT向量的方式适合将来的临床产品开发。 在这个项目中，我们建议研究由于α-1抗胰蛋白酶缺乏而导致的遗传性肺气肿 （AATD）。 AATD既作为遗传疾病都相当普遍，并且是更多的模型 慢性阻塞性肺疾病（COPD）的常见原因。在提案中，我们将使用 高级基因编辑工具，以创建AATD的一般定义的动物模型（称为 转基因动物），无论是小鼠还是在雪貂中，这是研究肺部疾病的好模型。 在研究过程中，我们将同时使用其他转基因方法和修改 重组病毒（RAAV）在这些模型中对AATD进行基因处理，并研究我们如何 设计可能会评估未来基因疗法对遗传性肺气肿的有效性。