A Hybrid Viral/Nonviral Vector for CFTR Delivery to CF Pig Airways

用于 CFTR 递送至 CF 猪气道的混合病毒/非病毒载体

• 批准号: 9923461
• 负责人: PATRICK L SINN
• 金额: $ 57.52万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9923461
• 关键词: Address; Adenovirus Vector; Animal Model; Anions; Basal Cell; Biliary cirrhosis; Categories; Cell surface; Cells; Characteristics; Clinical; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; DNA; DNA Transposons; Data; Defect; Disease; Disease Outcome; Disease Progression; Dose; Electrolytes; Emerging Technologies; Engineering; Epithelial Cells; Exhibits; Exocrine pancreatic insufficiency; Failure; Family suidae; Gene Delivery; Gene Expression; Gene Transfer; Gene therapy trial; Genes; Genetic Diseases; Genome; Genomics; Goals; Guide RNA; Host Defense; Human; Hybrids; Ileus; In Vitro; Infection; Inflammation; Intestines; Ion Transport; Lentivirus Vector; Life; Lobe; Lung; Lung diseases; Measures; Meconium; Mendelian disorder; Methods; Modeling; Mucous body substance; Mus; Neonatal; Newborn Infant; Non-Viral Vector; Nose; Outcome; Phenotype; Population; Pulmonary Cystic Fibrosis; Quality of life; Reagent; Reporter Genes; Reporting; Research; Research Personnel; Structure of respiratory epithelium; Surface; System; Testing; Therapeutic; Transgenes; Transposase; Treatment Efficacy; Viral; Viral Vector; adeno-associated viral vector; aerosolized; airway epithelium; airway obstruction; base; cell type; cellular transduction; cystic fibrosis mouse; disease phenotype; disease-causing mutation; disorder prevention; efficacy testing; experimental study; falls; gene replacement; gene therapy; genetic approach; gutless adenoviral vector; improved; in vivo; innovation; knockout gene; novel; novel strategies; overexpression; preclinical study; prevent; progenitor; promoter; stem cells; success; therapeutic evaluation; therapeutic gene; tool; uptake; vector

Project Summary/Abstract Gene therapy was first proposed as a viable therapeutic strategy for genetic disease nearly 30 years ago. There are many notable recent successful gene therapy trials for multiple diverse diseases; however, gene therapy for cystic fibrosis (CF) has presented many unexpected roadblocks. For example, 1) airway cells have evolved many barriers to prevent uptake of foreign DNA, 2) mouse models of CF generally lack lung disease, and 3) each of the standard gene delivery vehicles have pros and cons. We have a demonstrated track record of utilizing many categories of viral and non-viral based reagents for gene delivery to the airways. Viral based vectors are the most efficient way to deliver genes to airway cells. Our long-term goal is to engineer a reagent for gene delivery to cells that is an effective therapeutic for CF lung disease, yet there is a critical need for improved gene delivery tools. Here, we develop a novel reagent. We propose to demonstrate that our engineered vector can integrate into the genome of progenitor cells in the airways and express the transgene in the appropriate surface cells. To this end, we have combined the efficiency of an adenoviral-based vector with the persistent expression of a DNA transposon-based non-viral vector. In addition, we test this `hybrid' vector in a relevant large animal model of CF. Here we demonstrate that this vector will: 1) deliver CFTR to CF cells and correct the anion transporter defect in vitro using a novel strategy to focus cellular integration and expression, 2) verify that the necessary levels of gene transfer are achievable in pigs airways in vivo, and 3) correct the anion channel defect and disease progression in the airways of a pig CF model. We investigate a gene delivery tool that can be used to address questions that no other reagent can address. There are >2000 known disease causing mutations in CFTR. Our goal is to provide a life-long gene replacement strategy that would be efficacious regardless of the disease causing mutation. This proposed research is highly innovative. Combining the emerging technologies of DNA transposon-based vectors with well-studied adenoviral delivery presents an exciting new opportunity to improve the utility of delivery vehicles for therapeutic genes such as CFTR. The reagents, methods, and data generated by these experiments will provide guidance for gene therapies for other monogenic disorders, thereby significantly advancing the gene therapy field.

项目总结/摘要 基因治疗是近30年前首次提出的一种可行的治疗遗传性疾病的策略。 最近有许多值得注意的成功的基因治疗试验用于多种不同的疾病;然而，基因治疗是一种非常有效的方法。 囊性纤维化（CF）的治疗出现了许多意想不到的障碍。例如，1）气道细胞具有 进化出许多屏障以防止外来DNA的摄取，2）CF的小鼠模型通常没有肺部疾病， 和3）每种标准基因递送工具都有优点和缺点。我们有良好的记录 利用许多种类的基于病毒和非病毒的试剂将基因递送到气道。基于病毒的 载体是将基因递送到气道细胞的最有效方式。我们的长期目标是设计一种试剂 对于将基因递送至细胞，这是CF肺病的有效治疗方法，但迫切需要 改进的基因传递工具。在这里，我们开发了一种新的试剂。我们建议证明我们的 工程化载体可以整合到气道中的祖细胞的基因组中并表达转基因 在适当的表面细胞中。为此，我们结合了基于腺病毒载体的效率 与基于DNA转座子的非病毒载体的持续表达。此外，我们测试这种“混合” 载体在CF的相关大型动物模型中。在这里，我们证明了这个载体将：1）将CFTR传递给CF 细胞和纠正阴离子转运蛋白缺陷，在体外使用一种新的策略，集中细胞整合， 表达，2）证实在猪体内气道中可实现必要水平的基因转移，和3） 纠正猪CF模型气道中的阴离子通道缺陷和疾病进展。我们研究了 基因递送工具，可用于解决其他试剂无法解决的问题。>2000 导致CFTR突变的已知疾病。我们的目标是提供一种终生的基因替换策略， 无论致病突变是什么都是有效的。这项研究具有很强的创新性。 将基于DNA转座子的载体的新兴技术与充分研究的腺病毒递送相结合 提供了一个令人兴奋的新机会，以提高治疗基因，如 CFTR。这些实验产生的试剂、方法和数据将为基因工程提供指导。 其他单基因疾病的治疗，从而显着推进基因治疗领域。