Epitope targeted AAVS for improved airway delivery

描述(由申请人提供)： 摘要囊性纤维化是高加索人最常见的致死性遗传病。由于CFTR基因的突变，大多数CF患者死于进行性肺部疾病，目前还没有治愈的方法。仅在6-10%的呼吸道上皮细胞中表达CFTRcDNA就可以纠正阴离子转运缺陷，因此基因治疗这种常染色体隐性遗传病具有很大的前景。病毒载体如腺相关病毒(AAV)是将矫正货物运送到呼吸道的最安全的工具之一，然而，低效的运送仍然限制了这一领域的发展。我们建议通过使用新的多肽表位来克服这一限制，这些表位可以有效地结合到分化良好的人和新的CF猪模型的原代CF气道上皮细胞的表面。我们的总体假设是，与CF气道上皮细胞顶面具有亲和力的多肽基序可以是 通过噬菌体淘洗鉴定，并整合到AAV衣壳中，以改善呼吸道的媒介趋向性。新设计的载体将用于通过在分化良好的上皮细胞中添加基因来纠正CF表型。这一合作方案结合了CF和呼吸道上皮细胞生物学方面的专业知识(McCray博士和Zabner博士)以及噬菌体淘洗和载体工程专业知识(Davidson博士和Zabner博士)。这些PI还在基因治疗方面拥有相当多的专业知识。这项建议包括两个目标，在目标1中，我们将使用噬菌体展示文库进行平移，以确定与分化良好的CF呼吸道上皮细胞的粘膜表面具有亲和力的多肽基序。我们将测试将这些基序插入到通过衣壳洗牌策略出现的新的AAV衣壳中，并显示改进的转导轮廓，以改善对CF呼吸道上皮细胞顶面的靶向。在目标2中，我们将使用多肽配体修饰的AAVs来纠正CFTR阴离子转运和宿主防御缺陷在CF气道上皮细胞中的存在。

DESCRIPTION (provided by applicant): Abstract Cystic fibrosis (CF) is the most common lethal genetic disorder among Caucasians. As a consequence of mutations in the CFTR gene, most CF patients die from progressive lung disease for which there is no curative treatment. Expression of the CFTR cDNA in as few as 6- 10% of respiratory epithelia can correct the anion transport defect, and therefore gene therapy holds great promise for this autosomal recessive disease. Viral vectors such as adeno- associated virus (AAV) are among the safest tools available to deliver a corrective cargo to the airways, however, inefficient delivery continues to limit the field. We propose to overcome this limitation by using novel peptide epitopes that bind efficiently to the surface of well-differentiaed primary CF airway epithelia from humans and from a novel CF pig model. Our overall hypothesis is that peptide motifs with affinity to the apical surface of CF airway epithelia can be identified via phage panning, and incorporated into an AAV capsid to improve vector tropism for the airways. The newly engineered vectors will be used to correct the CF phenotype by gene addition in well-differentiated epithelia. This collaborative proposal combines expertise in CF and airway epithelial cell biology (Drs. McCray and Zabner) with expertise in phage panning and vector engineering (Drs. Davidson and Zabner). These PIs also have considerable expertise in gene therapy. This proposal encompasses two aims, In Aim 1, we will use panning with a phage display library to identify peptide motifs with affinity to the mucosal surface of well-differentiaed CF airway epithelia. We will test insert those motifs into new AAV capsids that emerged through capsid shuffling strategies and show improved transduction profiles, for improved targeting to the apical surface of CF airway epithelia. In aim 2, we will use the peptide ligand-modified AAVs to correct the CFTR anion transport and host defense defects in CF airway epithelia.