Determinants of AAV Lung Tropism

摘要 呼吸道病原体，如人类副流感病毒和SARS冠状病毒有效 通过利用多种细胞表面聚糖和糖蛋白受体感染人类气道 在气道细胞类型中遇到。在腺相关病毒（AAV）的情况下，血清型1、5、6和更多 最近，已经显示AAV 9在体外和体内有效地阻断气道，尽管具有显著的 物种特异性和模式特异性差异。这项提案的目标是阐明分子和 AAV血清型中气道向性的细胞决定因素。为此，我们制定了一套全面的 该方法通过诱变、生化试剂和生物化学试剂对AAV衣壳进行分子操作， 用于鉴定介导AAV气道细胞进入的聚糖和共受体以及相关的体外模型， 人体呼吸道。本文所述的策略将（a）定义AAV衣壳结构元件， 确定气道向性氨基酸水平，和（B）能够鉴定细胞表面组分 包括决定AAV对不同气道细胞类型的向性的聚糖和整联蛋白亚单位。拟议 这些研究也将有助于全面了解AAV进入气道细胞的机制 为深入了解AAV气道向性的物种特异性差异提供了依据。如果成功的话，这些知识可以 促进AAV载体的改进以及集中于基因治疗的临床前/临床研究的设计。 治疗呼吸道疾病，如囊性纤维化。

ABSTRACT Respiratory tract pathogens such as the human parainfluenza virus and SARS coronavirus effectively infect human airways by exploiting a diverse set of cell surface glycans and glycoprotein receptors encountered in airway cell types. In case of adeno-associated viruses (AAV), serotypes 1, 5, 6 and more recently, AAV9 have been shown to efficiently transduce airways in vitro and in vivo, albeit with striking species-specific and serotype-specific differences. The goal of this proposal is to elucidate molecular and cellular determinants of airway tropism in AAV serotypes. To achieve such, we have devised a comprehensive approach that hinges on molecular manipulation of AAV capsids through mutagenesis, biochemical reagents for identification of glycans and co-receptors that mediate AAV airway cell entry and relevant in vitro models of the human respiratory tract. The strategies described herein will (a) define AAV capsid structural elements at the amino acid level that determine airway tropism, and (b) enable identification of cell surface components including glycans and integrin subunits that dictate AAV tropism for different airway cell types. The proposed studies will help provide a comprehensive picture of the mechanisms underlying AAV airway cell entry as well as provide insight into species-specific differences in AAV airway tropism. If successful, this knowledge may facilitate improvements in AAV vectors as well as in the design of preclinical/clinical studies focused on gene therapy of airway diseases such as cystic fibrosis.