Determinants of AAV Tropism

描述（由申请人提供）：重组AAV载体是用于几种人类遗传性疾病的基因治疗的有希望的候选物。尽管在临床试验中取得了早期的成功，但我们对AAV宿主和组织嗜性的机制的理解是不完整的。在上一个资助期间，我们在绘制新型AAV-聚糖受体相互作用和设计一类新的肝脏去靶向AAV载体方面取得了重大进展。此外，我们还优化了结构工具、体外细胞试验和体内动物研究， AAV载体的表征。尽管这些信息提供了对某些AAV血清型的生物学的进一步了解，但我们对目前在临床研究中测试的许多其他AAV毒株的理解存在重大差距。在目前的提案中，我们已经组建了一个由结构病毒学，糖生物学，血管生物学和AAV载体领域经验丰富的研究人员组成的团队，以剖析几种天然存在的以及新的实验室衍生的AAV突变体的生物学。具体来说，我们将使用一系列分子建模、计算配体对接、聚糖阵列和分子克隆工具来绘制AAV-聚糖相互作用的结构决定因素。然后，我们提出在不同的AAV血清型上重新设计聚糖结合足迹，以产生具有改变的组织向性的新突变体。还将使用小鼠模型进行调查不同组织聚糖以及整合素共受体对AAV载体的向性、生物分布、药代动力学和经血管转运的作用的综合研究。所提出的研究的长期目标是获得对不同宿主中的AAV生物学的透彻理解。如果成功，目前的建议可以提供重要的新见解聚糖和整合素对AAV组织向性的影响。除了为改进的AAV载体的结构驱动设计提供路线图之外，所提出的研究还可以指导选择适当的AAV株用于进一步的临床开发。

DESCRIPTION (provided by applicant): Recombinant AAV vectors are promising candidates for gene therapy of several human genetic disorders. Despite early successes in clinical trials, our understanding of the mechanisms underlying AAV host and tissue tropism is incomplete. During the previous funding period, we made significant progress in mapping novel AAV-glycan receptor interactions and engineering a new class of liver-detargeted AAV vectors. In addition, we optimized structural tools, in vitro cellular assays and in vivo animal studies pertinent to the characterization of AAV vectors. Although this information provided further insight into the biology of certain AAV serotypes, significant gaps exist in our understanding of numerous other AAV strains currently being tested in clinical studies. In the current proposal, we have assembled a team of experienced investigators in the fields of structural virology, glycobiology, vascular biology and AAV vectors to dissect the biology of several naturally occurring as well as new, lab-derived AAV mutants. Specifically, we will map the structural determinants of AAV-glycan interactions using a battery of molecular modeling, computational ligand docking, glycan array and molecular cloning tools. We then propose to reengineer glycan binding footprints on different AAV serotypes to generate novel mutants with altered tissue tropism. Comprehensive studies investigating the role of different tissue glycans as well as integrin co-receptors on the tropism, biodistribution, pharmacokinetics, and transvascular transport of AAV vectors using mouse models will also be undertaken. The long term goal of the proposed studies is to obtain a thorough understanding of AAV biology in different hosts. If successful, the current proposal could provide significant new insight into the influence of glycans and integrins on AAV tissue tropism. In addition to providing a roadmap for structure-driven design of improved AAV vectors, the proposed studies could guide the selection of appropriate AAV strains for further clinical development.