Novel AAVs for Gene Therapy

用于基因治疗的新型 AAV

• 批准号: 7154979
• 负责人: Mark A Kay
• 金额: $ 41.47万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-29 至 2006-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7154979
• 关键词: adeno associated virus group; biotechnology; capsid; cooperative study; dogs; gene delivery system; gene therapy; hemophilia As; hemophilia B; immune response; laboratory mouse; neutralizing antibody; protein sequence; recombinant proteins; transfection /expression vector

We have demonstrated that AAV-2 vectors can transduce human hepatocytes in vivo. However, AAV-mediated gene transfer is limited due to pre-existing humoral immunity and transgene expression is short-lived because of an immune response directed against the capsid containing hepatocytes. The goal of this project is to develop a novel class of human gene transfer vector based on adeno-associated virus (AAV) that combines high efficiency in vivo transduction of target tissues, with the capability to circumvent or overcome host immunologic responses in humans including the existence of neutralizing antibodies in the human population that may limit their efficacy. These novel vectors will be derived by 'molecular evolution' of DNA from an array of naturally occurring AAV serotypes. This array will comprise previously reported AAV variants from humans and primates, as well as new AAVs of non-primate origin whose isolation is our primary specific aim. The genomes of the various AAV isolates will be 'shuffled' to generate a DNA pool from which we will then generate a library of recombinant AAV particles having unique capsids. It is our idea that those capsids will combine the low reactivity with human sera as it is typical for non-primate AAVs, with the high efficiency of in vivo transduction as reported for primate/human AAVs. This AAV library will then be mixed with antibodies raised against primate/human AAV serotypes and used to transduce mouse liver in vivo. It is hoped that this form of selective pressure will enable us to isolate AAV capsid sequences allowing efficient in vivo transduction while evading neutralization from pre-existing immunity. We plan to test these new vectors in animal models of hemophilia. Our ultimate goal is to utilize any sequences that we can identify for development of novel improved AAV vectors for use in clinical trials.

我们已经证明，AAV-2载体可以在体内转导人肝细胞。然而，由于预先存在的体液免疫力，AAV介导的基因转移受到限制，并且由于针对含有含肝细胞的衣壳的免疫反应，因此短暂的转基因表达是短暂的。该项目的目的是基于腺相关病毒（AAV）开发一种新型的人类基因转移载体（AAV），结合了靶组织体内转导的高效率，并能够规避或克服人类中的宿主免疫反应，包括人类中和人群中中和抗体的存在，可能会限制其效率。这些新颖的矢量将通过DNA的“分子进化”来得出，从一系列天然存在的AAV血清型中。该阵列将包含先前报道的人类和灵长类动物的AAV变体，以及新的非灵活性AAV，它们的隔离是我们的主要特定目标。各种AAV分离株的基因组将被“洗牌”以生成一个DNA池，然后我们将产生一个具有独特的衣壳的重组AAV颗粒库。我们的想法是，这些衣壳将低反应性与人类血清相结合，因为它是非成就AAV的典型代表，而在灵长类动物/人类AAV的报道中，体内转导的效率很高。然后，该AAV库将与针对灵长类动物/人类AAV血清型提出的抗体混合，并用于在体内转导小鼠肝脏。希望这种选择性压力能够使我们能够隔离AAV capsid序列，从而使体内转导有效，同时避免免于预先存在的免疫力中和。我们计划在血友病动物模型中测试这些新媒介。我们的最终目标 是利用我们可以识别出的任何序列来开发新的改进的AAV矢量，以用于临床试验。