Engineering AAV Vectors to Evade Antibody Neutralization

设计 AAV 载体以逃避抗体中和

• 批准号: 7442123
• 负责人: DAVID V SCHAFFER
• 金额: $ 36.12万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-06-08 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7442123
• 关键词: Address; Animals; Antibodies; Biology; CD8B1 gene; Capsid; Capsid Proteins; Clinic; Complement; Dependovirus; Disease; Engineering; Erythropoietin; Gene Delivery; Genes; Human; Immune; Immune response; Immune system; Immunology; In Vitro; Individual; Laboratories; Libraries; Liver; Mediating; Medicine; Molecular; Molecular Biology; Muscle; Personal Satisfaction; Phenotype; Point Mutation; Population; Property; Resistance; Role; Safety; Serotyping; Serum; T-Lymphocyte; Technology; Therapeutic; Variant; Virus; Work; adeno-associated viral vector; base; cellular transduction; directed evolution; gene therapy; high throughput screening; in vivo; insight; mutant; novel; novel strategies; receptor binding; response; therapeutic gene; vector; viral gene delivery; virology

DESCRIPTION (provided by applicant): Gene therapy has vast potential for treating and potentially curing a wide variety of disorders. However, gene delivery technologies require significant improvements in safety, efficiency, and expression stability before the majority of these diseases can be treated. Vectors based on adeno-associated virus (AAV) have proven themselves to be highly promising, both in the laboratory and the clinic, but they still suffer from several shortcomings. In particular, the majority of the human population has been exposed to AAV serotype 2, as well as other serotypes, and as a result the immune system is primed to neutralize AAV. Antibody neutralization of AAV vectors is an established problem, and cellular immune responses may also be a challenge. We will attempt to solve the former problem and will further investigate basic mechanisms involved in the latter. For the former, we have developed novel directed evolution technology to generate new mutants of AAV with new properties. Specifically, large libraries of virus with random point mutations in the capsid gene encoding the viral coat protein are generated, and variants with novel properties are selected using high throughput screens. We have utilized this approach to generate variants with altered receptor binding properties, as well as variants that escape neutralization by antibodies that greatly inhibit AAV gene delivery by the wild type capsid or coat proteins. We will study the potential of human antibody evading variants to mediate high efficiency gene delivery of the therapeutic gene erythropoietin to the muscle and liver of animals carrying anti-AAV antibodies. In addition, while AAV neutralization by antibodies is an established problem, much less is known about AAV interactions with other components of the immune system. Therefore, the basic mechanisms of immune neutralization of this virus by complement [and T cells] will be investigated to both in vitro and in vivo. In summary, viruses have naturally evolved for their own ends, which do not always meet the needs of a human therapeutic. The novel approaches developed in this work to re-evolve viruses into enhanced human therapeutics will therefore have broad and general impact on the molecular engineering of enhanced viral gene delivery vehicles, including alternate AAV serotypes as well as other vectors. Furthermore, it will yield insights into the responses of other immune system components to AAV.

描述（由申请人提供）：基因治疗具有治疗和潜在治疗多种疾病的巨大潜力。但是，在可以治疗大多数疾病之前，基因递送技术需要显着改善安全性，效率和表达稳定性。在实验室和诊所中，基于腺相关病毒（AAV）基于腺相关病毒（AAV）的载体已经非常有前途，但它们仍然遭受了几个缺点。特别是，大多数人口已暴露于AAV血清型2以及其他血清型，因此免疫系统被启动以中和AAV。 AAV载体的抗体中和是一个已建立的问题，细胞免疫反应也可能是一个挑战。我们将尝试解决以前的问题，并将进一步研究涉及后者的基本机制。对于前者，我们开发了新颖的定向进化技术，以生成具有新属性的AAV的新突变体。具体而言，在编码病毒外套蛋白的衣壳基因中具有随机点突变的病毒库，并产生具有新型特性的变体。我们已经利用这种方法来生成具有改变受体结合特性的变体，以及通过抗体中和的变体，这些抗体极大地抑制了野生型衣壳或外套蛋白的AAV基因递送。我们将研究人类抗体逃避变体的潜力，以介导携带抗AAV抗体的动物的肌肉和肝脏的治疗基因促红细胞生成素的高效率基因递送。另外，尽管抗体中和是一个已建立的问题，但对于与免疫系统的其他成分的AAV相互作用知之甚少。因此，通过补体[和T细胞]对该病毒免疫中和的基本机制将被研究到体外和体内。总而言之，病毒自然而然地出于自己的目的而发展，这并不总是满足人类治疗的需求。因此，在这项工作中开发的新方法将病毒重新进化为增强的人类治疗剂将对增强的病毒基因输送车的分子工程产生广泛而普遍的影响，包括替代AAV血清型以及其他载体。此外，它将对其他免疫系统组件对AAV的反应产生见解。