Tropism-modified AAV vectors for neonatal gene transfer

用于新生儿基因转移的趋向性修饰 AAV 载体

• 批准号: 6620368
• 负责人: THOMAS J DALY
• 金额: $ 12.52万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-01 至 2003-10-10
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6620368
• 关键词: adeno associated virus group; bacterial virus; biotechnology; enzyme deficiency; fluorimetry; gene expression; gene therapy; genetic library; genetic manipulation; genetic screening; genetic transduction; histochemistry /cytochemistry; human genetic material tag; inborn metabolism disorder; laboratory mouse; liver; newborn animals; postdoctoral investigator; recombinant virus; southern blotting; tissue /cell culture; transfection /expression vector; virus infection mechanism; virus receptors

The amino acid disorders are a family of diseases characterized by genetic defects in a primary metabolic pathway. For most of these syndromes, current treatment is limited to supportive care for acute episodes, and dietary restriction to avoid utilization of the defective metabolic pathway. The long-term success of these strategies varies greatly between diseases. Because many of these disorders are caused by single gene defects in hepatic enzymes, they are appealing targets for neonatal gene transfer strategies directed at the liver. Adeno-associated virus (AAV) has emerged as a promising vector that can achieve long- term hepatic expression in animal models. However, its clinical potential for neonatal gene transfer to the liver is limited by non- specific transduction of multiple organs in vivo, and by the dilution of transduced cells by the rapid growth of the newborn liver following treatment. Improvements in the efficiency of AAV targeting to the liver could increase the effective dose of vector delivered to the liver, while decreasing unwanted extrahepatic transduction. In this proposal we hypothesize that the binding properties of AAV can be modified to meet these goals. We will create bispecific targeting constructs directed at liver-specific receptors, and use phage library display technology to seek neonatal liver-specific receptors which may be targeted. We will then examine the effects of these modifications on AAV transduction of hepatic and non-hepatic tissues in vitro. Finally, we will examine changes in the biodistibution and persistence of expression following intravenous injection of AAV into newborn mice. Improvements in AAV tropism for the neonatal liver will provide a tool which will be widely applicable to a range of pediatric metabolic diseases that primarily affect the liver, including the amino acid disorders. In addition, the skills and interaction provided in this proposal will be an invaluable aid in the transition from post-doctoral research to a career as an independent investigator.

氨基酸紊乱是一类以主要代谢途径中的遗传缺陷为特征的疾病。对于这些综合征中的大多数，目前的治疗仅限于对急性发作的支持性护理，以及限制饮食以避免利用有缺陷的代谢途径。这些策略的长期成功程度因疾病而异。由于这些疾病中的许多是由肝酶中的单基因缺陷引起的，它们是针对肝脏的新生儿基因转移策略的吸引目标。腺相关病毒(AAV)是一种很有前途的载体，可以在动物模型中实现肝脏的长期表达。然而，其新生儿基因转移到肝脏的临床潜力受到体内多个器官的非特异性转导的限制，以及治疗后新生肝脏的快速生长对转导细胞的稀释。靶向肝脏的AAV效率的提高可以增加向肝脏输送载体的有效剂量，同时减少不必要的肝外转导。在这项提议中，我们假设AAV的结合属性可以被修改以满足这些目标。我们将创建针对肝脏特异性受体的双特异性靶向构建体，并利用噬菌体文库展示技术寻找可能靶向的新生儿肝脏特异性受体。然后，我们将研究这些修饰对体外肝和非肝组织AAV转导的影响。最后，我们将检查AAV静脉注射到新生小鼠后，表达的生物分布和持久性的变化。改善AAV对新生儿肝脏的趋向性将提供一种工具，将广泛适用于一系列主要影响肝脏的儿科代谢性疾病，包括氨基酸紊乱。此外，这项建议中提供的技能和互动将是从博士后研究过渡到独立研究人员职业的宝贵帮助。