Recombinant adeno-associated virus-mediated transgene delivery into porcine organs to allow immunological modifications of xenografts

重组腺相关病毒介导的转基因递送至猪器官以允许异种移植物的免疫学修饰

• 批准号: 5424673
• 负责人: Professor Dr. Michael Hallek
• 金额: --
• 依托单位: Hämatologikum der GSF (aufgelöst)
• 依托单位国家: 德国
• 项目类别: Research Units
• 财政年份: 2004
• 资助国家: 德国
• 起止时间: 2003-12-31 至 2007-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/5424673?language=en
• 关键词: Recombinant; adeno; associated; virus; mediated

The breeding of transgenic pigs brings the realization of xenotransplantation closer. However, the generation of transgenic pigs expressing multiple transgenes is time-consuming and expensive. Adeno-associated virus (AAV) can serve as an efficient and safe vector system for gene transfer into donor organs. AAV is characterized by low immunogenicity and allows long-term gene expression. Our research group is experienced in high titer production and purification of AAV. Targeting strategies have been developed that enhance the specificity of gene transfer towards distinct types of cells such as endothelial cells. A newly developed self-complementary AAV construct allows very rapid and elevated transgene expression. Eight AAV serotypes, which show different tissue and species specificities, have been characterized. On this background AAV vectors will be constructed that are optimized towards efficient gene transfer into porcine tissues. This will involve the testing of different AAV serotypes on porcine cells in vitro and the development of a targeting AAV vector specially tailored for an efficient gene transfer into porcine endothelial and heart muscle cells. Furthermore, a self complementary AAV coding the immunomodulatory factors CTLA4-Ig and interleukin-10 will be produced and used in a allogenic mouse heart transplantation model. The optimized AAV vector will finally be used for gene transfer into porcine hearts in vivo. This technique will allow further modifications of organs derived from transgenic animals by introducing additional immunomodulatory genes.

转基因猪的培育使异种器官移植的实现更近了一步。然而，产生表达多种转基因的转基因猪是耗时且昂贵的。腺相关病毒（AAV）可作为一种高效、安全的基因转移载体系统。AAV的特征在于低免疫原性，并允许长期基因表达。我们的研究小组在AAV的高效价生产和纯化方面经验丰富。已经开发了靶向策略，其增强了基因转移对不同类型的细胞如内皮细胞的特异性。新开发的自身互补AAV构建体允许非常快速和升高的转基因表达。八种AAV血清型，显示不同的组织和物种特异性，已被表征。在此背景下，将构建针对有效基因转移到猪组织中而优化的AAV载体。这将涉及在体外对猪细胞进行不同AAV血清型的测试，以及开发专门针对猪内皮细胞和心肌细胞的有效基因转移而定制的靶向AAV载体。此外，将产生编码免疫调节因子CTLA 4-IG和白细胞介素-10的自身互补AAV，并将其用于同种异体小鼠心脏移植模型。优化的AAV载体将最终用于体内基因转移到猪心脏中。该技术将允许通过引入额外的免疫调节基因来进一步修饰来自转基因动物的器官。