HUMAN CTL RESPONSES TO ADENOVIRUS GENE THERAPY VECTORS

人类 CTL 对腺病毒基因治疗载体的反应

• 批准号: 2791351
• 负责人: PHYLLIS Rudolph FLOMENBERG
• 金额: $ 18.62万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-01-15 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2791351
• 关键词: Adenoviridae; MHC class I antigen; cytotoxic T lymphocyte; fibroblasts; gene deletion mutation; human genetic material tag; human tissue; immediate early protein; immunogenetics; microorganism culture; microorganism immunology; mixed tissue /cell culture; regulatory gene; transfection /expression vector; tumor necrosis factor alpha; virus genetics; virus infection mechanism; virus protein

The goal of this proposal is to define the mechanisms involved in the generation of human adenovirus-specific cytotoxic T lymphocytes (CTLs) in order to help design new strategies for evading the immune response to adenovirus gene therapy vectors. Adenoviruses are under extensive investigation as gene therapy vectors for a broad spectrum of heart, lung, and blood diseases including cystic fibrosis, hemophilia, and atherosclerosis. However, data from animal models indicate that the immunogenicity of adenovirus vectors interferes with the efficacy of adenovirus-mediated gene therapy. Administration of early region 1(E1)-deleted adenovirus vectors to mice, a host in which adenovirus infection is naturally restricted, results in the generation of adenovirus-specific CTLs which destroy adenovirus-transduced cells within a few weeks. Further analysis of this problem requires study of human CTL responses against adenovirus. We have successfully amplified memory adenovirus-specific CTLs in peripheral blood mononuclear cells from healthy adults and documented that these responses are major histocompatiblity complex (MHC)-restricted and mediated by CD8+ T cells. Based on our studies, it is likely that the presence of memory cellular immune responses will pose a major additional obstacle for adenovirus-mediated gene therapy in man. We postulate, however, that human adenovirus-specific CTLs may be targeted against a limited number of immunodominant epitopes. Therefore, it may be possible to reduce the immunogenicity of adenovirus vectors by elimination of such epitopes. As a second approach, it may be possible to take advantage of mechanisms which adenovirus has developed to evade host immune responses. The Ad early region 3 (E3) codes for proteins which help make cells resistant to CTLs and tumor necrosis factor, but this region is deleted or poorly expressed from most adenovirus vectors. We postulate that constitutive expression of E3 region proteins may help reduce the immunogenicity of adenovirus vectors. We propose to address these hypotheses by analysis of human CTL responses against adenovirus in vitro. These studies will provide an experimental basis for the design of more effective adenovirus gene therapy vectors for future human trials.

该提案的目标是定义参与产生人腺病毒特异性细胞毒性T淋巴细胞（CTL）的机制，以帮助设计新的策略，以逃避腺病毒基因治疗载体的免疫应答。腺病毒作为广泛的心脏、肺和血液疾病（包括囊性纤维化、血友病和动脉粥样硬化）的基因治疗载体正在进行广泛的研究。然而，来自动物模型的数据表明腺病毒载体的免疫原性干扰腺病毒介导的基因治疗的功效。 将早期区域1（E1）缺失的腺病毒载体给予小鼠（其中腺病毒感染天然受限的宿主）导致产生腺病毒特异性CTL，其在几周内破坏腺病毒转导的细胞。 对这个问题的进一步分析需要研究针对腺病毒的人CTL应答。 我们已经成功地扩增了记忆腺病毒特异性CTL在外周血单核细胞从健康成人和记录，这些反应是主要组织相容性复合体（MHC）限制和介导的CD8+ T细胞。 根据我们的研究，记忆细胞免疫反应的存在可能会对腺病毒介导的基因治疗造成额外的障碍。然而，我们假设，人类腺病毒特异性CTL可能针对有限数量的免疫显性表位。因此，有可能通过消除这些表位来降低腺病毒载体的免疫原性。 作为第二种方法，可以利用腺病毒已经开发的逃避宿主免疫应答的机制。Ad早期区域3（E3）编码有助于使细胞对CTL和肿瘤坏死因子具有抗性的蛋白质，但该区域在大多数腺病毒载体中缺失或表达不足。 我们推测E3区蛋白的组成型表达可能有助于降低腺病毒载体的免疫原性。 我们建议解决这些假设，通过分析在体外对腺病毒的人CTL反应。 这些研究将为今后设计更有效的腺病毒基因治疗载体进行人体试验提供实验基础。