GENE TRANSFER FOR SPECIFIC IMMUNOTHERAPY OF MYASTHENIA

用于肌无力特异性免疫治疗的基因转移

• 批准号: 6702280
• 负责人: DANIEL B DRACHMAN
• 金额: $ 35.42万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-02-09 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6702280
• 关键词: T cell receptor; acetylcholine; antigen antibody reaction; antigen presenting cell; apoptosis; autoantigens; biotechnology; complementary DNA; disease /disorder model; gene expression; gene targeting; genetically modified animals; hemagglutinin; immunotherapy; laboratory mouse; laboratory rat; method development; model design /development; myasthenia gravis; neurotransmitter receptor; plasmids; tissue /cell culture; transfection; transfection /expression vector; vaccinia virus

DESCRIPTION (adapted from applicant's abstract): This is a proposal to develop and apply a unique and powerful new gene transfer strategy for specific immunotherapy of myasthenia gravis (MG). Ideally, therapy of MG should specifically inhibit the autoimmune response to the autoantigen, acetylcholine receptor (AChR), without otherwise interfering with the immune system. Because the antibody response to AChR is T cell dependant, elimination of the AChR-specific T cells interrupts the autoimmune disorder at a pivotal point, resulting in clinical benefit. This requires elimination of virtually all AChR-specific T cells. However, the marked heterogeneity of T cell responses to AChR in humans and in experimental animals presents a challenge in designing specific treatment capable of eliminating the entire repertoire of AChR-specific T cells. The investigator's targeting strategy is based on the fact that a myasthenic individual's own antigen presenting cells (APCs) can present AChR epitopes to that indivudual's entire repertoire of AChR specific T cells. They have adapted a method that induces the APCs to process and present AChR by inserting a cDNA construct that encodes the key immunogenic domain of the AChR flanked by signals that induce the APCs process and present it. These APCs target AChR-specific T cells highly effectively. In order to eliminate the targeted T cells, they utilize Fas ligands as a "warhead." When FasL interacts with Fas, which is abundantly expressed on activated T cells, it induces apoptosis and death of the T cells. Insertion of the gene for FasL in APCs induces them to express FasL, which effectively kills Fas-expressing target cells. It is essential to protect these APCs, which would die by FasL- induced "suicide," because they also express Fas. To protect these APCs, they insert a 3rd gene for a truncated form of FADD, which acts as a dominant negative inhibitor of Fas-mediated cell death, into the APCs. To insert all 3 genes simultaneously into APCs ex vivo, they have developed a vaccinia virus vector (vvv), which carries all 3 genes. This vaccinia vector can transfer multiple genes simultaneously, inducing high level production of the gene encoded proteins. When it is attenuated (by treatment with psoralen and UV light) the vvv does not replicate but does direct expression of the gene products. Finally, the expression of FasL directed by their vvv system should confer protection against an immune attack by the host's immune system on the virus vector itself. They now propose to optimize the system and develop it in two animal models: (1) experimental MG, including a new transgenic myasthenic mouse model; and (2) a transgenic mouse model of hemaglutinin (HA) sensitivity. The experiments described in the present proposal will develop this novel strategy in experimental animals, both for prevention and for treatment of EAMG. This method is designed to be suitable for treatment of human autoimmune diseases. When optimized in experimental animals, it will be adaptable for treatment of patients with MG and other autoimmune diseases.

描述（改编自申请人的摘要）：这是开发的建议 并应用独特而强大的新基因转移策略 肌无力的重症疗法（MG）。理想情况下，MG的治疗应 特别抑制对自身抗原，乙酰胆碱的自身免疫反应 受体（ACHR），没有其他干扰免疫系统。因为 对ACHR的抗体反应是T细胞依赖性的，消除了 ACHR特异性T细胞中断了关键点的自身免疫性疾病， 导致临床益处。这几乎需要消除所有 ACHR特异性T细胞。但是，T细胞反应的明显异质性 人类和实验动物中的ACHR提出了设计的挑战 能够消除整个曲目的特定处理 ACHR特异性T细胞。研究者的定位策略是基于 肌无力的人自己的抗原呈现细胞（APC）可以 当前ACHR的表现是该独立的ACHR特定t的整个曲目 细胞。他们已经改编了一种诱导APC进行处理并存在的方法 通过插入编码密钥免疫原性的cDNA构建体来ACHR ACHR的侧面是诱导APC过程并提出它的信号。这些 APC靶向ACHR特异性T细胞高度有效。为了消除 靶向T细胞，他们利用FAS配体作为“弹头”。当Fasl相互作用时 使用在活化的T细胞上大量表达的FAS，它诱导 T细胞的凋亡和死亡。在APC中插入FASL基因 诱导他们表达FASL，这有效地杀死了表达FAS的目标 细胞。保护这些APC至关重要，这些APC会因FASL诱导而死 “自杀”，因为他们也表达了FAS。为了保护这些APC，它们插入 FADD的截断形式的第三基因，该基因充当主要负面的基因 Fas介导的细胞死亡的抑制剂，进入APC。插入所有3个基因 同时，他们在apcs ex vivo中开发了疫苗病毒载体 （VVV），它带有所有3个基因。该离子载体可以转移多个 基因同时诱导了编码基因的高水平产生 蛋白质。当它被衰减时（通过用牛cor和紫外线处理） VVV不复制，而是直接表达基因产物。 最后，通过其VVV系统指导的FASL的表达应授予 防止宿主的免疫系统对病毒的免疫攻击 向量本身。 他们现在建议优化系统并在两个动物模型中开发它： （1）实验MG，包括新的转基因肌关系小鼠模型； （2） 血谷蛋白（HA）敏感性的转基因小鼠模型。实验 本提案中描述的将在 实验动物，用于预防和治疗EAMG。这 方法旨在适合治疗人自身免疫性疾病。 在实验动物中进行优化时，它将适应 MG和其他自身免疫性疾病的患者。