GENE THERAPY OF X-LINKED CHRONIC GRANULOMATOUS DISEASE

X连锁慢性肉芽肿性疾病的基因治疗

• 批准号: 6105676
• 负责人: Mary C Dinauer
• 金额: $ 12.08万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-15 至 1999-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6105676
• 关键词: Retroviridae; biomarker; bone marrow; chronic granulomatous disease; gene expression; gene therapy; genetic transduction; hematopoietic stem cells; hematopoietic tissue transplantation; human tissue; polymerase chain reaction; recombinant proteins; tissue /cell culture; tissue mosaicism; transfection /expression vector; xenotransplantation; Retroviridae; biomarker; bone marrow; chronic granulomatous disease; gene expression; gene therapy; genetic transduction; hematopoietic stem cells; hematopoietic tissue transplantation; human tissue; polymerase chain reaction; recombinant proteins; tissue /cell culture; tissue mosaicism; transfection /expression vector; xenotransplantation

X-linked chronic granulomatous disease (X-CGD) arises from defects in the gene encoding gp91/phox, a subunit of a phagocyte-specific cytochrome b that is essential for respiratory burst oxidase function. Affected patients lack a major antimicrobial pathway and develop recurrent, severe infections beginning in early childhood. The objective of the proposed research is to establish an experimental basis for gene replacement therapy of X-CGD using replication-defective retroviruses for expression of gp91/phox. The central hypothesis underlying this objective is that retroviral-mediated gene transfer of gp91/phox cDNA into X-CGD hematopoietic stem cells will restore respiratory burst activity in mature phagocytic leukocytes and correct the defect in host defense. In the proposed research plan, retroviral vectors containing the gp91 cDNA will be prepared utilizing designs previously shown by others to confer long-term expression in vivo of transferred gene sequences. These vectors will be tested for their ability to confer functional expression of gp91/phox in human X-CGD hematopoietic stem and progenitor cells. In addition to bone marrow cells, hematopoietic precursors isolated from peripheral blood will be evaluated as targets for gene transfer and also as candidates of ex vivo expansion prior to transduction. Protocols developed for efficient transduction of marrow cells will be modified, if necessary, for peripheral blood cell targets. Gene transfer and expression will be first investigated using in vitro clonogenic assays of stem and progenitor cells. A human- sheep xenograft model will be used to assess transduced target cells for long-term, in vivo, bone marrow populating capabilities and the functional expression of recombinant gp91/phox in mature phagocytic leukocytes. The work outlined in this subproject should aid in the development of clinical protocols using retroviral-mediated gene transfer as a therapeutic strategy in X- CGD and other single-gene defects of hematopoietic stem cells. More broadly, these studies should add to knowledge of how to introduce specific genetic modifications into hematopoietic stem cells while maintaining self-renewal and multipotentiality.

X连锁的慢性肉芽肿性疾病（X-CGD）源于该缺陷 编码gp91/phox的基因，吞噬细胞特异性细胞色素b的亚基 这对于呼吸爆发氧化酶功能至关重要。 做作的 患者缺乏主要的抗菌途径，并且会产生重复的，严重的 感染从幼儿开始。 提议的目标 研究是为基因置换的实验基础 使用复制缺陷逆转录病毒对X-CGD进行治疗以表达 gp91/phox的 这一目标的基础的中心假设是 逆转录病毒介导的基因转移GP91/Phox cDNA到X-CGD中 造血干细胞将恢复呼吸道爆发活性 成熟的吞噬白细胞并纠正宿主防御中的缺陷。 在 拟议的研究计划，含有GP91 cDNA的逆转录病毒载体 将准备利用其他人以前显示的设计进行准备 转移基因序列的体内长期表达。 这些 向量将通过其赋予功能表达的能力进行测试 人X-CGD造血干和祖细胞中GP91/PHOX的of。 在 除了骨髓细胞外，从中分离出的造血前体 外周血将作为基因转移的靶标评估，也将 作为转导之前的体内扩张的候选者。 协议 开发用于有效转导骨髓细胞的开发，将修改 如有必要，对于外周血细胞靶标。 基因转移和 表达将首先使用体外克隆分析进行研究 茎和祖细胞。 人类绵羊异种移植模型将是 用于评估长期体内骨的转导靶细胞 骨髓填充功能和功能表达 重组GP91/Phox在成熟的吞噬白细胞中。 工作概述了 在本次审计中，应有助于制定临床方案 使用逆转录病毒介导的基因转移作为X-的治疗策略 CGD和造血干细胞的其他单基因缺陷。 更多的 从广义上讲，这些研究应该增加有关如何介绍的知识 特定的遗传修饰为造血干细胞，而 保持自我更新和多能性。