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: 6879596
• 负责人: Mary C Dinauer
• 金额: $ 40.27万
• 依托单位: INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-02-29 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6879596
• 关键词: NAD(P)H dehydrogenase; Retroviridae; biotechnology; cell line; chronic granulomatous disease; disease /disorder model; gene therapy; genetic transduction; green fluorescent proteins; hematopoietic stem cells; hematopoietic tissue transplantation; human genetic material tag; laboratory mouse; nonhuman therapy evaluation; recombinant proteins; sex linked trait; stem cell transplantation; transfection /expression vector

X-linked chronic granulomatous disease (X-CGD) arises from defects in the gene encoding gp91phox, 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 gp91phox. The central hypothesis underlying this objective is that retroviral-mediated gene transfer of gp91phox 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 will be prepared utilizing designs shown by others to confer long-term expression in vivo of transferred gene sequences. A mouse model of X-CGD that has been recently developed by gene targeting will be utilized to examine retroviral-mediated gp91phox expression and respiratory burst oxidase function in vivo and to evaluate the impact of gene replacement therapy on the X-CGD phenotype. Promising vectors will also be tested for their ability to confer functional expression of gp91phox 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 for ex vivo expansion prior to transduction. Protocols developed for efficient transduction of marrow cells will be modified, if necessary, for peripheral blood cell targets. In addition to in vitro colony assays, a human-more than 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 gp91phox 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连锁慢性肉芽肿病（X-CGD）是由细胞缺陷引起的 编码 gp91phox 的基因，吞噬细胞特异性细胞色素 b 的一个亚基 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出现复发性、严重的 感染始于儿童早期。 拟议的目标 研究目的是为基因替代建立实验基础 使用复制缺陷型逆转录病毒进行表达的 X-CGD 治疗 gp91phox。 这一目标的核心假设是 逆转录病毒介导的 gp91phox cDNA 基因转移至 X-CGD 造血干细胞将恢复呼吸爆发活性 成熟的吞噬白细胞并纠正宿主防御的缺陷。 在 拟议的研究计划，将利用逆转录病毒载体来制备 其他人展示的设计可在体内长期表达 转移的基因序列。 X-CGD 小鼠模型 最近开发的基因打靶将用于检查 逆转录病毒介导的 gp91phox 表达和呼吸爆发氧化酶 体内功能并评估基因替代疗法的影响 关于 X-CGD 表型。 有前途的载体也将接受测试 在人类 X-CGD 中赋予 gp91phox 功能表达的能力 造血干细胞和祖细胞。 除了骨髓 从外周血中分离出的细胞、造血前体细胞将 被评估为基因转移的靶标以及离体的候选者 转导前的扩增。 为高效而开发的协议 如有必要，将修改骨髓细胞的转导 外周血细胞目标。 除了体外菌落分析之外， 人类-超过绵羊的异种移植模型将用于评估转导 具有长期体内骨髓增殖能力的靶细胞 以及重组gp91phox在成熟细胞中的功能表达 吞噬性白细胞。 该子项目中概述的工作应该有助于 使用逆转录病毒介导的基因制定临床方案 转移作为 X-CGD 和其他单基因缺陷的治疗策略 造血干细胞。 更广泛地说，这些研究应该补充 了解如何将特定的基因修饰引入 造血干细胞同时保持自我更新和 多重潜能。