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连锁的慢性肉芽肿性疾病（X-CGD）源于该缺陷 编码gp91phox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 gp91phox。 这一目标的基础的中心假设是 逆转录病毒介导的GP91Phox cDNA转移到X-CGD 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 拟议的研究计划将使用逆转录病毒载体使用 他人显示的设计以赋予体内长期表达 转移的基因序列。 X-CGD的鼠标模型 基因靶向最近开发的将用于检查 逆转录病毒介导的GP91PHOX表达和呼吸爆发氧化酶 在体内功能并评估基因替代疗法的影响 在X-CGD表型上。 有希望的向量也将对他们的 能够在人X-CGD中赋予gp91phox的功能表达 造血干和祖细胞。 除了骨髓 细胞，从外周血分离的造血前体将是 评估为基因转移的靶标，并作为离体的候选者评估 转导之前的扩展。 为高效而开发的协议 如有必要，将修改骨髓细胞的转导 外周血细胞靶标。 除了体外菌落测定外， 与绵羊异种移植模型相比，人摩尔将用于评估转导的 靶细胞长期，体内，骨髓填充能力 重组GP91Phox在成熟中的功能表达 吞噬白细胞。 该子标记中概述的工作应有助于 在使用逆转录病毒介导的基因的临床方案开发中 作为X-CGD和其他单基因缺陷中的治疗策略转移 造血干细胞。 更广泛地，这些研究应该增加 了解如何将特定遗传修饰引入 造血干细胞，同时保持自我更新和 多能性。