GENE THERAPY FOR IMMUNE DEFICIENCIES

免疫缺陷的基因治疗

• 批准号: 3768877
• 负责人: H L MALECH
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3768877
• 关键词: NAD(P)H dehydrogenase; Retroviridae; adolescence (12-20); autosome; cell differentiation; child (0-11); chronic granulomatous disease; clinical trials; complementary DNA; cytochrome b; disease /disorder model; gene therapy; genetic transduction; human subject; human therapy evaluation; leukocyte oxidative burst; lymphocyte proliferation; monocyte; myeloid stem cell; neutrophil; sex linked trait; tissue /cell culture; transfection /expression vector

This project studies gene therapy for chronic granulomatous diseases (CGD) and other inherited immune diseases affecting human phagocytes. CGD are a group of 4 distinct genetic disorders with common phenotype characterized by life-threatening recurrent infections caused by failure of blood neutrophils and monocytes to produce superoxide and hydrogen peroxide. CGD results from the failure to produce any of the components of the NADPH oxidase required for superoxide generation. We have now successfully used retrovirus mediated gene transfer techniques to functionally correct 3 forms of CGD, the autosomal p47phox deficient CGD and the X-linked and autosomal forms of CGD involving abnormalities of flavocytochrome b558 subunits gp91phox and p22phox. We have developed two in vitro model systems to demonstrate the feasibility of genetic correction of CGD. The first system uses Epstein-Barr virus transformed B-lymphocyte cell lines (EBV-BL). EBV-BL from normal individuals produce small amounts of superoxide when stimulated, while those derived from CGD patients do not produce superoxide and are missing the oxidase component characteristic of that genetic form of CGD. The retrovirus vector, MFG, which produces high viral titers and higher protein production was used to transduce p47phox, gp91phox and p22phox cDNAs into EBV-BL to correct the CGD defect routinely obtaining a 1 to 15% correction rate without selection for transduced cells. This trait was stable for months of subsequent culture. Correction was specific to transduction of the defective component cDNA. In the second model system, marrow-like myeloid progenitor cells harvested from the peripheral blood could be induced to proliferate and differentiate in vitro to mature neutrophils capable of superoxide production. Similar cells from CGD patients could not produce superoxide, but transduction of these precursors from CGD patients with the MFG retrovirus containing the normal version of the specific CGD gene resulted in correction of superoxide production in 4-20% of resultant neutrophils. Theoretically, the peripheral blood progenitor model system of gene transfer could be scaled up to a procedure capable of being used for in vivo gene therapy for immune system and other disorders affecting bone marrow cells.

该项目研究慢性肉芽肿性疾病的基因治疗 （CGD）和其他影响人吞噬细胞的遗传免疫疾病。 CGD是一组常见表型的4种不同的遗传疾病 由失败引起的威胁生命的复发感染的特征 血液中性粒细胞和单核细胞产生超氧化物和氢 过氧化物。 CGD因未能产生任何组件而导致 超氧化物产生所需的NADPH氧化酶的数量。 我们现在有 成功使用逆转录病毒介导的基因转移技术 在功能上正确正确的CGD形式，常染色体P47PHOX缺陷CGD 以及CGD的X连锁和常染色体形式，涉及异常 黄素色素B558亚基GP91Phox和p22phox。 我们已经发展了 两个体外模型系统，以证明遗传的可行性 校正CGD。 第一个系统使用Epstein-Barr病毒转化 B-淋巴细胞细胞系（EBV-BL）。 来自正常人的EBV-BL会产生 刺激时少量的超氧化物，而这些超氧 CGD患者不产生超氧化物，并且缺少氧化酶 CGD的遗传形式的成分特征。 逆转录病毒 载体，MFG，产生高病毒滴度和较高蛋白质 生产用于将P47Phox，GP91Phox和P22Phox cDNA传递到 EBV-bl以纠正CGD缺陷通常获得1-15％ 校正率无用于转导细胞的选择。 这个特征是 稳定在几个月后的文化中。 更正是特定的 有缺陷的成分cDNA的转导。 在第二个模型中 系统，骨髓状的髓样祖细胞从 可以诱导外周血增殖和区分 体外到能够产生超氧化物的成熟中性粒细胞。 相似的 来自CGD患者的细胞不能产生超氧化物，而是转导的细胞 来自含MFG逆转录病毒的CGD患者的这些前体 特定CGD基因的正常版本导致校正 4-20％的中性粒细胞中的超氧化物产生。从理论上讲， 基因转移的外周血祖细胞模型系统可能是 扩展到能够用于体内基因治疗的过程 用于免疫系统和其他影响骨髓细胞的疾病。