GENE TRANSFER FOR THERAPY OF IL 2 RECEPTOR GAMMA CHAIN DEFICIENT X LINKED SCID

用于治疗 IL 2 受体伽玛链缺陷型 X 连锁 SCID 的基因转移

• 批准号: 2576539
• 负责人: J M PUCK
• 金额: --
• 依托单位: NATIONAL HUMAN GENOME RESEARCH INSTITUTE
• 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/2576539
• 关键词: SCID mouse; cytokine receptors; dogs; gene expression; gene mutation; gene therapy; hematopoietic stem cells; human genetic material tag; human tissue; immunofluorescence technique; immunopathology; interleukin 2; newborn human (0-6 weeks); nonhuman therapy evaluation; severe combined immunodeficiency; sex linked trait; tissue /cell culture; transfection /expression vector; umbilical cord

The gamma chain gene of the IL-2 receptor (IL2RG) has been identified as responsible for X-linked severe combined immunodeficiency (SCID). This is the most common form of human SCID. The only available lifesaving treatment at the present time is bone marrow transplantation, but drawbacks of this procedure include the lack of an HLA matched donor in over 2/3 of cases and poor post-transplant B cell function, primarily due to non-engraftment of B cells. The previously proven negative selection against cells expressing a mutant IL2RG, resulting in skewed X chromosome inactivation in female carriers, makes X-linked SCID a promising candidate for therapy with retrovirally transduced autologous repopulating stem cells. This strategy is being pursued first by comparing different retrovirus constructs for titer, gamma chain expression in transduced cell lines, correction of the functional defect in EBV-B cell lines from SCID-affected patients with defined IL2RG defects, and monitoring the development of cord blood stem cells transduced with IL2RG retroviruses in SCID mice. These experiments have shown significant cell surface expression of the gamma chain protein by immunofluorescence in previously deficient cell lines from SCID patients. The availability of a large panel of patient cell lines will permit evaluation of potential dominant negative mutations. SCID patients prenatally diagnosed with defined IL2RG mutations have had their cord blood collected at birth for experimental transduction and evaluation of lymphocytogenesis. Canine spontaneous gamma chain mutations resulting in SCID dogs are being studied in collaboration with Paula Henthorn, at the U. of PA School of Vet. Med. A marking experiment is in progress in which two retroviral constructs were introduced into bone marrow of an unaffected dog pretreated with cytokines. The marrow was infused into a MHC-matched irradiated recipient, who at 8 weeks demonstrates substantial blood and bone marrow expression of human gamma chain. Follow-up experiments in SCID dogs are planned.

IL-2受体的γ链基因（IL2RG）已被鉴定为 X连锁严重联合免疫缺陷（SCID）。这 是人类严重联合免疫缺陷最常见的形式。唯一可用的救生 目前的治疗方法是骨髓移植，但 这种方法的缺点包括缺乏HLA匹配的供体， 超过2/3的病例和移植后B细胞功能差，主要是由于 到B细胞的非植入。先前证明的负选择 针对表达突变型IL2RG的细胞，导致X染色体偏斜 在女性携带者中失活，使X连锁SCID成为一种有前途的 用逆转录病毒转导的自体 再生干细胞这一战略首先由以下方面执行： 比较不同逆转录病毒构建体的滴度、γ链 在转导细胞系中的表达，功能缺陷的校正 在来自患有确定的IL2RG的SCID患者的EBV-B细胞系中 脐带血干细胞的培养 在SCID小鼠中用IL2RG逆转录病毒转导。这些实验 显示γ链蛋白的显著细胞表面表达， 在来自SCID患者的先前缺陷的细胞系中的免疫荧光。 大量患者细胞系的可用性将允许 潜在显性负突变的评估。scid患者 产前诊断为确定的IL2RG突变的患者， 出生时采集的血液用于实验转导和评估 淋巴细胞生成犬自发性γ链突变导致 目前正在与Paula Henthorn合作研究SCID犬， 联合一个标记实验正在进行中， 将两种逆转录病毒构建体导入骨髓， 用细胞因子预处理的未受影响的狗。骨髓被注入一个 MHC匹配的辐射受体，在8周时表现出实质性的 人γ链的血液和骨髓表达。后续行动 计划在SCID狗中进行实验。