Gene Therapy for Athabascan SCID

阿萨巴斯卡 SCID 基因治疗

• 批准号: 7336836
• 负责人: R. Scott McIvor
• 金额: $ 30.63万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7336836
• 关键词: Allogenic; Animals; Autologous; B-Lymphocytes; Bone Marrow Transplantation; Cell Transplants; Cells; Code; Common Lymphoid Progenitor; Complementary DNA; Condition; Development; Disease; Disruption; Embryo; Engraftment; Evaluation; Experimental Models; Fibroblasts; Gene Rearrangement; Gene Transfer; Generations; Genes; Genetic; Goals; HIV-1; Hematopoietic; Hematopoietic Stem Cell Transplantation; Hematopoietic stem cells; Human; Immune; Immune response; Immunologic Deficiency Syndromes; Laboratories; Lead; Lentivirus Vector; Lymphoid Cell; Marrow; Mediating; Molecular; Morbidity - disease rate; Mus; Native Americans; Nonhomologous DNA End Joining; Patients; Population; Proteins; Research Personnel; Risk; Role; Severe Combined Immunodeficiency; Sleeping Beauty; Stem cell transplant; Stem cells; System; Systems Integration; T-Lymphocyte; Testing; Therapeutic; Transplantation; V(D)J Recombination; Viral; Viral Genes; allogeneic disease; artemis; base; cellular transduction; design; embryonic stem cell; gene therapy; insight; mortality; mouse model; progenitor; restoration; vector

DESCRIPTION (provided by applicant): SCIDA (T-B-NK+ severe combined immunodeficiency of Athabascan-speaking Native Americans) is a primary immune deficiency of humans resulting from absence of Artemis protein, required for nonhomologous end-joining in V(D)J gene rearrangement. Currently, the only effective therapy for SCIDA is allogeneic bone marrow transplant, for which there is a high risk of morbidity and mortality. Recently, the laboratory of collaborating investigator Dr. Mort Cowan, generated a mouse model of SCIDA by homologous disruption of the murine Scida gene in embryonic stem cells. Here we propose to use this Scida+ mouse model as a test system to evaluate two different gene therapy approaches for SCIDA. In Aim 1, lentiviral vectors based on HIV-1 will be designed for optimized expression of the Scida cDNA sequence in lymphoid cell populations. These vectors will first be tested in Scida -/-mouse embryo fibroblasts (MEF's) for transduction and restoration of non-homologous end-joining activity. These vectors will then be used to introduce the Scida gene into Scida -/- marrow cells, evaluating recipient animals for the presence and selective outgrowth of positively transduced cell populations as well as for correction of immunodefiency. In Aim 2, nonviral vectors based on the Sleeping Beauty transposon system will be designed for introduction and optimized expression of the Scida coding sequence, first testing for the feasibility of transposition in Scida -/- mouse embryo fibroblasts. These vectors will then be electroporated into Scida -/- marrow cells (unenriched marrow, marrow enriched for stem cells, and marrow enriched for common lymphoid progenitor cells) with subsequent evaluation for engraftment, selective outgrowth and correction of immunodeficiency after transplantation into Scida -/- recipients. Results from these studies are anticipated to provide insight into the conditions under which introduction of the SCIDA gene into appropriate lympho-hematopoietic progenitors can lead to correction of immunodeficiency in scida -/- animals, with the ultimate goal of applying such gene transfer approaches to the treatment of SCIDA in humans.

描述（由申请人提供）：SCIDA（阿萨巴斯卡语美洲原住民的 T-B-NK+ 严重联合免疫缺陷）是由于缺乏 Artemis 蛋白而导致的人类原发性免疫缺陷，Artemis 蛋白是 V(D)J 基因重排中非同源末端连接所必需的。目前，SCIDA唯一有效的治疗方法是同种异体骨髓移植，其发病率和死亡率较高。最近，合作研究员 Mort Cowan 博士的实验室通过同源破坏胚胎干细胞中的小鼠 Scida 基因，建立了 SCIDA 小鼠模型。在这里，我们建议使用 Scida+ 小鼠模型作为测试系统来评估两种不同的 SCIDA 基因治疗方法。在目标 1 中，将设计基于 HIV-1 的慢病毒载体，以优化 Scida cDNA 序列在淋巴细胞群中的表达。这些载体将首先在 Scida -/-小鼠胚胎成纤维细胞 (MEF's) 中进行测试，用于转导和恢复非同源末端连接活性。然后，这些载体将用于将 Scida 基因引入 Scida -/- 骨髓细胞，评估受体动物的阳性转导细胞群的存在和选择性生长以及免疫缺陷的纠正。在目标 2 中，将设计基于睡美人转座子系统的非病毒载体，用于 Scida 编码序列的引入和优化表达，首先测试 Scida -/- 小鼠胚胎成纤维细胞中转座的可行性。然后将这些载体电穿孔到 Scida -/- 骨髓细胞（未富集的骨髓、富集干细胞的骨髓和富集共同淋巴祖细胞的骨髓）中，随后在移植到 Scida -/- 受体后评估植入、选择性生长和免疫缺陷的纠正。这些研究的结果预计将深入了解将 SCIDA 基因引入适当的淋巴造血祖细胞中可以纠正 scida -/- 动物的免疫缺陷，最终目标是将这种基因转移方法应用于人类 SCIDA 的治疗。