STEM CELL ADHESION IN GROWTH AND TRANSDUCTION

干细胞生长和转导中的粘附

• 批准号: 6879595
• 负责人: David Collin Williams
• 金额: $ 35.69万
• 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/6879595
• 关键词: Retroviridae; biotechnology; cell adhesion; cell cycle; chimeric proteins; cytokine receptors; fibronectins; gene therapy; genetic transduction; hematopoietic stem cells; hematopoietic tissue transplantation; messenger RNA; microarray technology; stem cell transplantation; tissue /cell culture; transfection /expression vector

Severe combined immunodeficiency disease (SCID) is a fatal congenital disease of children. Approximately 1/3 of cases are associated with the deficiency of adenosine deaminase (ADA) and some of the remaining cases with the X-linked variant have mutations of the IL-2 gamma receptor gene. The severe nature of this illness, the deficiency of single peptide proteins, and the ability to correct the disease by allogeneic bone marrow transplantation have led many investigators to explore the use of gene transfer technology as a therapeutic option for the treatment of SCID. The long term goal of the work proposed here is to utilize gene transfer technology to effect efficient transfer into reconstituting hematopoietic stem cells and stable expression in progeny of these cells to genetic sequences defective in SCID in order to provide life-long cure of this disease. The basis of the approaches to gene transfer proposed in this grant include the realization that published data, including from our own laboratory, demonstrate that current protocols for infection of reconstituting stem cells of large animals species are not optimal. Furthermore, a wealth of experimental data suggests that the hematopoietic microenvironment is a source of both positive and negative regulators of hematopoiesis and these signals may be important for successful and efficient transduction of primitive hematopoietic stem cells. The hypothesis to be tested in this proposal is that transduction of long term reconstituting stem cells can be accomplished in large animal transplant models by retroviral infection in the presence of critical components of the hematopoietic microenvironment. We will utilize our previously characterized simplified retroviral vector which expresses the ADA cDNA and a new set of vectors which encode the IL-2 gamma receptor cDNA for these experiments. We will utilize both in vitro and in vivo assays for human stem cells to analyze transduction efficiency and expression of introduced sequences. In addition, we will continue to use primate transplants to evaluate gene technology in a large animal autologous transplant model.

严重的联合免疫缺陷疾病（SCID）是致命的先天性 儿童疾病。 大约有1/3的案例与 腺苷脱氨酶（ADA）和剩余的一些 具有X连锁变体的病例具有IL-2伽马受体的突变 基因。 这种疾病的严重性质，单身的不足 肽蛋白以及通过同种异体纠正疾病的能力 骨髓移植使许多研究人员探索 将基因转移技术用作治疗的治疗选择 scid。 这里提出的工作的长期目标是利用基因 转移技术以有效地转移到重建 造血干细胞和这些细胞后代的稳定表达 遗传序列在SCID中有缺陷以提供终身治愈 这种疾病。 提出的基因转移方法的基础 在这笔赠款中包括已发布数据的认识，包括 我们自己的实验室证明了当前感染的方案 重建大型动物物种的干细胞不是最佳的。 此外，大量的实验数据表明 造血微环境是阳性和阴性的来源 造血和这些信号的调节剂对 原始造血茎的成功有效转导 细胞。 在此提案中要检验的假设是转导 长期重建干细胞可以在大的 在存在的情况下，通过逆转录病毒感染的动物移植模型 造血微环境的关键成分。 我们将 利用我们先前表征的简化逆转录病毒矢量 表达ADA cDNA和一组新编码IL-2的向量 这些实验的伽马受体cDNA。 我们将在体外使用 并在体内分析人类干细胞分析转导 引入序列的效率和表达。 此外，我们将 继续使用灵长类动物移植来评估基因技术 大动物自体移植模型。