TRANSDUCTION OF BABOON HEMATOPOIETIC STEM CELLS

狒狒造血干细胞的转导

• 批准号: 6494850
• 负责人: ROBERT GOFF ANDREWS
• 金额: $ 20.94万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-01 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6494850
• 关键词: CD34 molecule; Retroviridae; baboons; biotechnology; bone marrow; cell differentiation; cell growth regulation; cell population study; colony stimulating factor; cytokine; flow cytometry; gene expression; gene therapy; genetic transduction; hematopoiesis; hematopoietic growth factor; hematopoietic stem cells; stem cell factor; stem cell transplantation; tissue /cell culture; transfection /expression vector

The effective application of gene therapy to heritable diseases of the hematopoietic system remains largely unrealized. Clinical studies demonstrate that the transplantation of allogeneic stem cells can correct many of these diseases. Therefore the goal of this project is to enhance gene transfer into marrow repopulating cells in baboons as a relevant, non-human primate, pre-clinical model for stem cell transplantation. Using a competitive repopulation strategy to study different variables related to gene transfer in baboons we have demonstrated 1) that Gibbon Ape Leukemia Virus-pseudotype vectors; 2) that efficient transduction of marrow repopulating cells can be accomplished by culturing cells from over the recombinant human fibronectin fragment CH-296; 3) that retrovirus mediated gene transfer is enhanced in G-CSF plus SCF primed (mobilized) peripheral blood and marrow progenitors. However, in spite of transient levels of marking in blood of between 10% and 20% during the initial post-transplant period, long-term gene transfer levels still are not high enough for most genetic diseases. Therefore, further improvements are needed in the transduction and then the engraftment of long-term repopulating stem cells. We propose to establish optimal methods for mobilization of repopulating cells that can be efficiently transduced and determine whether conditioning can be reduced without inhibiting in vivo repopulation by transduced cells. In addition, as the non-human primate model is logistically limited, we will determine whether identical information is provided using an immune-deficient mouse model. Finally, this project will extend findings from other projects. Studies will focus on the potential of novel hybrid vectors to transduce marrow repopulating cells and for ligands for the RAR and Notch families of receptors to enhance gene transfer into these repopulating cells. Non-human primates provide a highly relevant model in which these strategies can be directly tested before being introduced into human clinical trials.

基因治疗在造血系统遗传性疾病中的有效应用在很大程度上仍未实现。临床研究表明，同种异体干细胞移植可以纠正许多此类疾病。因此，该项目的目标是增强狒狒骨髓再生细胞的基因转移，作为干细胞移植的相关非人类灵长类动物临床前模型。使用竞争性繁殖策略来研究与狒狒基因转移相关的不同变量，我们已经证明：1）长臂猿白血病病毒假型载体； 2)可以通过培养来自重组人纤连蛋白片段CH-296的细胞来实现骨髓再生细胞的有效转导； 3)逆转录病毒介导的基因转移在G-CSF加SCF引发的（动员的）外周血和骨髓祖细胞中得到增强。然而，尽管在移植后的最初阶段，血液中的标记水平短暂在 10% 到 20% 之间，但对于大多数遗传疾病来说，长期基因转移水平仍然不够高。因此，需要进一步改进转导以及长期再生干细胞的植入。我们建议建立可有效转导的再增殖细胞动员的最佳方法，并确定是否可以在不抑制转导细胞体内再增殖的情况下减少调节。此外，由于非人类灵长类动物模型在逻辑上受到限制，我们将确定使用免疫缺陷小鼠模型是否提供相同的信息。最后，该项目将扩展其他项目的发现。研究将集中于新型杂交载体转导骨髓再生细胞的潜力，以及 RAR 和 Notch 受体家族的配体增强基因转移到这些再生细胞中的潜力。非人类灵长类动物提供了一个高度相关的模型，在将这些策略引入人类临床试验之前可以直接测试这些策略。