TRANSDUCTION OF BABOON HEMATOPOIETIC STEM CELLS

狒狒造血干细胞的转导

• 批准号: 6358972
• 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: 2000
• 资助国家: 美国
• 起止时间: 2000-09-01 至 2001-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6358972
• 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.

基因疗法在造血系统的可遗传疾病中的有效应用仍然很大程度上未实现。临床研究表明，同种异体干细胞的移植可以纠正许多这些疾病。因此，该项目的目的是将基因转移到狒狒中的骨髓再植物中，作为一种相关的非人类灵长类动物，用于干细胞移植的临床前模型。使用竞争性的重生策略来研究与狒狒基因转移相关的不同变量，我们证明了1）长臂猿白血病病毒型媒介； 2）可以通过从重组人纤连纤维蛋白片段CH-296中培养细胞来实现骨髓再现细胞的有效转导； 3）逆转录病毒介导的基因转移在G-CSF加上SCF（动员）外周血和骨髓祖细胞中得到了增强。然而，尽管在最初的移植期间，血液的瞬时标记水平在10％至20％之间，但对于大多数遗传疾病而言，长期基因转移水平仍然不够高。因此，在转导，然后植入长期重塑干细胞时需要进一步改进。我们建议建立最佳的方法来动员重型细胞，这些方法可以有效地转导，并确定是否可以降低调理而不抑制转导细胞的体内重摄取。另外，由于非人类灵长类动物模型在逻辑上有限，我们将确定是否使用免疫缺陷的小鼠模型提供了相同的信息。最后，该项目将扩展其他项目的发现。研究将侧重于新型混合载体传递骨髓再现细胞的潜力，以及用于RAR和Notch受体家族的配体，以增强基因转移到这些重塑细胞中。非人类灵长类动物提供了一个高度相关的模型，在该模型中可以直接测试这些策略，然后再将其引入人类临床试验。