Gene Therapy in Hematopoietic Cells

造血细胞基因治疗

• 批准号: 7304848
• 负责人: HANS-PETER KIEM
• 金额: $ 56.17万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-15 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7304848
• 关键词: CD34 molecule; Lentivirus; biotechnology; blood cell count; busulfan; disease /disorder model; dogs; flow cytometry; gammaretrovirus; gene therapy; hematopoietic stem cells; hematopoietic tissue transplantation; longitudinal animal study; method development; nonhuman therapy evaluation; nucleic acid repetitive sequence; pharmacokinetics; polymerase chain reaction; pyruvate kinase deficiency; stem cell transplantation; transfection /expression vector; virus integration

The overall objective of this project is to develop strategies, which will allow the efficient and safe treatment of hematopoietic diseases by hematopoietic stem cell (HSC) gene therapy. Unfortunately neither in vitro studies nor studies in the mouse model have been predictive for safety and efficacy of gene transfer in large animals or humans, thus we will use the dog model which will also allow us to evaluate improved strategies directly in a disease model. While gene transfer efficiencies into HSCs in the dog model and other large animals have improved significantly over the past several years, the development of leukemia in 3 patients in the French X-linked severe combined immunodeficiency (XSCID) gene therapy trial has shifted the emphasis from efficacy to safety. Thus, we will use the dog model to study the safety of 3 commonly used integrating vector systems: gammaretrovirus, lentivirus, and foamy virus vectors. Using these vector systems, we have been able to achieve efficient transduction of canine long-term repopulating cells with stable gene transfer levels >5% in a significant number of dogs. Thus, we now have a unique resource available to study in Specific Aim 1 the safety of HSC gene transfer with these vectors. In Specific Aim 2, we will further optimize transduction conditions for foamy and lentiviral vectors to minimize risks from insertional mutagenesis. We will focus on lentivirus and foamy virus vectors since, in contrast to gammaretroviral vectors, these vector systems allow for efficient transduction using short transduction cultures. This is particularly important for stem cell gene therapy in a nonmyeloablative transplant setting where maintenance of stem cells is crucial for the ability to compete with surviving endogenous stem cells. Thus, in Specific Aim 3 we will use these vector systems in a nonmyeloablative setting and also explore whether in vivo selection strategies can improve gene transfer levels after nonmyeloablative conditioning. Finally, in Specific Aim 4 we will test improved gene transfer protocols in a canine genetic disease model. The availability of a clinically relevant large animal model should allow us to quickly translate our findings to clinical HSC gene therapy studies.

该项目的总体目标是制定策略，使高效和安全的 通过造血干细胞（HSC）基因疗法治疗造血系统疾病。不幸的是都没有 体外研究和小鼠模型研究都可以预测基因转移的安全性和有效性 在大型动物或人类中，因此我们将使用狗模型，这也将使我们能够评估改进的 直接在疾病模型中制定策略。虽然在狗模型和其他模型中将基因转移到 HSC 中的效率 大型动物在过去几年中得到显着改善，白血病的发展在3年 法国 X 连锁严重联合免疫缺陷 (XSCID) 基因治疗试验中的患者已发生变化 强调从功效到安全。因此，我们将使用狗模型来研究3种常见的安全性 使用的整合载体系统：γ逆转录病毒、慢病毒和泡沫病毒载体。使用这些向量 系统，我们已经能够实现犬长期再生细胞的有效转导 在大量狗中，稳定的基因转移水平> 5%。因此，我们现在拥有独特的资源 可用于在特定目标 1 中研究使用这些载体进行 HSC 基因转移的安全性。在具体目标 2 中，我们 将进一步优化泡沫和慢病毒载体的转导条件，以尽量减少插入的风险 诱变。我们将重点关注慢病毒和泡沫病毒载体，因为与伽马逆转录病毒相比 载体，这些载体系统允许使用短转导培养物进行有效的转导。这是 对于非清髓性移植环境中的干细胞基因治疗尤其重要 干细胞的维持对于与存活的内源干细胞竞争的能力至关重要。因此，在 具体目标 3 我们将在非清髓性环境中使用这些载体系统，并探讨是否 体内选择策略可以提高非清髓性调理后的基因转移水平。最后，在 具体目标 4 我们将在犬遗传病模型中测试改进的基因转移方案。这 临床相关的大型动物模型的可用性应该使我们能够快速将我们的发现转化为 临床HSC基因治疗研究。