Selectable Gamma-Globin Lentiviral Vectors for SCD

用于 SCD 的可选伽马珠蛋白慢病毒载体

• 批准号: 6508640
• 负责人: DEREK A PERSONS
• 金额: $ 33.75万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6508640
• 关键词: Lentivirus; bone marrow; disease /disorder model; drug design /synthesis /production; drug resistance; erythrocytes; flow cytometry; gene expression; gene induction /repression; gene therapy; genetic enhancer element; genetic promoter element; globin; green fluorescent proteins; hematopoietic stem cells; hemoglobin F; laboratory mouse; messenger RNA; methyltransferase; nonhuman therapy evaluation; sickle cell anemia; tissue /cell culture; transfection /expression vector

DESCRIPTION (provided by applicant): This project is focused on the development of a gene therapy approach to sickle cell disease using a gamma-globin lentiviral vector with the capacity to permanently integrate into the genome of hematopoietic stem cells (HSCs), thereby providing the possibility of a lifelong cure. Our efforts will concentrate on satisfying two critical requirements for the eventual success of this approach. The first is achieving high level, sustained erythroid-specific expression of a transferred gamma-globin expression cassette. Recently, we have developed a gamma-globin lentiviral vector with the capacity to achieve HbF level of 10 percent in the red cells of mice. Since it is likely that a therapeutic impact for sickle cell disease will require higher levels of HbF per cell, the first specific aim is centered on modifying our first generation vector to further increase expression. This will be done through a series of carefully planned alterations designed to boost both the level and persistence of gamma-globin expression. Vectors modified to augment transcriptional activity and dampen position effect variegation and silencing will be evaluated in both in vitro and in vivo studies. These experiments will culminate in testing the therapeutic efficacy of optimized vectors in two murine models of sickle cell disease, which we have acquired. The second specific aim focuses on developing a gamma-globin vector also containing a selectable gene (methylguanine methyltransferase, MGMT), previously shown to enable in vivo selection of HSCs. We estimate that at least 10-20 percent of HSCs capable of giving rise to gamma-globin expressing red cells will be required for a therapeutic effect in sickle cell disease. Therefore, it is likely that in vivo selection will be needed in a human therapeutic trial to increase the subtherapeutic, small proportion of transduced HSCs in recipients that will result from limited gene transfer efficiency and the preferable use of non-myeloablative conditioning. In vivo selection experiments in both normal mice and in the two sickle cell murine models are proposed in Specific Aim 2. The ultimate goal is to obtain therapeutic in vivo selection of y-globin expressing cells in the sickle cell disease models. Progress in these two areas would have substantial impact on the planning of initial clinical trials and would bring gene therapy closer to being a potential treatment for sickle cell disease in the near future.

描述（由申请人提供）：该项目的重点是开发镰状细胞病的基因治疗方法，该方法使用能够永久整合到造血干细胞（HSC）基因组中的γ-珠蛋白慢病毒载体，从而提供终身治愈的可能性。我们的努力将集中在满足这种方法最终成功的两个关键要求上。第一个是实现转移的γ-珠蛋白表达盒的高水平、持续的红细胞特异性表达。最近，我们开发了一种γ-珠蛋白慢病毒载体，能够在小鼠红细胞中达到10%的HbF水平。由于镰状细胞病的治疗效果可能需要每个细胞更高水平的HbF，因此第一个具体目标集中在修饰我们的第一代载体以进一步增加表达。这将通过一系列精心计划的改变来实现，这些改变旨在提高γ-珠蛋白表达的水平和持久性。将在体外和体内研究中评价经修饰以增强转录活性并抑制位置效应杂色和沉默的载体。这些实验将最终测试优化的载体在两个镰状细胞病小鼠模型中的治疗效果，我们已经获得了。第二个具体目标集中于开发还含有可选择基因（甲基鸟嘌呤甲基转移酶，MGMT）的γ-珠蛋白载体，所述可选择基因先前显示能够在体内选择HSC。我们估计，至少10- 20%的造血干细胞能够产生γ-珠蛋白表达的红细胞将需要在镰状细胞病的治疗效果。因此，很可能在人类治疗试验中需要体内选择以增加受体中亚治疗性的、小比例的转导的HSC，这将由有限的基因转移效率和优选使用非清髓性调节引起。具体目标2中提出了正常小鼠和两种镰状细胞小鼠模型中的体内选择实验。最终目标是在镰状细胞病模型中获得γ-珠蛋白表达细胞的治疗性体内选择。这两个领域的进展将对初步临床试验的规划产生重大影响，并将使基因疗法在不久的将来更接近于成为镰状细胞病的潜在治疗方法。