STEM CELL GENE THERAPY FOR AIDS USING AN ANTI-HIV ENVELOPE ANTISENSE MOLECULE

使用抗 HIV 包膜反义分子治疗艾滋病的干细胞基因疗法

• 批准号: 7349499
• 负责人: Stephen E. Braun
• 金额: $ 13.48万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-05-01 至 2007-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7349499
• 关键词: STEM; CELL; GENE; THERAPY; AIDS

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The introduction of inhibitory genes into hematopoietic stem cells offers the potential for long-lived immune reconstitution with progeny cells resistant to HIV-1 replication. However, successful development of stem cell gene therapy for AIDS is likely to require preclinical testing in the rhesus macaque model. We examined the ability of a HIV-1-based lentiviral vector (VRX494) encoding a 937 bp antisense HIV-1 envelope sequence to inhibit viral replication. Because HIV-1 does not replicate in rhesus macaques, chimeric SIV/ HIV-1 viruses with the HIV-1 envelope were used to determine the efficacy of VRX494. Challenge of VRX494-transduced CEMx174 cells resulted in potent inhibition of HIV-1 and several SHIV strains. To evaluate the efficacy of the VRX494 in CD4+ T cells derived from transduced CD34+ cells, rhesus CD34+ bone marrow cells were transduced with VRX494 and then cultured on rhesus fetal thymus stromal cells to induce T cell differentiation. Transduction conditions for CD34+ cells were optimized so as to yield relatively high levels of transduction efficiency (greater than 50 per cent), with minimal effective multiplicity of infection. Purified CD4+GFP+ T cells derived from VRX494-transduced cells strongly inhibited SHIV HXBc2P 3.2 and SHIV 89.6P replication compared to controls. Southern blot analysis of T cell clones derived from transduced CD34+ cells revealed a subset of cells with multiple proviral copies per cell highlighting the importance of optimizing transduction conditions to minimize the possibility of multiple integration events per cell. These results indicate that a lentiviral vector expressing an HIV-1 antisense sequence strongly inhibits HIV-1 and SHIV replication and that the SHIV macaque model should serve as a useful preclinical model to evaluate stem cell gene therapy for AIDS.

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子弹和调查员（PI）可能已经从其他NIH来源获得了主要资金，因此可以在其他清晰的条目中代表。列出的机构适用于该中心，这不一定是调查员的机构。将抑制性基因引入造血干细胞中，具有抗HIV-1复制具有抵抗力的后代细胞的长寿命免疫重建的潜力。然而，成功开发用于艾滋病的干细胞基因治疗可能需要在恒河猴模型中进行临床前测试。我们检查了基于HIV-1的慢病毒载体（VRX494）编码937 bp反义HIV-1包膜序列抑制病毒复制的能力。由于HIV-1在恒河猕猴中不复制，因此使用HIV-1包膜的嵌合SIV/ HIV-1病毒用于确定VRX494的功效。 VRX494转导的CEMX174细胞的挑战导致HIV-1和几种SHIV菌株的有效抑制。为了评估VRX494在源自转导的CD34+细胞的CD4+ T细胞中的疗效，用VRX494转导恒河猴CD34+骨髓细胞，然后在恒和胎儿基质细胞上培养以诱导T T细胞分化。优化了CD34+细胞的转导条件，以产生相对较高的转导效率（大于50％），并且感染的有效多样性最小。与对照组相比，纯化的CD4+ GFP+ T细胞强烈抑制SHIV HXBC2P 3.2和SHIV 89.6p复制。从转导的CD34+细胞中得出的T细胞克隆的Southern印迹分析显示，每个细胞中有多个前病毒拷贝的细胞集，强调了优化转导条件的重要性，以最大程度地减少每个细胞的多个整合事件的可能性。这些结果表明，表达HIV-1反义序列的慢病毒载体强烈抑制HIV-1和SHIV复制，并且SHIV猕猴模型应作为评估AIDS干细胞基因治疗的有用临床前模型。