TARGETED VECTORS TO BLOCK HIV1 REPLICATION

阻断 HIV1 复制的靶向载体

• 批准号: 6078547
• 负责人: RALPH C DORNBURG
• 金额: $ 24.32万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-15 至 2004-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6078547
• 关键词: AIDS; AIDS therapy; HIV infections; SCID mouse; cell type; electroporation; fluorescence microscopy; gene expression; gene therapy; genetic manipulation; genetic transcription; human immunodeficiency virus 1; nonhuman therapy evaluation; polymerase chain reaction; transfection /expression vector; virus replication

This proposal seeks funding for the systemic build-up and testing of retroviral vectors, which transduce anti-HIV-1 therapeutic genes into human hematopoietic cells relevant in vivo gene therapy of HIV-1 infection. In the past years, our laboratories have made significant progress in the development of two important areas in gene therapy for HIV-1 infection: (1) Dr. Pomerantz's lab has developed a series of anti- HIV-1 single chain antibodies, which significantly inhibit HIV-1 replication at early stages of the retroviral life cycle. (2) Dr. Dornburg's laboratory has developed retroviral vectors, which display single chain antibodies (scAs) or other ligands on the viral surface of spleen necrosis virus, SV. Such vectors enabled an efficient (up to 10/6 cfu/ml), cell-type- specific gene transfer into various human hematopoietic cells. Cell-type- specificity was mediated by the scA. Now, experiments will be performed to combined these two technologies to establish potent anti-HIV-1 therapeutic gene transfer systems for in vivo gene therapy for HIV-1 infection. (1) Recently, three different SNV-derived packaging systems have been developed to transduce gene specifically into various human T- cells or CD34-positive hematopoietic stem cells with efficiencies above 10/6 cfu/ml. Using these systems, vectors will be developed and systematically improved to transduce anti-HIV-1 therapeutic genes developed and tested in Dr. Pomerantz's laboratory into human T-cells. The efficiency of the gene transfer and the level of protection against HIV-1 will be determined and compared to other vector systems. (2) Cell- type specificity and the efficiency of infection of all targeting vectors will be tested in SCID mice model systems. E.G., human target and non-target cells will be implanted into SCID mice, followed by the injection of retroviral vector preparations, specific for one cell type. First, human cells as well as various mouse tissues will be investigated for the presence of marker genes transduced by the targeting vector particles. Next, the experiments will be expanded to test the therapeutic effect of anti-HIV-1 genes in vivo.

该提案寻求资金用于全身积累和测试逆转录病毒载体，该载体将抗HIV-1治疗基因转导至HIV-1感染的体内基因疗法中的人类造血细胞。 在过去的几年中，我们的实验室在HIV-1感染基因治疗的两个重要领域的发展方面取得了重大进展：（1）Pomerantz博士实验室开发了一系列抗HIV-1单链抗体，这些抗体在逆转录病毒生命周期的早期阶段显着抑制了HIV-1复制。 （2）Dornburg博士的实验室已经开发了逆转录病毒载体，该载体在脾脏坏死病毒的病毒表面上显示单链抗体（SCA）或其他配体。这种载体使得有效（最多10/6 CFU/mL），细胞类型 - 特异性基因转移到各种人造血细胞中。细胞类型特异性是由SCA介导的。现在，将进行实验以组合这两种技术，以建立用于HIV-1感染体内基因治疗的有效的抗HIV-1治疗基因转移系统。 （1）最近，已经开发出三种不同的SNV衍生包装系统，以将基因专门转导至各种人类T细胞或CD34阳性造血干细胞，其效率高于10/6 CFU/ML。 使用这些系统，将开发并系统地改进向量，以转导Pomerantz博士的实验室中开发和测试的抗HIV-1治疗基因。基因转移的效率和针对HIV-1的保护水平将被确定并与其他向量系统进行比较。 （2）将在SCID小鼠模型系统中测试所有靶向载体的细胞类型特异性和所有靶向载体的感染效率。例如，人类靶标和非目标细胞将植入SCID小鼠，然后注射逆转录病毒载体制剂，特异于一种细胞类型。首先，将研究人类细胞以及各种小鼠组织，以实现靶向载体颗粒转导的标记基因。接下来，将扩展实验以测试体内抗HIV-1基因的治疗作用。