DUAL INHIBITOR GENE THERAPY STRATEGIES FOR AIDS

这个子项目是利用资源的许多研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， NCRR赠款不直接向子项目或子项目工作人员提供资金。 即使抗逆转录病毒药物疗法取得了成功，但毒性、病毒抗药性或潜伏感染等相当多的问题也为开发新的治疗方式提供了动力。 基因治疗已被提出作为一种替代方法。已经开发了靶向病毒生命周期所有阶段的多种抑制机制（即核酶、反义、适体、RNAi、病毒诱饵、锌指核酸酶、显性阴性蛋白和胞内抗体）;然而，靶向多个点的抑制剂可能会提供更稳健的抑制作用，降低病毒逃逸的可能性。在RP约翰逊的实验室在NEPRC，我们测试和比较多个载体系统，抑制剂，和目标人群。 作为这项工作的延伸，我正在开发一种两相病毒抑制剂，通过结合来自CD 4的gp 120结合结构域与七肽重复序列。 由于gp 120和gp 41在小瓶结合和融合过程中经历了广泛的构象变化，我们假设双重抑制剂与病毒包膜的结合将打开gp 120并暴露gp 41中的脆弱序列，以通过七肽重复序列进行结合和抑制。 这种类型的抑制剂既可以作为可溶性蛋白发挥作用，也可以拴在细胞膜上。 在这些抑制剂的变体完成克隆和测序后，将在T细胞系、原代T细胞和恒河猴中评估它们阻断病毒复制的功效。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Even with the success of antiretroviral drug therapy, considerable problems such as toxicity, viral resistance or latent infections provide an impetus to develop new therapeutic modalities. Gene therapy has been proposed as one alternative method. Multiple mechanism of inhibition have been developed (i.e. ribozymes, antisense, aptamers, RNAi, viral decoys, zinc finger nucleases, dominant negative proteins, and intrabodies) that target all phases of the viral lifecycle; however, it may be that an inhibitor that targets multiple points will provide more robust inhibition with reduced potential for viral escape. In RP Johnson's laboratory at NEPRC, we tested and compared multiple vector systems, inhibitors, and target populations. As an extension of this work, I am developing a two-phase viral inhibitor by combining the gp120-binding domain from CD4 with the heptad repeat sequences. Because gp120 and gp41 undergo extensive conformation changes during vial binding and fusion, we are hypothesizing that binding of the duel inhibitor to the viral envelope will open up gp120 and expose vulnerable sequences in gp41 for binding and inhibition by the heptad repeat sequences. This type of inhibitor could function either as a soluble protein, or tethered to the cell membrane. After variations of these inhibitors are finished being cloned and sequenced, they will be evaluated in T cell lines, primary T cells, and in rhesus macaque for efficacy in blocking viral replication.