Humanized Rag2-/-gammac-/-mice for HIV infection and gene therapy

描述（由申请人提供）：使用合适的动物模型，可以模拟正常功能的人类造血系统所提供的生理条件，将极大地促进HIV发病机制和HIV-1基因治疗结构的临床前评估的研究。由于先前使用的异种移植SCID-hu小鼠模型缺乏产生初级免疫反应的能力，因此只能在缺乏主动免疫反应的情况下评估病毒对人类细胞的致病作用。因此，在这些系统中缺少一个重要的生理维度。这种缺陷现在可以通过使用新出现的具有免疫能力的人源化小鼠模型来潜在地纠正。将人CD34造血干细胞注射到免疫缺陷的新生Rag2-/-yc-/-小鼠体内，可产生多系人造血和功能性免疫系统。最近的证据表明，这些人源化小鼠允许传播HIV感染。该模型具有多谱系人类细胞移植的能力和对HIV感染的易感性，在基于干细胞的环境中评估抗HIV基因治疗构建具有很大的潜力。基因修饰的细胞现在可以测试它们的免疫重建能力。我们最近在构建和评估几种有前途的抗hiv RNA抑制分子（如sirna）方面取得了进展，并建议利用这种新的小鼠模型来回答与基因治疗方法成功相关的几个重要问题。我们的具体目标是：1。进一步建立人源化Rag2共同γ链双敲除（Rag2-/-yc-/-）小鼠HIV感染模型，评价抗HIV基因治疗构建；评估抗hiv基因转导的CD34造血祖细胞在人源化Rag2-/-yc-/-小鼠中的移植潜力和多谱系分化，2。测定终分化T细胞和巨噬细胞中抗hiv基因的表达，并评估它们在体内外对HIV-1的抵抗。4. 评估转基因造血细胞亚群的功能能力和携带转基因细胞的人源化Rag2-/-yc-/-小鼠产生正常免疫反应的能力：这些研究将有助于建立新的HIV/AIDS研究动物模型，以及评估新的基因治疗方法来控制HIV感染。

DESCRIPTION (provided by applicant): Studies on HIV pathogenesis and preclinical evaluation of HIV-1 gene therapeutic constructs would be greatly facilitated by the use of a suitable animal model that can mimic the physiological conditions afforded by a normally functioning human hematopoietic system. As the previously used xenograft SCID-hu mouse models lacked the capacity for generating primary immune responses, viral pathogenic effects on the human cells could only be evaluated in the context of the absence of an active immune response. Thus an important physiological dimension was missing in these systems. This deficiency can now be potentially rectified by the use of a newly emerged immunocompetent humanized mouse model. Injection of human CD34 hematopoietic stem cells into immunodeficient neonatal Rag2-/-yc-/- mice results in multi-lineage human hematopoiesis and the generation of a functional immune system. Recent evidence showed that these humanized mice are permissive for disseminated HIV infection. With the capacity for multi-lineage human cell engraftment and susceptibility for HIV infection, this model shows great potential to evaluate anti-HIV gene therapeutic constructs in a stem cell-based setting. Gene modified cells can now be tested for their capacity for immune reconstitution. We recently made progress in the construction and evaluation of several promising anti-HIV RNA inhibitory molecules such as siRNAs and propose to utilize this new mouse model to answer several important questions relevant for the success of gene therapy approaches. Our specific aims are to: 1. Further develop the humanized Rag2 common gamma chain double knock out (Rag2-/-yc-/-) mouse model for HIV infection to evaluate anti-HIV gene therapeutic constructs, 2. Evaluate the engraftment potential and multi-lineage differentiation of anti-HIV gene transduced CD34 hematopoietic progenitor cells in humanized Rag2-/-yc-/- mice, 3. Determine the expression of anti-HIV genes in terminally differentiated T cells and macrophages and evaluate their resistance to HIV-1 challenge in vitro and in vivo. 4. Evaluate the functional competence of transgenic hematopoietic cell subsets and the capacity of humanized Rag2-/-yc-/- mice bearing transgenic cells to give rise to normal immune responses: These studies will be helpful in developing new animal models for HIV/AIDS research, and in evaluating novel gene therapy approaches to control HIV infection.