Anti-HIV Gene Delivery to Human Hematopoietic Cells

描述（由申请方提供）：本提案的目的是研究慢病毒载体递送的、经证实的基于基因的HIV-1破坏模式在人造血干细胞（HSC）、骨髓和CD 4 + T细胞中的毒性和功能。据估计，30-45%的接受高效抗逆转录病毒治疗（HAART）的艾滋病患者具有可证实的病毒，具有某种形式的耐药性。因此，抗药性HIV-1的不断出现使得开发具有新作用机制的治疗药物势在必行。我们认为，通过采用经证实的、基于基因的HIV-1破坏模式的组合，以及使用慢病毒载体递送至CD 34+和T细胞，抗HIV-1基因阳性细胞可能对感染难治，不能有效地复制病毒，并且受保护的细胞将随着时间的推移而富集。我们的长期目标是提供关于病毒和细胞基因破坏策略的基础研究结果，这些策略可能会为HAART失败或失败的患者提供自体T和CD 34+细胞或同种异体CD 34+细胞移植来源。类似于用于HAART的组合药物策略，我们正在完成一种慢病毒递送载体，该载体含有靶向细胞和HIV信息和蛋白质的多个基因，从而破坏病毒进入、整合、病毒和细胞功能。此外，已经选择了靶位点，如果发生突变，所产生的病毒应该不太适合。 我们的目标将通过成功完成四个高度相互作用的特定目标来实现：1）包含多个HIV-1破坏基因的完整慢病毒载体。2)抗HIV慢病毒载体在不破坏正常细胞功能的情况下是否能保护细胞免受HIV-1感染？3)抗HIV基因的表达是否改变人类造血和胸腺生成？4)含有抗HIV基因的HSC是否会产生受HIV-1攻击保护的髓样和CD 4 + T细胞？ 这些研究完成后，将提供关于含有靶向HIV-1及其细胞通路的多个抗病毒基因的慢病毒载体是否破坏造血功能的信息。我们的研究结果也将提供深入了解是否选定的抗病毒基因的组合赋予细胞保护和选择性的优势，在HIV-1感染的存在。最后，预计在我们的研究期间开发和验证的许多基因破坏和慢病毒载体递送策略将适用于其他基础研究和临床用途。 项目叙述：据估计，30-45%的接受高效抗逆转录病毒治疗（HAART）的艾滋病患者具有可证实的病毒，具有某种形式的耐药性。因此，抗药性HIV-1的不断出现需要新的治疗方法。我们的长期目标是提供关于病毒和细胞抗HIV-1策略的基因递送的基础研究结果，使血细胞抵抗HIV-1感染并减少HIV-1的生长。这些新疗法可用于HAART失败或即将失败的患者的T细胞和CD 34+细胞移植来源。

DESCRIPTION (provided by applicant): The goals for this proposal are to investigate toxicity and function of lentiviral vector delivered, proven gene- based HIV-1 disruption modalities in human hematopoietic stem cells (HSCs), myeloid and CD4+ T cells. It is estimated that 30-45% of AIDS patients on Highly Active Anti-Retroviral Therapy (HAART) having demonstrable virus have some form of drug resistance. Thus, the continuing emergence of drug resistant HIV-1s makes it imperative to develop therapeutics with novel mechanisms of action. We posit that by employing a combination of proven, gene-based HIV-1 disruption modalities, and the use of lentiviral vectors for delivery to CD34+ and T cells, anti-HIV-1 gene positive cells may be refractory to infection, not replicate virus efficiently, and protected cells will enrich over time. Our long-term goals are to provide basic research findings on viral and cellular gene disruption strategies that may find use for autologous T and CD34+ cell or allogenic CD34+ cell transplant sources for patients that have or are going to fail HAART. Analogous to combination drug strategies used for HAART, we are completing a lentiviral delivery vector containing multiple genes that target cellular and HIV messages and proteins, thereby disrupting viral entry, integration, viral and cellular function. Moreover, target sites have been selected that if mutations arise the resulting virus should be less fit. Our goals will be achieved by the successful completion of four highly interactive Specific Aims: 1) Complete lentiviral vectors containing multiple HIV-1 disruption genes. 2) Do anti-HIV lentiviral vectors provide protection from HIV-1 while not disrupting normal cellular function? 3) Does expression of anti-HIV genes alter human hematopoiesis and thymopoiesis? 4) Do HSCs containing anti-HIV genes give rise to myeloid and CD4+ T cells that are protected from HIV-1 challenge? When completed these studies will provide information on whether lentiviral vectors containing multiple anti- viral genes targeting HIV-1 and its cellular pathways disrupt hematopoietic function. Our findings will also provide insight into whether selected combinations of anti-viral genes confer a cell protective and selective advantage in the presence of HIV-1-infection. Lastly, it is anticipated that many of the gene disruption and lentiviral vector delivery strategies developed and validated during our studies will be applicable for other basic research and clinical uses. Project Narrative: It is estimated that 30-45% of AIDS patients on Highly Active Anti-Retroviral Therapy (HAART) having demonstrable virus have some form of drug resistance. Thus, the continuing emergence of drug resistant HIV-1s requires new therapies for treatment. Our long-term goals are to provide basic research findings on gene delivery of viral and cellular anti-HIV-1 strategies that make blood cells resistant to HIV-1 infection and reduce growth of HIV-1. These new therapies may be utilized for T cell and CD34+ cell transplant sources for patients that have or are going to fail HAART.