Anti-HIV Gene Delivery to Human Hematopoietic Cells

描述（由申请人提供）：本提案的目标是研究慢病毒载体传递的毒性和功能，证明基于基因的HIV-1破坏方式在人造血干细胞（hsc），髓细胞和CD4+ T细胞中的作用。据估计，在接受高效抗逆转录病毒治疗（HAART）的艾滋病患者中，有30-45%的人有某种形式的耐药性。因此，耐药hiv -1的持续出现使得开发具有新的作用机制的治疗方法势在必行。我们假设，通过采用经过验证的、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到CD34+和T细胞，抗HIV-1基因阳性细胞可能难以感染，不能有效地复制病毒，并且受保护的细胞会随着时间的推移而丰富。我们的长期目标是提供病毒和细胞基因破坏策略的基础研究结果，这些策略可能会为已经或即将失败的HAART患者提供自体T和CD34+细胞或同种异体CD34+细胞移植来源。与HAART中使用的联合药物策略类似，我们正在完成一个包含多个基因的慢病毒传递载体，这些基因靶向细胞和HIV信息和蛋白质，从而破坏病毒的进入、整合、病毒和细胞功能。此外，如果发生突变，所选择的靶位点应该不太适合病毒。

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.