Improving gene transfer to provide intracellular immuniz

摘要：NHGRI校内项目已经扩大了研究工作，将病毒载体整合到成人造血干细胞（HSC）中，以对抗HIV感染。HSC是所有外周血细胞的最终祖细胞，包括被HIV感染的CD4+淋巴细胞和可以在组织中长时间隔离HIV的巨噬细胞。如果抗HIV元素可以被引入到HSC中，它将被传递给该HSC的所有后代，从而确保连续的、终生的抗HIV细胞的产生。如果这种治疗完全成功，这种治疗将永久地保护患者免受艾滋病毒的传播，并且不需要进一步的治疗。用病毒载体成功修饰HSC有三个重要步骤。首先，病毒载体必须与HSC表面的特定受体结合。我们已经确定两种新型逆转录病毒RD114和FeLV-C的受体在HSC上大量存在，而传统的逆转录病毒受体在HSC上不存在。我们利用小鼠和绵羊异种移植模型，利用带有这些新包膜的逆转录病毒载体，证明了高水平的基因转移到人类HSC中。在接下来的一年里，我们将扩展这些研究，包括携带抗HIV基因的假分型慢病毒载体。HSC修饰的第二个重要步骤是将载体整合到目标细胞的DNA中。由于基因治疗试验中插入性白血病发生的新安全性问题，NHGRI的内部研究人员正在比较慢病毒和逆转录病毒载体的整合位点。我们也在研究调节造血细胞周期进程的几个基因。用抗HIV元素成功修饰HSC的最后一个重要步骤是，必须在成熟的HSC后代中始终产生抗HIV元素，以便细胞始终准备好中断HIV感染。许多研究小组已经表明，随着时间的推移，病毒载体会变得沉默，并停止制造关键元素。为了打击沉默，NHGRI的内部研究人员正在开发和评估含有防止沉默的遗传元素的不同病毒载体。正在开发的病媒中包括含有绝缘体成分的病媒，这些成分可防止与基因沉默有关的变化类型。新型绝缘体正被纳入载体，以评估小鼠模型中的基因沉默。这三项举措的结合将显著提高基于HSC的艾滋病基因治疗成功的前景。预计这些研究结果可以在不久的将来结合到艾滋病基因治疗的临床试验中。NHGRI已经启动了艾滋病的遗传方法，充分利用了校内NHGRI临床研究项目的优势。NHGRI临床研究人员研究遗传性免疫疾病，建立知识，这将影响HIV疫苗的最终生产，并提供对HIV如何使免疫系统失活的理解。NHGRI遗传咨询计划将发起一项努力，为艾滋病患者提供咨询和教育。遗传咨询项目已经完成了对遗传性免疫疾病家庭最有效咨询技术的广泛研究。NHGRI艾滋病倡议将比较已证明对遗传性免疫缺陷疾病有效的咨询方法与对艾滋病家庭有效的咨询方法。这样的心理学研究有望揭示是否需要不同的方法来帮助家庭应对艾滋病及其后果。

Summary: The NHGRI intramural program has expanded research efforts to integrate virus vectors into adult hematopoietc stem cells (HSC) to combat HIV infection. The HSC is the ultimate progenitor of all peripheral blood cells, including the CD4+ lymphocytes that are infected by HIV and the macrophages that can sequester HIV for long periods of time in tissues. If an anti-HIV element can be introduced into HSC, it will be passed along to all of the progeny of that HSC, ensuring the continuous, life-long production of HIV resistant cells. If this therapy were completely successful, this treatment would permanently protect the patient from HIV spread and no further treatments would be required. The successful modification of HSC by viral vectors has three important steps. First the viral vector must bind to a specific receptor on the surface of the HSC. We have determined that the receptors for two novel retroviruses RD114 and FeLV-C are plentiful on HSC, unlike the conventional retrovirus receptors, which are not present on HSC. We have demonstrated high levels of gene transfer into human HSC using retrovirus vectors with these new envelopes using mouse and sheep xenograft models. In the coming year, we will extend these studies to include pseudotyping lentivirus vectors that carry anti HIV genes. The second important step in the modification of HSC requires the vector to become integrated into the DNA of the target cell. Because of new safety concerns regarding insertional leukemogenesis in a gene therapy trial, NHGRI intramural researchers are comparing the integration sites of lentivirus and oncoretrovirus vectors. We are also studying several genes that regulate hematopoietic cell cycle progression. The final important step for successful modification of HSC with anti-HIV elements is that the anti-HIV elements have to be produced in the mature progeny of the HSC at all times so that the cells are always prepared to interrupt HIV infection. Many groups have shown that viral vectors can become silenced over time and stop making the critical elements. To combat silencing, NHGRI intramural investigators are developing and evaluating different virus vectors that contain genetic elements to prevent silencing. Among the vectors under development are vectors containing insulator elements that prevent the type of changes associated with gene silencing. Novel insulators are being incorporated into vectors to evaluate gene silencing in mouse models. The combination of these three initiatives should significantly improve the prospects for successful HSC based AIDS gene therapy. It is anticipated that the results of these studies can be combined into a clinical trial for AIDS gene therapy in the near future. NHGRI has initiated a genetic approach to AIDS that takes full advantage of the strengths of the intramural NHGRI clinical research program. NHGRI clinical researchers study inherited immune disorders building knowledge that will bear on the ultiumate production of HIV vaccines and provide an understanding of how HIV deactivates the immune system. The NHGRI Genetic Counseling program will initiate an effort to counsel and educate individuals with AIDS. The Genetic Counseling program has completed an extensive study of the most effective counseling techniques for families with inherited immune disorders. The NHGRI AIDS initiative will compare counseling approaches that have been shown to be effective in inherited immunodeficiency diseases with counseling for AIDS families. Such psychological studies are expected to reveal whether different approaches are necessary to help families cope with AIDS and its consequences.