TCR GENE USAGE AND QUANTITATION OF HIV SPECIFIC CTL

尽管存在一种强有力的和广泛反应性的HIV-1特异性抗体， 受感染个体的CTL应答，几乎进展为AIDS 不可避免地发生。 导致疾病进展的因素 这种强CTL应答的设定还没有被很好地定义。 一 一种假说表明，关键表位内的突变可能允许 HIV-1逃避细胞毒性T淋巴细胞的免疫识别。 在这 在这种情况下，宿主产生新的CTL应答的能力可能是一种 抑制病毒复制的关键因素。 另一个假说 提示HIV特异性CTL克隆的缺失在HIV感染过程中， 疾病，即“克隆衰竭”可能是一个因素解释下降， 伴随疾病进展的CTL活性。 我们已经开发 对HIV-1特异性CTL的T细胞受体基因进行测序的方法， 已经表明，在感染个体中强烈的CTL应答可能是 由于效应细胞的寡克隆扩增， 几年 我们还发现了HIV-1变异 在CTL表位内的一个突变导致先前CTL识别的丧失， 从该个体分离，但大多数CTL应答 仍然针对较小的体内序列变体。 的能力 通过分析跟踪特定HIV表位特异性CTL的群体 将使我们能够确定这些细胞的命运， 感染的过程。 本提案中的实验目的 是使用T细胞受体（TCR）序列分析作为评估 对已知HIV-1表位的CTL应答的多样性，并评估 在这些表位中的HIV-1序列变异， 不同的病程。 我们的目标是检验以下假设： a）针对多种HIV的广泛定向的HIV特异性CTL应答 表位与缓慢的疾病进展相关。B）强CTL 非进展性疾病受试者的反应将由以下因素介导： 具有异质性TCR的CTL克隆群，其能够更好地 科普HIV-1序列变异。 c）如果存在HIV-1序列变异 在这些受试者的这些表位中，存在CTL群体， 能够识别这些变异，表明有能力应对艾滋病毒 变化量 具体而言，我们建议1）确定表位 人HIV特异性CTL克隆特异性和TCR异质性 匹配三个或更多I类等位基因但具有不同疾病状态 和病毒载量。 2)确定定义的克隆应答的寿命 通过测量CTL前体频率和通过探测TCR cDNA文库， 特异性寡核苷酸探针。 3)关联大小和 存在体内病毒的特异性克隆应答的异质性 在不同疾病结果的人群中。 了解 HIV-1特异性CTL应答对长期非免疫应答的贡献 进行性感染可能会导致增加这只手臂的实际策略 免疫系统的反应。

Despite the presence of a vigorous and broadly reactive HIV-1 specific CTL response in infected individuals, progression to AIDS almost inevitably occurs. The factors that allow disease progression in the setting of this strong CTL response have not been well defined. One hypothesis suggests that mutations within critical epitopes may allow HIV-1 to escape immune recognition by cytotoxic T lymphocytes. In this case the ability of the host to generate new CTL responses may be a critical factor in containing viral replication. Another hypothesis suggests that the deletion of HIV-specific CTL clones over the course of disease i.e. "clonal exhaustion" may be a factor explaining the decline in CTL activity that accompanies disease progression. We have developed methods to sequence the T cell receptor genes of HIV-1-specific CTL and have shown that the vigorous CTL response in an infected individual may be due to an oligoclonal expansion of effector cells that can persist for several years. We have also found instances in which HIV-1 variation within CTL epitopes leads to a loss of recognition by CTL previously isolated from that individual, yet the majority of the CTL response remains directed at a minor in vivo sequece variant. Our ability to follow population of particular HIV epitope-specific CTL by the analysis of their TCR usage will allow us to determine the fate of these cells over the course of infection. The purpose of the experiments in this proposal is to use T cell receptor (TCR) sequence analysis as a means to assess the diversity of the CTL response to known HIV-1 epitopes and to assess variation of HIV-1 sequences within these epitopes in patients with different disease courses. Our aim is to test the following hypotheses: a) A broadly directed HIV-specific CTL response, against multiple HIV epitopes correlates with slow disease progression. b) The vigorous CTL responses in subjects with non-progressing illness will be mediated by a population of CTL clones with heterogeneous TCR that are better able to cope with HIV-1 sequence variation. c) If HIV-1 sequence variation exists within these epitopes in these subjects, CTL populations exist that are able to recognize these variants, indicating an ability to response to HIV variation. Specifically we propose to 1) Determine the epitope specifically and TCR heterogeneity of HIV-specific CTL clones in persons matched at three or more class I alleles but with different disease states and viral loads. 2) Determine the longevity of - defined clonal responses by measuring CTL precursor frequency and by probing TCR cDNA libraries with specific oligonucleotide probes. 3) Correlate the magnitude and heterogeneity of specific clonal responses with the in vivo virus present in persons with different disease outcomes. Understanding the contribution of the HIV-1- specific CTL response to long-term non progressing infection may lead to practical strategies to augment this arm of the immune response.