TCR GENE USAGE AND QUANTITATION OF HIV SPECIFIC CTL

尽管存在一种强有力的和广泛反应的HIV-1特异性 感染者的CTL反应，几乎进展为艾滋病 不可避免地会发生。导致疾病进展的因素 这种强烈的CTL反应的设置还没有很好地定义。一 假说表明，关键表位的突变可能会导致 HIV-1逃避细胞毒性T淋巴细胞的免疫识别。在这 如果主机生成新的CTL响应的能力可以是 遏制病毒复制的关键因素。另一种假设 建议在整个过程中删除HIV特异性CTL克隆 疾病，即“克隆性衰竭”，可能是导致 伴随疾病进展的CTL活动。我们已经开发出 方法对HIV-1特异性CTL的T细胞受体基因进行测序 已经表明，感染者体内强烈的CTL反应可能是 由于效应细胞的寡克隆性扩增可以持续 好几年了。我们还发现了HIV-1变异的例子 在CTL表位内导致先前CTL识别的丢失 与该个体隔离，但大多数CTL反应 仍然是针对体内的一个微小的序列变体。我们有能力 通过分析跟踪特定HIV表位特异性CTL的人群 它们的TCR使用率将使我们能够确定这些细胞的命运 感染的过程。本提案中实验的目的是 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反应的多样性和评估 慢性支气管炎患者这些表位中HIV-1序列的变异 不同的病程。我们的目标是检验以下假设： A)针对多种艾滋病毒的广泛定向艾滋病毒特异性CTL反应 表位与疾病进展缓慢相关。B)充满活力的CTL 非进展性疾病受试者的反应将通过 具有异种TCR的CTL克隆群体，能够更好地 应对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，CTL群体存在 能够识别这些变异，表明对艾滋病毒的反应能力 变种。具体地说，我们建议1)确定表位 人体内HIV特异性CTL克隆的特异性和TCR异质性 匹配三个或更多的I类等位基因，但疾病状态不同 和病毒载量。2)确定定义的克隆反应的寿命 通过测量CTL前体频率和探测TCR cDNA文库 用特定的寡核苷酸探针。3)将震级和 体内病毒特异性克隆反应的异质性 在有不同疾病结局的人中。了解 HIV-1特异性CTL应答对长期非 进行性感染可能会导致实际的策略来增强这一手臂 对免疫反应的影响。

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.