HIV Vaccine Induced CTL: Qualitative/Functional Analyses

当今疫苗学面临的主要科学挑战之一是开发一种有效的艾滋病疫苗。已有证据表明，MHC I类限制的CD8+细胞毒性T淋巴细胞(CTL)反应可能对HIV感染具有保护作用。随着测试候选艾滋病疫苗的I/II阶段临床试验的进展，基于金丝雀痘的疫苗策略已经在相当数量的未感染的免疫志愿者中诱导出抗HIV-1 CTL。虽然我们已经开始更好地了解激发的CTL反应的特异性和持续时间，但对这些反应的定性方面仍然知之甚少。基于与最近完成的试验相关联的初步研究，这一提议的具体目的旨在检验疫苗诱导的抗HIV CTL反应与自然感染HIV引起的反应在性质上不同的主要假设。研究的目的是从抗原特异性体外刺激(IVS)培养中鉴定CTL株，最终CTL克隆，关于亲和力、TCR受体宽度以及对感染代表基因多样性病毒分支的初级分离株感染的自体淋巴细胞靶细胞的细胞溶解和非细胞溶解病毒抑制反应。亲和力将通过传统的多肽滴定和多肽/MHC四聚体结合来测量。四聚体还将被用于跟踪疫苗接种者PBMC中的表位特异性CTL及其成熟阶段。还将在体外CTLp扩增水平上研究金丝雀痘载体对接种者CTL反应的偏向能力，使用一组不同的表达HIV的重组载体在不同的基于APC的抗原特异性刺激策略以及HIV-1特异性激活的水平上。最后，将进行比较研究，以确定疫苗诱导的CTL反应的质量是否更类似于在HIV-1感染的急性阶段发展的CTL反应，特别是在与长期病毒刺激和感染相关的免疫耗竭变得明显之前的结构化早期治疗中断有关的情况下。通过对疫苗诱导的CTL质量的新见解，这些研究可能极大地影响基于载体的疫苗的进一步发展。

One of the major scientific challenges in vaccinology today is the development of an effective AIDS vaccine Evidence has been accumulated that MHC class I restricted CD8+ cytotoxic T lymphocyte (CTL) responses may provide protection against HIV infection. As Phase I/II clinical trials for testing of candidate AIDS vaccines progresses, canarypox-based vaccine strategies have elicited anti-HIV-1 CTL in a significant number of uninfected immunized volunteers. Although we have begun to better understand the specificity and duration of the elicited CTL reactivities, little is still known about the qualitative aspects of these responses. Based on preliminary studies performed in association with recently completed trials, the specific aims of this proposal are designed to test the major hypothesis that vaccine induced anti-HIV CTL responses are qualitatively different from the responses elicited by natural HIV infection. Studies are designed to characterize CTL lines, and ultimately CTL clones, from antigen-specific in vitro stimulation (IVS) cultures with respect to avidity, TcR receptor breadth, as well as cytolytic and non-cytolytic virus suppressive reactivities against autologous lymphocyte target cells infected with primary isolates representing genetically diverse viral clades. Avidity will be measured both by conventional peptide titration as well as peptide/MHC tetramer binding. Tetramers will also be utilized to track epitope-specific CTL and their maturation stage in PBMC from vaccinees. The capacity of canarypox vectors to bias the CTL responses of vaccinees will also be studied at the level of in vitro CTLp amplification, using a panel of different HIV-expressing recombinant vectors in different APC-based antigen-specific stimulation strategies as well as HIV-1-specific activation. Lastly, comparative studies will be performed to determine whether the quality of vaccine- induced CTL reactivities more closely resemble CTL responses that develop during the acute phase of HIV-1 infection, especially in association with structured early treatment interruption before the immune depletion associated with prolonged viral stimulation and infection become manifest. By yielding new insights into the quality of vaccine-induced CTL, these studies could greatly impact the further development of vector-based vaccines.