Evaluating Relevant Vaccine Epitopes Displayed on HIV-Infected Cells

描述（由申请人提供）：艾滋病毒/艾滋病流行病继续在全球范围内肆虐，已成为有史以来最具破坏性的流行病之一。尽管在延长生命的抗逆转录病毒疗法方面取得了巨大进步，但很可能阻止艾滋病毒/艾滋病流行的唯一途径是开发有效的艾滋病毒疫苗。不幸的是，传统的艾滋病毒疫苗提供了很少或根本没有保护，这突出表明需要更好地了解艾滋病毒特异性免疫反应和新的艾滋病毒疫苗方法。研究已经证明CD8+ T细胞在控制HIV感染中的重要作用，但对于哪些病毒蛋白负责引发有效的CD8+ T细胞反应，目前还没有明确的共识。特别是，在HIV感染过程中，被感染的CD4+ T细胞所显示的T细胞特异性表位的时间、大小和相关性缺乏信息，然而，当考虑将特异性HIV蛋白作为基于疫苗的方法的成分或作为免疫治疗的靶标时，这些参数至关重要。拟议研究的目标是开发新的试剂和方法，可在普通实验室环境中进行，以评估感染细胞上的艾滋病毒抗原呈递谱，并将这些参数与艾滋病毒感染过程中早期发生的时间事件联系起来。我们一直在开发新的基于T细胞受体（TCR）的试剂和简单的方法，以更直接地定量和可视化病变细胞和组织（包括hiv感染细胞）上的肽抗原呈递。我们已经证明，可溶性单链TCR （scTCR）试剂可用于评估内源性抗原在肿瘤细胞上的递呈，最近又扩展了这些研究，以产生高亲和力的可溶性scTCR试剂，能够识别HIV肽表位及其已知的各自的表位变体。通过流式细胞术发现这些试剂能够识别HIV感染的CD4 T细胞上显示的HIV肽/HLA复合物，验证了该方法用于评估HIV抗原呈递谱的可行性。在此建议下，我们打算扩大HIV抗原识别特异性的试剂组合，并进一步优化分析HIV感染T细胞中抗原呈递水平和动力学的方法。为实现该项目的目标，将进行以下具体目标：1)生成并表征至少20种不同的HIV肽/HLA复合物特异性的可溶性scTCR试剂，2)利用这些试剂定量HIV感染细胞的HIV肽抗原呈递。这些研究的结果将有助于确定哪些特定的HIV表位可能在感染早期产生保护性CD8 T细胞反应中发挥作用，从而有助于为预防性疫苗选择最相关的HIV靶点，这是本项目的最终目标。与此同时，我们将迅速将hiv特异性TCR试剂商业化，作为我们现有研究试剂组合的一部分，以允许它们在本提案中考虑的研究之外使用。

DESCRIPTION (provided by applicant): The HIV/AIDS pandemic continues to wreak havoc on global scale and has become one of the most destructive pandemics in recorded history. Despite the tremendous improvements in life-extending antiretroviral therapy, it is likely that the only way by which the HIV/AIDS pandemic can be halted is through the development of an effective HIV vaccine. Unfortunately, conventional vaccines against HIV have provided little or no protection, highlighting the need for a better understanding of HIV-specific immune responses and for novel HIV vaccine approaches. Studies have demonstrated the important role of CD8+ T cells in controlling HIV infections, yet there is no clear consensus as to which viral proteins are responsible for eliciting effective CD8+ T cell responses. Particularly, information is lacking on the timing, magnitude and relevance of T cell- specific epitopes displayed by the infected CD4+ T cells during the course of HIV infection, yet these parameters are of crucial importance when considering specific HIV proteins as components in vaccine-based approaches or as targets for immunotherapy. The goal of the proposed research is to develop new reagents and methods, which can be performed in an ordinary laboratory setting, to assess the HIV antigen presentation profile on infected cells and relate these parameters with the temporal events occurring early during the course of HIV infection. We have been developing novel T cell receptor (TCR)-based reagents and simple methods to more directly quantitate and visualize peptide antigen presentation on diseased cells and tissues, including HIV-infected cells. We have shown that soluble single-chain TCR (scTCR) reagents can be used to evaluate endogenous antigen presentation on tumor cells and recently have extended these studies to generate high- affinity soluble scTCR reagents that are capable of recognizing HIV peptide epitopes as well as their known respective epitope variants. These reagents were found to be capable of recognizing HIV peptide/HLA complexes displayed on HIV-infected CD4 T cells by flow cytometry, verifying the feasibility of this approach for evaluating the HIV antigen presentation profile. Under this proposal, we intend to expand the reagent portfolio of HIV antigen recognition specificities and further optimize methods for analyzing antigen presentation levels and kinetics in HIV-infected T cells. The following specific aims will be conducted to achieve the goals of this project: 1) Generate and characterize a panel of soluble scTCR reagents specific for at least 20 different well-characterized HIV peptide/HLA complexes and 2) utilize these reagents to quantitate HIV peptide antigen presentation by HIV-infected cells. The results of these studies will help establish which specific HIV epitopes may play a role in generating protective CD8 T cell responses early in infection and thus should aid in the selection of the most relevant HIV targets for preventative vaccines, an ultimate goal of this project. In parallel, we will rapidly commercialize the HIV-specific TCR reagents as part of our existing research reagent portfolio to allow their use beyond the studies contemplated in this proposal. PUBLIC HEALTH RELEVANCE: Current information is lacking on the timing, magnitude and relevance of HIV antigens displayed by patient's T cells during the course of viral infection, yet these parameters are of crucial importance when considering specific HIV proteins as components in vaccine-based approaches. The goal of the proposed research is to develop approaches to assess the HIV antigen presentation profile on infected cells and relate these parameters with the temporal events occurring early during the course of HIV infection. Ultimately, these studies will help establish which specific HIV antigens play a role in generating protective CD8+ T cell responses early in infection and thus should aid in the selection of the most relevant HIV targets for preventative vaccines.