Antigen stability as a parameter in rational T(CD8+)-targeting vaccine design

描述（由申请人提供）：CD 8 + T细胞（TCD 8+）通过消除病毒感染的细胞（在其表面展示病毒），在限制病毒传播中发挥关键作用 衍生肽与主要组织相容性复合物I类（MHCI）分子的组合。由于许多病毒在感染后数小时内复制，有效的TCD 8+作用取决于病毒蛋白质合成后不久通过仍然知之甚少的机制进行的催化。我们对这些机制的追求揭示了肽在细胞表面展示的开始，因此，肽/MHCI复合物的密度根据抗原的性质而变化很大。因此，不能有效地易位到内质网中的胞质蛋白质和携带信号序列（SS）的蛋白质迅速出现，而有效易位的蛋白质的加工实质上被延迟。此外，未屏蔽的疏水性（例如未束缚的跨膜结构域）构成了足够快速的蛋白质催化的一个驱动因素。基于这些观察结果，我们假设，TCD 8+特异性的表位来源于最迅速降解的蛋白质是最具保护性的，由于中断病毒复制在感染后的早期时间点。巧合的是，我们预测这些相同的蛋白质在引发TCD 8+应答中是较差的，因为交叉呈递的过程-其中抗原从感染的细胞转移到树突细胞-取决于蛋白质的稳定性。通过扩展，我们提出了一种策略，用于开发TCD 8+靶向疫苗，其中包括，首先，鉴定病毒蛋白，在其自然状态下，与快速肽生成相关，因此更有效地清除病毒。在第二步中，抗原被工程化为当由疫苗表达时是稳定的，以优化交叉呈递。这些想法将在体外和体内实验的组合进行测试，使用一组主要由重组鼠痘病毒，一种天然的小鼠病原体和重组腺病毒表达的模型抗原时，异源表达将是必要的。这些实验的结果可能从根本上改变TCD 8+靶向疫苗的合理设计，这一目标与许多人类病原体，包括艾滋病毒，疱疹病毒和流感有关。 公共卫生相关性：免疫系统的细胞毒性T细胞在保护身体免受病毒感染方面起着重要作用，因为它们在新病毒产生之前识别并杀死受感染的细胞。不幸的是，我们仍然不知道如何配制疫苗以优化细胞毒性T细胞的性能;如果我们这样做，针对艾滋病毒和丙型肝炎病毒的疫苗以及针对流感病毒和疱疹病毒的更有效的疫苗可能是可能的。这里提出的研究计划将测试我们开发的一种新的优化策略。

DESCRIPTION (provided by applicant): CD8+ T cells (TCD8+) play critical roles in limiting viral spread through the elimination of virus-infected cells, which display at their surfaces virus derived peptides in combination with major histocompatibility complex class I (MHCI) molecules. Since many viruses replicate within hours after infection, effective TCD8+ action depends upon catabolism of viral proteins shortly after their synthesis through mechanisms that remain poorly understood. Our pursuit of these mechanisms has revealed that onset of peptide display at the cell surface and, consequently, the density of peptide/MHCI complexes varies considerably depending upon properties of the antigen. Thus, cytosolic proteins and signal sequence (SS)-bearing proteins that are not efficiently translocated into the endoplasmic reticulum appear rapidly while the processing of efficiently translocated proteins is substantially delayed. Furthermore, unshielded hydrophobicity (such as an untethered transmembrane domain) constitutes one driver of sufficiently rapid protein catabolism. Based upon these observations, we hypothesize that TCD8+ specific for epitopes derived from the most rapidly degraded proteins are most protective due to interruption of viral replication at early time points followin infection. Paradoxically, we predict that these same proteins are inferior in priming for a TCD8+ response, since the process of cross-presentation - in which antigen is transferred from an infected cell to the dendritic cell - depends upon stability of the protein. By extension, we propose a strategy for development of a TCD8+-targeting vaccine that involves, first, the identification of a viral protein which, in its natural state, is associated with rapid peptide generation and, therefore more efficient viral clearance. In second step, the antigen is engineered to be stable when expressed by a vaccine, in order to optimize cross- presentation. These ideas will be tested with a combination of in vitro and in vivo experiments using a panel of model antigens expressed primarily by recombinant ectromelia virus, a natural mouse pathogen, and recombinant adenovirus when heterologous expression will be necessary. Results of these experiments could fundamentally alter the rational design of TCD8+-targeting vaccines, a goal with relevance to many human pathogens, including HIV, herpesviruses, and influenza. PUBLIC HEALTH RELEVANCE: Cytotoxic T cells of the immune system play an important role in defending the body against viral infections because they recognize and kill infected cells before new viruses can be produced. Unfortunately, we still do not understand how vaccines should be formulated in order to optimize cytotoxic T cell performance; if we did vaccines against HIV and hepatitis C virus and more effective vaccines against influenza viruses and Herpesviruses might be possible. The research plan proposed here will test a novel optimization strategy that we have developed.