Tracking Immune Activation by Stress Proteins In Vivo

含有与应激蛋白相关的多肽的疫苗是已知的最有效的T细胞免疫原。 然而，它们激活免疫细胞的确切机制仍不清楚。为了解决这个问题，我们 将追踪应激融合抗原免疫的小鼠中活化的树突状细胞(DC)和CD8+T细胞 蛋白质HSP70，并确定哪些细胞因子和细胞表面受体是其反应所必需的。 我们的初步研究表明，hspTO融合疫苗可诱导更强、更持久的特异性CD8+T细胞 细胞反应优于单独使用抗原或与其他佐剂联合使用。我们的基本假设是抗原/HSP70 融合是一种有效的疫苗，因为它们刺激DC为CD8+T细胞提供最佳的微环境 细胞激活。这种微环境以高抗原密度和强共刺激作用为特征。 为了测试这一点，我们将在体内跟踪引流淋巴结中DC和CD8+T细胞的反应，以及由此产生的 外周有效应性和记忆性CD8+T淋巴细胞。我们提案的新颖之处包括：1) 定量活化DC提呈的抗原量，2)HSP70融合蛋白对 DC寿命，3)确定几个已报道的HSP70受体中哪些是必需的或足够的 在体内向DC传递激活信号，以及4)确定共刺激和细胞因子所需的 我们的疫苗的深远效果是具有CD8+T细胞记忆。 这些实验的结果将使我们能够理性地操纵应激蛋白诱导的反应 最大限度地将其用于癌症治疗的最终目标。

Vaccines containing peptides linked to stress proteins are the most potent known T cell immunogens. However, the precise mechanisms by which they activate immune cells remain unclear. To address this, we will track activated dendritic cells (DCs) and CD8+ T cells in mice immunized with antigens fused to the stress protein hsp70, and determine which cytokines and cell surface receptors are required for their responses. Our preliminary studies reveal hspTO fusion vaccines elicit much more potent and durable specific CD8+ T cell responses than antigen alone or with other adjuvants. Our underlying hypothesis is that antigen/hsp70 fusions are potent vaccines because they stimulate DCs to provide an optimal microenvironment for CD8+ T cell activation. This microenvironment is defined by high antigen density and strong costimulation. To test this, we will track in vivo DC and CD8+ T cell responses in draining lymph nodes, and the resultant effector and memory CD8+ T lymphocytes in the periphery. Novel aspects of our proposal include: 1) quantifying the amount of antigen presented by activated DCs, 2) the effect hsp70 fusion proteins have on DC lifespan, 3) determining which of the several reported hsp70-receptors are necessary or sufficient for delivering activation signals to DCs in vivo, and 4) determining the costimulatory and cytokines required for the profound effect our vaccine has CD8+ T cell memory. The results of these experiments will allow us to manipulate stress protein-induced responses rationally with the ultimate goal of maximizing their use for cancer therapy.