Tolerance mechanisms regulating the complete HER-2/neu CD+8 T cell repertoire

描述(由申请人提供)：这项建议的广泛目标是了解在不耐受和耐受小鼠中调节低亲和力和高亲和力肿瘤抗原特异性T细胞激活状态和功能的分子机制。识别这些机制是开发方法的第一步，这种方法可以更有效地激活具有一系列癌症抗原特异性的T细胞。许多病毒免疫的成功部分是由于在健康宿主中诱导了病毒特异性的、高亲和力的CD8+T细胞库和足够的记忆T细胞室。相比之下，在携带癌症的宿主中成功接种疫苗需要绕过多种免疫耐受机制。许多已知的肿瘤抗原是过度表达的组织特异性抗原。必须存在T细胞耐受机制，以防止自身抗原的自身免疫。这些建立和维持自我耐受的机制可能是癌症患者体内T细胞缺乏有效性的原因之一。耐受诱导的含义是，癌症特异性高亲和力T细胞群被删除或被抑制，癌症疫苗必须打破无能T细胞的耐受性，或激活低亲和力T细胞群，基于其较弱的反应性，可能逃脱主动耐受诱导。直到最近，还没有工具来了解高亲和力和低亲和力的癌症特异性CD8+T细胞在荷癌宿主中的命运。自发性乳腺肿瘤的HER-2/neu转基因(neu-N)小鼠模型提供了使用天然肿瘤抗原HER-2/neu(Neu)评估这一问题的机会。在免疫亲本FVB/N小鼠中，大多数CD8+T细胞具有高亲和力，并针对免疫优势抗原RNEU420-429。相比之下，在接种了neu-N的小鼠中，大多数T细胞反应都很弱，亲和力低，并且针对多个表位。为了了解低亲和力和高亲和力CD8+T细胞激活和功能的调节机制，我们建立了高亲和力和低亲和力RNEU420-429特异性TCR转基因小鼠。在目标1中，我们将比较和表征高亲和力和低亲和力的RNEU420-429特异性CD8+T细胞在体内的命运，并确定调节性T细胞在体内控制激活和记忆的高亲和力和低亲和力T细胞的功能中所起的作用。在目标2中，我们将评估microRNA(MiRNA)作为T细胞功能转录后调节因子在高亲和力和低亲和力RNEU420-429特异性CD8+T细胞中的作用。在目标3中，我们将评估高亲和力和低亲和力RNEU420-429特异性CD8+TCR转基因T细胞T细胞信号通路的变化。公共卫生相关性：这项建议的广泛目标是了解在不耐受和耐受小鼠中调节低亲和力和高亲和力肿瘤抗原特异性T细胞激活状态和功能的分子机制。具体地说，我们将评估一些可能对低亲和力T细胞和高亲和力T细胞进行差异调节的机制，包括：调节性T细胞、T细胞信号通路和转录后调节。

DESCRIPTION (provided by applicant): The broad objective of this proposal is to understand the molecular mechanisms that regulate the activation status and function of low versus high avidity tumor-antigen specific T cells in non-tolerized and tolerized mice. Identification of these mechanisms is the first step toward developing approaches that can allow for more effective activation of T cells with a range of avidities specific for cancer antigens. The success of immunization against many viruses is in part due to the induction of a virus specific, high avidity CD8+ T cell repertoire with a sufficient memory T cell compartment in healthy hosts. In contrast, successful vaccination in cancer bearing hosts requires bypassing multiple mechanisms of immune tolerance. Many of the known tumor antigens are over-expressed tissue- specific antigens. Mechanisms of T cell tolerance must exist to prevent autoimmunity against self- antigens. These same mechanisms that establish and maintain self-tolerance are likely contributors to the lack of effectiveness of T cells in vivo that is often observed in cancer patients. The implications of tolerance induction are that cancer-specific high avidity T cell populations are deleted or suppressed, and that cancer vaccines must either break tolerance in anergic T cells or activate populations of low avidity T cells which, based on their weak reactivity, may escape active tolerance induction. Until recently, the tools to understand the fate of the high avidity and low avidity cancer specific CD8+ T cell repertoires in cancer bearing hosts were not available. The HER-2/neu transgenic (neu-N) mouse model of spontaneous mammary tumors provides the opportunity to evaluate this issue using a natural tumor antigen, HER-2/neu (neu). In vaccinated parental FVB/N mice, the majority of CD8+ T cells are of high avidity and directed against an immunodominant antigen, RNEU420-429. In contrast, in vaccinated neu-N mice, most T cell responses are weak, of low avidity, and directed against multiple epitopes. To understand the mechanisms that regulate the activation and function of CD8+ T cells of both low and high avidity, we have developed high and low avidity RNEU420-429-specific TCR transgenic mice. In aim 1 we will compare and characterize the fate of the high avidity versus the low avidity RNEU420-429-specific CD8+ T cells in vivo, and determine the role regulatory T cells (Tregs) play in controlling the function of activated and memory high avidity versus low avidity T cells in vivo. In aim 2, we will evaluate the role of microRNA (miRNA) as post- transcriptional regulators of T cell function in the high avidity versus low avidity RNEU420-429-specific CD8+ T cells. In aim 3, we will evaluate the high avidity versus low avidity RNEU420-429-specific CD8+ TCR transgenic T cells for changes in T cell signaling pathways. PUBLIC HEALTH RELEVANCE: The broad objective of this proposal is to understand the molecular mechanisms that regulate the activation status and function of low versus high avidity tumor-antigen specific T cells in non-tolerized and tolerized mice. Specifically, we will evaluate a number of mechanisms that may differentially regulate low versus high avidity T cells including: regulatory T cells, T cell signaling pathways, and post-transcriptional regulation.