COBRE: DMS: MECHANISMS OF CONCOMITANT TUMOR IMMUNITY

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。伴随的肿瘤免疫被定义为对携带进行性原发肿瘤的宿主的继发肿瘤的排斥。我们最近发现，免疫原性差的黑色素瘤如果在肿瘤进展过程中缺乏CD4+CD25+调节性T细胞，则存在伴随免疫。这种保护性的抗肿瘤免疫是由CD8+T细胞介导的，可以通过实验完全耗尽CD4+T细胞来诱导。免疫原性低的黑色素瘤的伴随免疫也以打破对肿瘤表达的自身抗原的免疫耐受为特征。除了直接耗尽CD4+调节性T细胞外，我们还发现了其他诱导伴随免疫启动的治疗方法。其中包括(1)化疗淋巴滤除，(2)通过TNFR家族成员GITR刺激，以及(3)局部表达GM-CSF。这些疗法中的每一种都会在原发荷瘤宿主中引起继发性肿瘤的排斥反应，尽管它们的作用机制尚不清楚。淋巴消融导致所有淋巴亚群的动态平衡增殖，但也可能对调节性T细胞有选择性作用。同样，GITR的刺激也与抑制功能的阻断和CD8T细胞的过度激活有关。GM-CSF激活的DC可以直接作用于调节性T细胞，也可以通过直接刺激CD8T细胞来克服局部抑制。解开这些机制将为打破对低免疫原性肿瘤的免疫耐受的方法提供坚实的理解。我们假设，伴随免疫，其特征是对肿瘤表达的自身抗原的免疫识别，可以通过消除CD4调节性T细胞和/或通过驱动进展性肿瘤宿主的CD8T细胞的扩张来诱导。具体目标1：确定淋巴耗竭宿主诱导伴随免疫的机制。用环磷酰胺治疗小鼠会导致大量淋巴枯竭和强大的伴随免疫。这是高度依赖于治疗方案的，当原发肿瘤生长和淋巴间隔重新聚集同时发生时，可获得最佳效果。这一观察结果与之前一篇关于CD8稳态扩张可导致肿瘤免疫识别的报告一起表明，CD8快速增殖有助于伴随免疫。然而，调节性T细胞也可能发挥作用。我们假设环磷酰胺通过耗尽调节性T细胞和诱导CD8T细胞稳态来诱导伴随免疫。特异性目标2：确定GITR刺激调节性T细胞和/或CD8T细胞是否诱导伴随免疫。用刺激性抗GITR的DTA-1单抗治疗黑色素瘤宿主可获得有效的伴随免疫。DTA-1最初是通过与调节性T细胞结合和阻断抑制功能发挥作用的，但现在已知该抗体还与表达GITR的活化的CD4和CD8T细胞群结合，从而诱导过度激活和增殖。我们推测DTA-1通过对调节性T细胞和CD8T细胞的作用而诱导伴随免疫。具体目标3：明确肿瘤GM-CSF的产生是否通过激活树突状细胞与CD8 T细胞交叉激活来诱导伴随免疫，并表征这些树突状细胞对调节性T细胞的作用。肿瘤GM-CSF表达的主要作用是树突状细胞(DC)的局部募集。DC可能通过一条或多条途径发挥作用。首先，它们可能通过交叉递呈肿瘤抗原直接促进伴随免疫，从而扩大引流淋巴结中的CD8T细胞效应器。其次，GM-CSF募集的树突状细胞可能通过阻断调节性T细胞功能直接发挥作用。我们假设，在携带B16-GMCSF肿瘤的小鼠中，树突状细胞负责局部阻断调节性T细胞功能，以及有效的肿瘤抗原交叉呈递，并且这种结合诱导伴随免疫的启动。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Concomitant tumor immunity is defined as the rejection of a secondary tumor in a host bearing a progressive primary tumor. We have recently shown that concomitant immunity exists for poorly-immunogenic melanoma if CD4+ CD25+ regulatory T cells are absent during tumor progression. This protective anti-tumor immunity is mediated by CD8+ T cells, and can be induced experimentally by total depletion of CD4+ T cells. Concomitant immunity for poorly-immunogenic melanoma is also characterized by the breaking of immunological tolerance to tumor-expressed self antigens. In addition to direct depletion of CD4+ regulatory T cells, we have discovered other treatments that induce priming of concomitant immunity. Among these are (1) chemotherapeutic lymphodepletion, (2) stimulation through the TNFR family member GITR, and (3) local expression of GM-CSF. Each of these treatments elicits rejection of secondary tumors in primary tumor-bearing hosts, although the mechanisms by which they function remain unknown. Lymphoablation causes homeostatic proliferation of all lymphoid subsets, but may also have selective effects on regulatory T cells. Likewise, stimulation of GITR has been implicated in both blockade of suppressor function and hyperactivation of CD8 T cells. GM-CSF activated DCs may act directly on regulatory T cells, or may overcome local suppression by direct stimulation of CD8 T cells. Unraveling these mechanisms will provide a solid understanding of methods for breaking immunological tolerance to poorly-immunogenic tumors. We hypothesize that concomitant immunity, characterized by immune recognition of tumor-expressed self antigens, can be induced by elimination of CD4 regulatory T cells and/or by driving expansion of CD8 T cells in hosts with progressive tumors. Specific Aim 1: Determine the mechanism for induction of concomitant immunity in lymphodepleted hosts. Treatment of mice with cyclophosphamide results in massive lymphodepletion and potent concomitant immunity. This is highly schedule dependent, with optimal effects achieved when primary tumor growth and re-population of lymphoid compartments occur simultaneously. This observation, together with a previous report that CD8 homeostatic expansion can lead to immune recognition of tumors, suggests that rapid CD8 proliferation contributes to concomitant immunity. However regulatory T cells are also likely to play a role. We hypothesize that cyclophosphamide elicit concomitant immunity through both depletion of regulatory T cells and the induction of CD8 T cell homeostasis. Specific Aim 2: Determine if concomitant immunity is induced by stimulation of GITR on regulatory T cells and/or on CD8 T cells. Melanoma-bearing hosts treated with stimulatory DTA-1 monoclonal antibody to GITR mount potent concomitant immunity. It was originally described that DTA-1 functions by binding to regulatory T cells and blocking suppressive function, but it is now known that this antibody also binds GITR-expressing activated CD4 and CD8 T cell populations, thereby inducing hyperactivation and proliferation. We hypothesize that DTA-1 induces concomitant immunity through its effects on regulatory T cells and CD8 T cells. Specific Aim 3: Define if tumor GM-CSF production induces concomitant immunity by activating dendritic cells to cross-prime CD8 T cells, and characterize the action of these dendritic cells on regulatory T cells. The predominant effect of tumor GM-CSF expression is the local recruitment of dendritic cells (DC's). DC's may function through one or more pathways. First, they may directly contribute to concomitant immunity by cross-presenting tumor antigens, thus expanding CD8 T cell effectors in draining lymph nodes. Secondly, GM-CSF-recruited dendritic cells may act directly by blocking regulatory T cell function. We hypothesize that, in mice bearing B16-GMCSF tumors, dendritic cells are responsible for local blockade of regulatory T cell function, as well as efficient tumor antigen cross-presentation, and that this combination induces priming of concomitant immunity.