COBRE: DMS: MECHANISMS OF CONCOMITANT TUMOR IMMUNITY

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。伴发肿瘤免疫被定义为携带进展性原发肿瘤的宿主对继发肿瘤的排斥反应。我们最近的研究表明，如果在肿瘤进展过程中缺乏CD4+ CD25+调节性T细胞，则存在低免疫原性黑色素瘤的伴随免疫。这种保护性抗肿瘤免疫是由CD8+ T细胞介导的，并且可以通过实验诱导CD4+ T细胞的完全耗尽。低免疫原性黑色素瘤的伴随免疫也以破坏对肿瘤表达的自身抗原的免疫耐受为特征。除了直接消耗CD4+调节性T细胞外，我们还发现了其他诱导启动伴随免疫的治疗方法。其中包括(1)化疗性淋巴细胞清除，(2)通过TNFR家族成员GITR刺激，(3)GM-CSF的局部表达。尽管这些治疗方法的作用机制尚不清楚，但每种治疗方法都会引起原发肿瘤宿主对继发性肿瘤的排斥反应。淋巴消融术引起所有淋巴亚群的稳态增殖，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CD8 T细胞的过度活化有关。GM-CSF激活的dc可直接作用于调节性T细胞，或通过直接刺激CD8 T细胞克服局部抑制。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，以肿瘤表达的自身抗原的免疫识别为特征，可以通过消除CD4调节性T细胞和/或通过驱动CD8 T细胞在进行性肿瘤宿主中的扩增来诱导。特异性目的1：确定在淋巴缺失宿主中诱导伴随免疫的机制。用环磷酰胺治疗小鼠可导致大量淋巴细胞耗竭和强大的伴随免疫。这是高度时间表依赖性的，当原发肿瘤生长和淋巴细胞室重新聚集同时发生时，效果最佳。这一观察结果，加上之前关于CD8稳态扩增可导致肿瘤免疫识别的报道，表明CD8的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消耗调节性T细胞和诱导CD8 T细胞稳态来引发伴随免疫。特异性目的2：确定GITR对调节性T细胞和/或CD8 T细胞的刺激是否会诱导伴随免疫。DTA-1单克隆抗体对黑素瘤宿主抗GITR负载的强效伴随免疫。最初的描述是DTA-1通过结合调节性T细胞和阻断抑制功能发挥作用，但现在已知该抗体也结合表达gitr的活化CD4和CD8 T细胞群，从而诱导过度活化和增殖。我们假设DTA-1通过其对调节性T细胞和CD8 T细胞的作用诱导伴随免疫。特异性目标3：确定肿瘤GM-CSF的产生是否通过激活树突状细胞交叉原质CD8 T细胞诱导伴随免疫，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可能通过一种或多种途径起作用。首先，它们可能通过交叉呈递肿瘤抗原直接促进伴随免疫，从而扩大CD8 T细胞在引流淋巴结中的效应。其次，gm - csf募集的树突状细胞可能通过阻断调节性T细胞功能直接起作用。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递呈，并且这种组合诱导了伴随免疫的启动。

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.