Development and specificity of endogeneous tumor-infiltrating regulatory T cells

描述（由申请人提供）：Foxp3 调节性 T 细胞（“Treg”）对于免疫稳态、器官特异性耐受和炎症的调节至关重要。由于Tregs在免疫调节中的核心作用，以及Tregs在各种人类癌症中的普遍存在，许多新兴的治疗策略都集中在疫苗接种的同时调节或消耗Tregs，以刺激有效的抗肿瘤免疫反应。然而，人们对肿瘤浸润性Tregs的发育起源、特异性和原位功能知之甚少。迄今为止，对与肿瘤相关抗原有反应的 Foxp3 Tregs 的研究依赖于模型外源抗原在肿瘤细胞上的异位表达。然而，由于 Treg 识别的天然抗原未知，这些实验系统可能无法正确概括 Treg 生物学的几个基本方面。因此，为了了解肿瘤相关 Treg 的真实性质，开发直接分析内源性抗原特异性 Treg 群体的新系统至关重要。在我们正在进行的工作中，我们已经鉴定出一群自然存在的表达保守 TCR 的 Foxp3 Tregs？通过在患有前列腺癌的小鼠中进行抗原驱动的选择而高度富集这种特异性的细胞（此后，这种特异性的细胞将被称为“RT3”T细胞），并产生表达这种RT3“Treg TCR”的转基因小鼠。该提案的总体目标是了解内源性 RT3 Treg 反应的发展、谱系可塑性和“特异性”。我们的中心假设是，RT3 T 细胞是在胸腺中发育的“天然”Foxp3 Tregs，并且这些 Tregs 对非前列腺癌特有的共享抗原和常见刺激有反应。为了解决这个假设，我们将使用补充方法：a）分析纯化的 TCR 转基因细胞群转移到荷瘤宿主后的发育、分化和稳定性，以及 b）典型 RT3 TCR 的量化？使用“深度”TCR 测序对荷瘤小鼠的内源性 T 细胞亚群进行转录。我们计划通过追求以下三个具体目标来测试我们的中心假设并实现此应用程序的目标。在目标 1 中，我们将检验以下假设：由于 Foxp3 Treg 分化为 Foxp3neg 细胞，Foxp3neg 效应 T 细胞和具有相同 RT3 特异性的 Foxp3 Tregs 在荷瘤小鼠中同时发育。在目标 2 中，我们将检验这样的假设：Foxp3 RT3 Tregs 是在胸腺中发育的“天然”Treg 群体，并且通过抗原驱动的选择在前列腺肿瘤中高度富集。最后，在目标 3 中，我们将检验这样的假设：RT3 Tregs 不是前列腺癌特异性的，而是对共享抗原和常见刺激有反应。这些目标的完成将为调节性 T 细胞的生物学提供基础见解，这将指导旨在通过选择性调节或耗竭肿瘤相关 Tregs 诱导癌症消退的治疗策略的开发。

DESCRIPTION (provided by applicant): Foxp3+ regulatory T cels ("Tregs") are critical for the regulation of immune homeostasis, organ-specific tolerance, and inflammation. Because of the central role of Tregs in immune modulation, and the prevalence of Tregs in various human cancers, many emerging therapeutic strategies have focused on the modulation or depletion of Tregs concomitant with vaccine administration, in an effort to stimulate effective anti-tumor immune responses. However, little is known about the developmental origins, specificity, and in situ function of tumor-infiltrating Tregs. To date, studies of Foxp3+ Tregs reactive to tumor-associated antigens have relied on the ectopic expression of model foreign antigens on tumor cells. However, since the natural antigens recognized by Tregs are unknown, these experimental systems may not correctly recapitulate several fundamental aspects of Treg biology. Therefore, in order to understand the true nature of tumor-associated Tregs, it is paramount to develop novel systems by which to directly analyze endogenous, antigen-specific Treg populations. In our ongoing work, we have identified a population of naturally occurring Foxp3+ Tregs expressing a conserved TCR?? that is highly enriched by antigen-driven selection in mice with prostate cancer (hereafter, cells of this specificity will be referred to as "RT3" T cells), and generated transgenic mice expressing this RT3 "Treg TCR". The overall objective of this proposal is to understand the development, lineage plasticity, and "specificity" of the endogenous RT3 Treg response. It is our central hypothesis that RT3 T cells are "natural" Foxp3+ Tregs that develop in the thymus, and that these Tregs are responsive to shared antigens and common stimuli that are not specific to prostate cancer. To address this hypothesis, we will use complementary approaches: a) analysis of the development, differentiation, and stability of purified TCR transgenic cell populations following transfer into tumor-bearing hosts, and b) quantification of canonical RT3 TCR? transcripts in endogenous T cell subsets from tumor-bearing mice using "deep" TCR sequencing. We plan to test our central hypothesis and accomplish the objectives of this application by pursuing the following three specific aims. In Aim 1, we will test the hypothesis that Foxp3neg effector T cells and Foxp3+ Tregs of the same RT3 specificity develop concurrently in tumor-bearing mice, as a result of Foxp3+ Treg differentiation into Foxp3neg cells. In Aim 2, we will test the hypothesis that Foxp3+ RT3 Tregs are a population of "natural" Tregs that develop in the thymus, and are highly enriched in prostate tumors by antigen-driven selection. Finally, in Aim 3, we will test the hypothesis that RT3 Tregs are not prostate cancer-specific, but are instead responsive to shared antigens and common stimuli. Completion of these aims will provide fundamental insight into the biology of regulatory T cells, which will guide the development of therapeutic strategies aimed at the induction of cancer regression through the selective modulation or depletion of tumor-associated Tregs.