Impact of TCR-pMHC affinity on prostate-specific CD4+ T cell development and function

项目摘要/摘要 Foxp3+CD4+调节性T细胞(Treg细胞)对自身免疫的预防和调节至关重要 抗肿瘤免疫，部分通过抑制传统T细胞的激活和功能来发挥作用。特雷格 细胞的许多生物学方面都依赖于T细胞受体(TCR)的刺激，包括胸腺发育， 外周分化，抑制功能。在这一点上，TCR与自体肽的结合亲和力 与MHC-II类分子复合(自身pMHCII)被认为是Treg细胞的主要决定因素 发展和功能。现有的胸腺选择模型假设TCR-pMHCII中间亲和力- 在积极选择和消极选择之间的阈值-促进最佳胸腺树突状细胞(TTreg)发育， 允许Treg细胞克隆分化，同时避免克隆删除。此外，有人建议， Treg细胞对自身pMHC-II配体可能比Foxp3neg常规的CD4+T(Tconv)细胞更敏感。 同样的特异性，可能有助于Treg介导的外周抑制。然而，这些 提出的原则在很大程度上源于对转基因表达的外源模型的CD4+T细胞反应的研究 抗原。由于这种工程抗原的表达模式、配体密度和TCR-pMHCII亲和力 系统可能不会反映内源性自身抗原的系统，目前尚不清楚 亲和力模型适用于对天然自身配体有反应的Treg细胞克隆。为了解决这个问题，我们有 确定了一组对同一内源性组织表现出不同敏感性的CD4+T细胞克隆 抗原(TRA)，Taf3646-658(C4多肽)。C4是一种未经修饰的I-Ab限制性前列腺自体多肽，可驱动 典型Treg细胞克隆MJ23的胸腺分化。在其他工作中，我们也证明了 在癌基因驱动的前列腺癌中，单克隆MJ23 Treg细胞高度富含，表明C4 多肽是一种天然存在的肿瘤相关抗原。我们使用C4/I-Ab四聚体的新数据表明 在前列腺癌荷瘤小鼠中，C4反应性Treg细胞和Tconv细胞共存于肿瘤引流淋巴中 结节，但只有前者在肿瘤浸润性病变中被检测到。TCR-pMHCII亲和力是否有助于 Tconv细胞被排除在前列腺癌之外仍不清楚。因此，这项建议旨在界定TCR的作用-- PMHCII亲和力在胸腺和外周血中对天然TRA和肿瘤反应的CD4+T细胞的选择 相关抗原C4。目标1中的研究将提供第一个证据来测试T细胞的“TCR亲和力”模型 细胞克隆对一种引导tTreg细胞分化的天然自体肽起反应，提供了一个独特的机会 验证和/或颠覆现有的胸腺选择范例。在Aim 2小鼠身上的研究将产生独特的 对驱动TRA反应性Treg和Tconv细胞外周选择的机械性见解，以及 前者在肿瘤微环境中的选择性浓缩。总体而言，这项工作将利用 新的和生理相关的TCR-配体系统，以了解TCR-pMHCII亲和力对 建立和加强免疫耐受的基本机制。

PROJECT SUMMARY/ABSTRACT Foxp3+ CD4+ regulatory T (Treg) cells are critical for the prevention of autoimmunity and the regulation of antitumor immunity, functioning in part by suppressing the activation and function of conventional T cells. Treg cells depend on T cell receptor (TCR) stimulation for many facets of their biology, including thymic development, peripheral differentiation, and suppressive function. In this regard, the affinity of TCR binding to self-peptides complexed with MHC class II molecules (self-pMHCII) is thought to be a primary determinant of Treg cell development and function. Existing models of thymic selection posit that intermediate TCR-pMHCII affinities – at the threshold between positive and negative selection – promote optimal thymic Treg (tTreg) development, allowing Treg cell clones to differentiate while also evading clonal deletion. In addition, it has been suggested that Treg cells may be more sensitive to self-pMHC-II ligand than Foxp3neg conventional CD4+ T (Tconv) cells of the same specificity, potentially contributing to Treg-mediated suppression in the periphery. However, these proposed principles largely stem from the study of CD4+ T cells reactive to transgene-expressed foreign model antigens. As the expression patterns, ligand densities, and TCR-pMHCII affinities of such engineered antigen systems may not reflect those of endogenous self-antigens, it remains unclear whether the predictions of the affinity model apply to Treg cell clones reactive to natural self-ligands. To address this question, we have identified a panel of CD4+ T cell clones exhibiting varying sensitivities to the same endogenous tissue-restricted antigen (TRA), Tcaf3646-658 (“C4” peptide). C4 is an unmodified I-Ab restricted prostatic self-peptide that drives the thymic differentiation of a canonical Treg cell clone, MJ23. In other work, we also demonstrated that monoclonal MJ23 Treg cells are highly enriched in oncogene-driven prostate tumors, indicating that the C4 peptide is a naturally occurring tumor-associated antigen. Our new data using C4/I-Ab tetramers demonstrate that, in prostate-tumor-bearing mice, C4-reactive Treg and Tconv cells coexist in the tumor-draining lymph nodes, yet only the former is detected within the tumor infiltrate. Whether TCR-pMHCII affinity contributes to Tconv cell exclusion from prostate tumors remains unclear. Thus, this proposal seeks to define the role of TCR- pMHCII affinity in the thymic and peripheral selection of CD4+ T cells reactive to the natural TRA and tumor- associated antigen C4. The studies in Aim 1 will provide the first evidence testing the "TCR affinity" model for T cell clones reactive to a natural self-peptide that directs tTreg cell differentiation, offering a unique opportunity to validate and/or upend existing paradigms of thymic selection. The studies in Aim 2 mice will yield unique mechanistic insights into the factors driving the peripheral selection of TRA-reactive Treg and Tconv cells, and the selective enrichment of the former into the tumor microenvironment. Collectively, this work will leverage a novel and physiologically relevant TCR-ligand system to understand the impact of TCR-pMHCII affinity on the fundamental mechanisms by which immune tolerance is established and enforced.