Regulatory T Cell Control of Intestinal Tumorigenesis

肠道肿瘤发生的调节性 T 细胞控制

• 批准号: 9379022
• 负责人: Alfred LM Bothwell
• 金额: $ 12.62万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9379022
• 关键词: APC gene; Acids; Address; Adenomatous Polyposis Coli; Affect; ApcMin/+ mice; Biopsy; Biopsy Specimen; CD4 Positive T Lymphocytes; Cell Proliferation; Cells; Clinic; Colon; Colorectal Cancer; Development; Disease; Disease model; Down-Regulation; Emulsions; FOXP3 gene; Flow Cytometry; Gene Mutation; Human; Immune; Immune system; Immunologic Surveillance; In Vitro; Inflammatory; Inflammatory Bowel Diseases; Inflammatory disease of the intestine; Inherited; Interleukin-10; Interleukin-17; Intestinal Neoplasms; Intestinal Polyps; Intestines; Knock-in Mouse; LGR5 gene; Lead; Malignant Neoplasms; Measures; Mediating; Modeling; Monitor; Mus; Mutation; Pathway interactions; Patients; Polyps; Process; Regulatory T-Lymphocyte; Role; Serum; Signal Pathway; Signal Transduction; Source; Stem cells; Study models; T-Lymphocyte; Techniques; Testing; Therapeutic; Translations; Tumor Expansion; WNT Family Gene; WNT Signaling Pathway; Work; base; cell type; colon cancer patients; cytokine; defined contribution; deoxycholate; experimental study; functional disability; in vitro Assay; in vivo; insight; intestinal homeostasis; mouse development; mouse model; mutant; nanoparticle; prevent; public health relevance; receptor; response; therapeutic development; tumor; tumor microenvironment; tumorigenesis

DESCRIPTION (provided by applicant): Mutations in the Apc gene occur in many cancers but are especially frequent in colorectal cancer (CRC). The ApcMin/+ mouse is a highly studied model of intestinal tumorigenesis since the Apc gene mutation results in dysregulation of the Wnt signaling pathway. Our recent work using IL-17A deficient (KO) ApcMin/+ mice has identified IL-17A as a proinflammatory cytokine affecting tumorigenesis. Introduction of wildtype (WT) Tregs into ApcMin/+ mice regressed tumors markedly while Tregs from ApcMin/+ mice could not, suggesting an important factor from Tregs for tumor regression lacking in ApcMin/+ Tregs. In ApcMin/+ mice, the expression of Gata-3 was decreased in Tregs as well as effector T cells. Our analysis of Tregs and other immune cell types has identified the Wnt antagonist Dikkopf-1(Dkk1) as the most abundant Wnt family gene produced by Tregs. Importantly, IL-17A effectively inhibited Dkk1 expression in ApcMin/+ Tregs but not in WT Tregs. Dkk1 could induce Gata-3 and also IL-10, which was downregulated in ApcMin/+ mice. Dkk1 also stimulated Treg proliferation but did not impair Treg function. The lack of Treg-derived Dkk1 blocked the ability of Treg to suppress inflammatory bowel disease (IBD) in a standard murine IBD model. Based on these results, first we hypothesize that Treg-derived Dkk1 suppresses intestinal inflammation that is fueled by IL-17A under homeostatic conditions. Second, we argue that the loss of Dkk1 expression in ApcMin/+ Treg leads to the loss of IL-10, and also fails to regulate the Wnt pathway controlling ApcMin/+ intestinal stem cell proliferation. Third, we hypothesize that Treg-derived Dkk1 and IL-17A from Th17 cells are crucial regulators of the tumor microenvironment, directly targeting ISCs carrying the ApcMin/+ mutation. Four aims will focus on testing these three hypotheses. First, Dkk-1 deficient Treg will be used in in vivo tumor regression experiments and in in vitro biologic studies to define the contribution of Dkk1 to Treg function. The mechanism that Dkk1 induces Treg proliferation will be studied in three different signaling pathways. Second, the role of IL-17A to downregulate Dkk1 in ApcMin/+ Tregs will be studied by utilizing IL-17 receptor-deficient Foxp3+ ApcMin/+ Tregs in vivo and in vitro. The mechanism which IL-17A and Dkk1 regulates IL-10 will be studied as well. Third, the effects of IL-17A and Dkk1 on intestinal stem cells (ISC) carrying the mutation in the Apc gene will be characterized. Conditional deletion of the IL-17 receptor in ISCs in the ApcMin/+ background will be studied in vivo. We will deliver Dkk1 into the intestine in ApcMin/+ mice, testing the potential for Dkk1 as a therapeutic. Finally, we will extend our findings to human FAP Treg utilizing a Treg/T cell expansion technique from colon biopsies, and compare them with cells from sporadic colon cancer patients and healthy donors. These studies should give important mechanistic insight into the specific inflammatory molecules that can compromise cells of the immune system. This mechanistic insight should provide a rational approach to translation of these results to the clinic.

描述（由申请人提供）：Apc 基因突变发生在许多癌症中，但在结直肠癌 (CRC) 中尤其常见。 ApcMin/+ 小鼠是一种经过深入研究的肠道肿瘤发生模型，因为 Apc 基因突变导致 Wnt 信号通路失调。我们最近使用 IL-17A 缺陷 (KO) ApcMin/+ 小鼠进行的研究已确定 IL-17A 是影响肿瘤发生的促炎细胞因子。将野生型 (WT) Tregs 引入 ApcMin/+ 小鼠可使肿瘤显着消退，而来自 ApcMin/+ 小鼠的 Tregs 则不能，这表明缺乏 ApcMin/+ Tregs 的 Tregs 是肿瘤消退的重要因素。在 ApcMin/+ 小鼠中，Treg 细胞和效应 T 细胞中 Gata-3 的表达降低。我们对 Tregs 和其他免疫细胞类型的分析已确定 Wnt 拮抗剂 Dikkopf-1 (Dkk1) 是 Tregs 产生的最丰富的 Wnt 家族基因。重要的是，IL-17A 有效抑制 ApcMin/+ Tregs 中的 Dkk1 表达，但不抑制 WT Tregs 中的 Dkk1 表达。 Dkk1 可以诱导 Gata-3 和 IL-10，后者在 ApcMin/+ 小鼠中下调。 Dkk1 还刺激 Treg 增殖，但不损害 Treg 功能。在标准小鼠 IBD 模型中，Treg 衍生的 Dkk1 的缺乏阻碍了 Treg 抑制炎症性肠病 (IBD) 的能力。基于这些结果，我们首先假设 Treg 衍生的 Dkk1 能够抑制稳态条件下由 IL-17A 引发的肠道炎症。其次，我们认为 ApcMin/+ Treg 中 Dkk1 表达的缺失导致 IL-10 的缺失，并且也无法调节控制 ApcMin/+ 肠干细胞增殖的 Wnt 通路。第三，我们假设来自 Th17 细胞的 Treg 衍生的 Dkk1 和 IL-17A 是肿瘤微环境的关键调节因子，直接靶向携带 ApcMin/+ 突变的 ISC。四个目标将集中于检验这三个假设。首先，Dkk-1缺陷的Treg将用于体内肿瘤消退实验和体外生物学研究，以确定Dkk1对Treg功能的贡献。 Dkk1诱导Treg增殖的机制将通过三种不同的信号通路进行研究。其次，将利用IL-17受体缺陷的Foxp3+ ApcMin/+ Tregs在体内和体外研究IL-17A下调ApcMin/+ Tregs中Dkk1的作用。还将研究IL-17A和Dkk1调节IL-10的机制。第三，将表征IL-17A和Dkk1对携带Apc基因突变的肠干细胞(ISC)的影响。将在体内研究 ApcMin/+ 背景下 ISC 中 IL-17 受体的条件删除。我们将 Dkk1 递送到 ApcMin/+ 小鼠的肠道中，测试 Dkk1 作为 治疗性的。最后，我们将利用结肠活检中的 Treg/T 细胞扩增技术将我们的研究结果扩展到人类 FAP Treg，并将其与来自散发性结肠癌患者和健康捐赠者的细胞进行比较。这些研究应该能够对可能损害免疫系统细胞的特定炎症分子提供重要的机制见解。这种机制上的见解应该为将这些结果转化为临床提供合理的方法。