CCL11 as a New Therapeutic Target for Colitis and Colon Cancer

在160多万美国人中，炎症性肠病(IBD)会导致严重的发病率 并可发展为结肠癌。IBD有两种主要形式：溃疡性结肠炎(UC)和克罗恩病 疾病，以及相关的异常免疫反应继续进行调查，目的是 发现新的治疗方法或干预途径。我们之前已经表明，在一个预期的 收集的成年UC患者队列中，CCL11(也称为嗜酸性粒细胞趋化因子-1)是嗜酸性粒细胞的趋化剂 在所有疾病严重程度的42种细胞因子中，只有分析物在血清和组织中增加 与非IBD对照组患者比较，评估趋化因子。我们还观察到了显著增加的 UC患者与对照组的结肠组织中的嗜酸性粒细胞，以及嗜酸性粒细胞计数与组织CCL11的相关性 级别。研究表明，CCL11基因缺陷小鼠在急性硫酸葡聚糖溶液中可以预防结肠炎。 钠(DSS)诱导的结肠炎模型。然而，在缺乏嗜酸性粒细胞的小鼠身上进行的急性结肠炎研究一直是 好坏参半，说明它不只是明显减少了组织中的嗜酸性粒细胞浸润而导致好转。我们有 现在确定：1)CCL11 mRNA在分离的结肠上皮细胞和分离的板层中表达 固有细胞对慢性DSS和偶氮甲烷(AOM)-DSS结肠炎的反应 致癌(CAC)模型；2)上皮细胞和巨噬细胞表达CCL11受体；3)治疗 抗CCL11抗体在结肠炎的损伤和修复模型中具有保护作用；4)CCL11-/-小鼠表现出 AOM-DSS模型中肿瘤数量、肿瘤负荷和结肠嗜酸性粒细胞减少；5)骨髓 移植研究表明，作为造血和上皮来源的CCL11的关键介质的丧失 在AOM-DSS模型中看到的保护；6)在APC突变的小鼠肿瘤中CCL11的表达增加；以及 7)与野生型小鼠相比，CCL11-/-小鼠的粪便微生物组在基线时发生了改变。在这项提案中，我们 将剖析CCL11在调节上皮损伤和修复以及免疫反应中的作用 慢性结肠炎、CAC和散发性/基因介导的结肠癌。我们的假设是：CCL11是一个关键 结肠炎、上皮功能障碍和致癌风险的介质。最重要的是 目的是对阻断CCL11信号在两种疾病的治疗中的潜在好处建立新的见解 结肠炎与结肠癌的预防。具体目标是：1.确定其作用机制 CCL11缺失对结肠炎相关性癌症的保护作用。CCL11-/-小鼠在AOM中表现出保护作用- DSS模型与组织细胞因子水平改变和组织嗜酸性粒细胞减少相关。然而，急性 对嗜酸性粒细胞缺陷小鼠的结肠炎研究表明，这可能是一种不依赖于嗜酸性粒细胞的效应。WT，Ccl11fl/fl， CCL11∆MYE、CCL11∆GIepi和CCL11-/-暴露于AOM-DSS的小鼠将被用来检验这一假设 保护作用是由于嗜酸性粒细胞和/或上皮功能以外的免疫细胞群的改变所致 当CCL11通过评估肿瘤数量和大小、结肠免疫细胞组成/功能而丢失时，以及 上皮性功能。2.确定CCL11是否也是散发性/基因介导的结肠的修饰物 肿瘤发生学。在小鼠模型中，CCL11的缺失对CAC具有保护作用，并且CCL11的表达在 APC突变肿瘤。CDX2P-CreERT2Apcfl/fl和CDX2P-CreerT2Apcfl/flCcl11-/-小鼠发生结肠癌 将被用来检验CCL11缺失将通过减少肿瘤而阻止肿瘤发展的假设- 通过评估促进炎症：肿瘤数量和大小，结肠免疫细胞组成/功能，以及 上皮性功能。3.确定使用中和抗体靶向CCL11信号是否具有保护作用 与结肠癌的发生有关。暴露于AOM-DSS和CDX2P-CreERT2Apcfl/fl小鼠的WT小鼠将得到治疗 用CCL11中和抗体(或同型对照)来建立对：肿瘤数量和大小的影响， 结肠免疫细胞组成/功能和上皮功能。这些研究将提供新的机制 支持在结肠炎和结肠癌中使用基于抑制CCL11的治疗的见解。

With more than 1.6 million Americans affected, inflammatory bowel disease (IBD) causes significant morbidity and can progress to colon cancer. There are two main forms of IBD: ulcerative colitis (UC) and Crohn's disease, and the associated abnormal immune response continues to be investigated with the goal of discovering new therapeutics or avenues for intervention. We have previously shown that in a prospectively collected cohort of adult UC patients, CCL11 (also called eotaxin-1), a chemoattractant for eosinophils, was the only analyte increased in both serum and tissue at all levels of disease severity out of 42 cytokines and chemokines assessed, compared to non-IBD control patients. We also observed a significant increase in colonic tissue eosinophils in UC patients vs. controls, and eosinophil counts correlated with tissue CCL11 levels. It has been shown that Ccl11-deficient mice were protected from colitis in an acute dextran sulfate sodium (DSS)-induced colitis model. However, acute colitis studies in mice lacking eosinophils have been mixed, suggesting it is not clearly just decreased tissue eosinophil infiltration leading to improvement. We have now determined that 1) Ccl11 mRNA is expressed in isolated colonic epithelial cells and isolated lamina propria cells in response to the chronic DSS and the azoxymethane (AOM)-DSS colitis-associated carcinogenesis (CAC) models; 2) epithelial cells and macrophages express receptors for CCL11; 3) treatment with an anti-CCL11 antibody leads to protection in an injury and repair model of colitis; 4) Ccl11–/– mice exhibit decreased tumor number, tumor burden, and colonic eosinophils in the AOM-DSS model; 5) bone marrow transplant studies point to loss of both hematopoietic and epithelial-derived CCL11 as key mediators of the protection seen in the AOM-DSS model; 6) CCL11 expression is increased in Apc mutant mouse tumors; and 7) the fecal microbiome is altered in Ccl11–/– mice compared to wild-type mice at baseline. In this proposal, we will dissect the role of CCL11 in modulating epithelial injury and repair, and immune responses in models of chronic colitis, CAC, and sporadic/genetically-mediated colon cancer. Our Hypothesis is: CCL11 is a key mediator of colonic inflammation, epithelial dysfunction, and risk for carcinogenesis. The overarching goal is to establish new insights into the potential benefits of blocking CCL11 signaling in both the treatment of colitis and prevention of colon carcinogenesis. The Specific Aims are: 1. To determine the mechanism of protection in colitis-associated cancer with loss of CCL11. Ccl11–/– mice exhibit protection in the AOM- DSS model associated with altered tissue cytokine levels and decreased tissue eosinophils. However, acute colitis studies in eosinophil-deficient mice suggest this may be an eosinophil independent effect. WT, Ccl11fl/fl, Ccl11∆mye, Ccl11∆GIepi, and Ccl11–/– mice exposed to AOM-DSS will be used to test the hypothesis that protective effects are due to altered immune cell populations other than eosinophils and/or epithelial function when CCL11 is lost by assessing tumor number and size, colonic immune cell composition/function, and epithelial function. 2. To determine if CCL11 is also a modifier in sporadic/genetically-mediated colon tumorigenesis. Loss of CCL11 protects from CAC in murine modeling, and CCL11 expression is increased in Apc mutant tumors. CDX2P-CreERT2Apcfl/fl and CDX2P-CreERT2Apcfl/flCcl11–/– mice that develop colon tumors will be used to test the hypothesis that loss of CCL11 will abrogate tumor development by reducing tumor- promoting inflammation by assessing: tumor number and size, colonic immune cell composition/function, and epithelial function. 3. To determine if targeting CCL11 signaling with neutralizing antibodies protects from colon tumorigenesis. WT mice exposed to AOM-DSS and CDX2P-CreERT2Apcfl/fl mice will be treated with a CCL11 neutralizing antibody (or isotype control) to establish the effects on: tumor number and size, colonic immune cell composition/function, and epithelial function. These studies will provide new mechanistic insights to support the use of treatments based on inhibition of CCL11 in colitis and colon cancer.