Role of TLR4 in Colitis Associated Neoplasia

TLR4 在结肠炎相关肿瘤中的作用

• 批准号: 9102094
• 负责人: Maria Teresa Abreu
• 金额: $ 33.39万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9102094
• 关键词: 1-Phosphatidylinositol 3-Kinase; Address; Animal Model; Animals; Automobile Driving; Bacteria; Biological Response Modifiers; Bone Marrow; Cells; Chimera organism; Chronic; Clinical; Colectomy; Colitis; Colon; Colon Carcinoma; Colonic Neoplasms; Colorectal Cancer; Data; Development; Dysplasia; Epithelial; Epithelial Cell Proliferation; Epithelial Cells; Epithelium; Gene Targeting; Grant; Growth; Health; Health Care Costs; Human; Immune; In Vitro; Inflammation; Inflammatory; Intestinal Cancer; Intestinal Neoplasms; Intestines; Knockout Mice; Laboratories; Lead; Leukocyte L1 Antigen Complex; Ligands; Maintenance; Malignant - descriptor; Malignant Neoplasms; Mediating; Molecular; Mucous Membrane; Mus; Mutate; Myelogenous; Neoplasms; Neoplastic Cell Transformation; Oncogenes; Pathway interactions; Patient Care; Patients; Phenotype; Play; Precancerous Conditions; Quality of life; Reporter; Role; S100A8 gene; S100A9 gene; Scientist; Signal Transduction; Stem cells; Suppressor-Effector T-Lymphocytes; TLR4 gene; Techniques; Testing; Tissue Banks; Transgenic Organisms; Ulcerative Colitis; Undifferentiated; Work; Xenograft procedure; autocrine; beta catenin; cancer stem cell; colitis associated cancer; colon cancer cell line; improved; in vivo; in vivo Model; microbiota; mouse model; mouse toll-like receptor 4; neoplastic cell; overexpression; paracrine; prevent; progenitor; programs; promoter; receptor; self-renewal; stemness; tumor; tumorigenesis; villin

DESCRIPTION (provided by applicant): The proposed project aims to understand the molecular underpinnings of inflammation-associated neoplasia. Patients with IBD continue to need colectomies for dysplasia largely because of our inability to halt IBD progression to cancer. Our previous work has demonstrated that innate immune signaling by toll-like receptor 4 (TLR4) contributes importantly to the development of colitis-associated cancer (CAC). We identified TLR4 as an oncogene which promotes colonic malignancy. We have shown that TLR4 is over-expressed in UC- associated dysplasia and in almost all CACs. We showed that animals deficient in TLR4 (TLR4-/- mice) are protected from colitis-associated neoplasia and have generated a mouse in which the villin promoter drives transgenic intestinal TLR4 overexpression (V-TLR4). These mice are highly prone to colitis-associated neoplasia. Mechanistically, we have shown that TLR4 promotes neoplasia by activating ß-catenin signaling through PI3 kinase in colonic epithelial cells. We present preliminary data that show that TLR4 drives expansion of Lgr5+ stem cells in both intestinal neoplasia and in normal mucosa suggesting that TLR4 signaling is driving stem cell self-renewal in this model in vivo. We also show that UC mucosa and TLR4- mediated inflammatory neoplasia in mouse models are characterized by an infiltrate of S100A8/A9 expressing myeloid-derived suppressor cells (MDSCs). In the current proposal we will test the hypothesis that TLR4 signaling plays both an autocrine role in malignant epithelial transformation and a paracrine role through activation of MDSCs to stimulate ß-catenin activation, epithelial cell proliferation and cancer stem cell expansion. This is pursued i the following specific aims: 1) Determine the role of TLR4 signaling in activation of intestinal stem cells in inflammatory neoplasia. Using our various TLR4 expressing animal models crossed to stem cell reporter Lgr5-EGFP mice we will investigate if TLR4 is required for expansion of lineage committed progenitors and of colon cancer stem cells during inflammation and neoplastic transformation in vivo. 2) Determine the role of TLR4 signaling in promoting stemness of colonic epithelial cells. We will use primary colonoid cultures to test if TLR4 signaling alters the phenotype (growth, budding) of primary colon stem cells in culture. We will use colon cancer cell lines to test whether TLR4 signaling promotes a cancer stem cell phenotype in vitro and increases tumorgenicity in vivo using xenografts. 3) Determine if S100A8/9+ MDSCs drive TLR4-dependent ß-catenin activation, stem cell activation, and tumorigenesis. We will determine whether S100A8/9 MDSCs are required for intestinal stem cell expansion and CAC. We will test if S100A8 or 9 activate ß-catenin in colonic epithelia in a TLR4-dependent manner. Human UC-associated MDSCs will be isolated and studied for their ability to stimulate ß-catenin signaling in colonic epithelia in vitro. The work proposed herein aims to provide the mechanistic justification for subsequent human studies to target innate immune signaling as a means to halt progression UC/inflammation to dysplasia.

描述(由申请人提供)：拟议的项目旨在了解炎症相关肿瘤的分子基础。IBD患者仍然需要结肠切除术来治疗发育不良，这主要是因为我们无法阻止IBD向癌症的发展。我们以前的工作已经证明，Toll样受体4(TLR4)的先天免疫信号在结肠炎相关癌症(CAC)的发展中起着重要作用。我们发现TLR4是一个促进结肠恶性的癌基因。我们发现TLR4在UC相关的异型增生和几乎所有的癌组织中都有过度表达。我们发现，缺乏TLR4的动物(TLR4-/-小鼠)可以免受结肠炎相关肿瘤的保护，并产生了一种由绒毛蛋白启动子驱动转基因肠道TLR4过表达(V-TLR4)的小鼠。这些小鼠极易患结肠炎相关肿瘤。从机制上讲，我们已经证明TLR4通过激活结肠上皮细胞中的PI3激酶来激活？连环蛋白信号，从而促进肿瘤的发生。我们提供的初步数据表明，在肠道肿瘤和正常粘膜中，TLR4都驱动Lgr5+干细胞的扩张，这表明在活体模型中，TLR4信号正在驱动干细胞的自我更新。我们还发现，在小鼠模型中，UC粘膜和TLR4介导的炎性肿瘤的特征是S100A8/A9表达髓系来源的抑制细胞(MDSCs)。在目前的方案中，我们将检验TLR4信号在恶性上皮转化中既发挥自分泌作用，又通过激活MDSCs刺激ç-连环蛋白激活、上皮细胞增殖和肿瘤干细胞扩增的旁分泌作用的假设。这项研究的具体目的如下：1)确定TLR4信号在炎性肿瘤中肠道干细胞激活中的作用。使用我们的各种表达TLR4的动物模型与干细胞报告基因Lgr5-EGFP小鼠杂交，我们将研究TLR4是否在体内炎症和肿瘤转化过程中扩增谱系承诺的祖细胞和结肠癌干细胞所必需的。2)确定TLR4信号在促进结肠上皮细胞干化中的作用。我们将使用原代结肠样细胞培养来测试TLR4信号是否改变了原代结肠干细胞在培养中的表型(生长、萌发)。我们将使用结肠癌细胞系来测试TLR4信号在体外是否促进癌症干细胞的表型，并通过异种移植增加体内的致瘤性。3)确定S100A8/9+MDSCs是否驱动TLR4依赖的？连环蛋白激活、干细胞激活和肿瘤发生。我们将确定肠干细胞扩增和CAC是否需要S100A8/9 MDSCs。我们将测试S100A8或S100A9是否以TLR4依赖的方式激活结肠上皮细胞中的？连环蛋白。人UC相关的MDSCs将被分离出来，并在体外研究其刺激结肠上皮细胞内？连环蛋白信号的能力。本文提出的工作旨在为后续的人类研究提供机制上的理由，将先天免疫信号作为阻止UC/炎症进展到异型增生的一种手段。