Role of TLR4 in Colitis Associated Neoplasia

TLR4 在结肠炎相关肿瘤中的作用

• 批准号: 8761283
• 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/8761283
• 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; Commit; Data; Development; Dysplasia; Epithelial; Epithelial Cell Proliferation; Epithelial Cells; Epithelium; Gene Targeting; Grant; Growth; Health Care Costs; Human; Immune; In Vitro; Inflammation; Inflammatory; Intestinal Cancer; Intestinal Neoplasms; Intestines; 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; Techniques; Testing; Tissue Banks; Transgenic Organisms; Ulcerative Colitis; Undifferentiated; Work; Xenograft procedure; autocrine; cancer stem cell; colitis associated cancer; colon cancer cell line; improved; in vivo; in vivo Model; mouse model; mouse toll-like receptor 4; neoplastic cell; overexpression; paracrine; prevent; progenitor; programs; promoter; public health relevance; receptor; self-renewal; stemness; toll-like receptor 4; 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 ?-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 b-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 b-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 ?-catenin in colonic epithelia in a TLR4-dependent manner. Human UC-associated MDSCs will be isolated and studied for their ability to stimulate ?-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）：拟建项目旨在了解炎症相关肿瘤的分子基础。由于我们无法阻止IBD向癌症的发展，IBD患者仍然需要结肠切除术来治疗发育不良。我们之前的工作已经证明toll样受体4 （TLR4）的先天免疫信号在结肠炎相关癌症（CAC）的发展中起重要作用。我们发现TLR4是一种促进结肠恶性肿瘤的致癌基因。我们发现TLR4在UC相关的异常增生和几乎所有的cac中都过表达。我们发现TLR4缺失的动物（TLR4-/-小鼠）免受结肠炎相关肿瘤的影响，并产生了一种绒毛蛋白启动子驱动转基因肠道TLR4过表达（V-TLR4）的小鼠。这些小鼠极易发生结肠炎相关的肿瘤。在机制上，我们已经证明TLR4通过激活？-catenin信号通过PI3激酶在结肠上皮细胞中传递。我们提供的初步数据显示，TLR4在肠道肿瘤和正常粘膜中都驱动Lgr5+干细胞的扩增，这表明TLR4信号传导在体内模型中驱动干细胞自我更新。我们还发现，UC粘膜和TLR4介导的炎性瘤变在小鼠模型中以表达髓源性抑制细胞（MDSCs）的S100A8/A9浸润为特征。在目前的提案中，我们将验证TLR4信号在恶性上皮转化中发挥自分泌作用和旁分泌作用的假设，通过激活MDSCs刺激b-连环蛋白激活，上皮细胞增殖和癌症干细胞扩增。具体目的如下：1)确定TLR4信号在炎性肿瘤中肠干细胞激活中的作用。利用我们的各种TLR4表达动物模型与干细胞报告细胞Lgr5-EGFP小鼠杂交，我们将研究在体内炎症和肿瘤转化过程中，TLR4是否需要谱系承诺祖细胞和结肠癌干细胞的扩增。2)确定TLR4信号在促进结肠上皮细胞干性中的作用。我们将使用原代结肠腺培养来测试TLR4信号是否会改变培养中原代结肠干细胞的表型（生长、出芽）。我们将使用结肠癌细胞系来测试TLR4信号是否在体外促进癌症干细胞表型并在体内使用异种移植物增加肿瘤致癌性。3)确定S100A8/9+ MDSCs是否驱动tlr4依赖性b-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是否激活？-catenin在结肠上皮中以tlr4依赖的方式表达。人类uc相关的MDSCs将被分离并研究其刺激？-catenin信号在结肠上皮细胞中的作用。本文提出的工作旨在为随后的人类研究提供机制依据，以先天免疫信号为目标，作为阻止UC/炎症进展为不典型增生的手段。