The role of colitic stem cell niche in oncogenesis

描述（由申请人提供）：我们的长期目标是阐明炎症和癌症之间的联系，特别是预防结肠炎相关癌症（CAC）。 由于CAC的发病机制尚不清楚，我们的直接目标是确定机制。 因此，对于CAC，我们将确定以下因素的作用：（1）基因缺陷，（2）癌症干细胞（CSC），特别是其克隆性和人类结直肠癌的重演，以及（3）基质成纤维细胞，特别是在结肠癌起始细胞（CCIC）的产生中。 为了探究这些关系，CCIC和结肠炎基质是强大的工具，而球状体培养物（球体）和小鼠异种移植物是强大的癌症模型。 我们的中心假设是，上述三个因素是CAC发展的关键。 这项拟议中的研究与NIH的使命有关，即开发可能有助于减轻人类残疾和疾病负担的基础知识。 我们的初步数据表明，我们有能力（1）在人类溃疡性结肠炎（UC）的一部分非异型增生结肠中富集前体CCIC（pCCIC）;（2）使pCCIC衍生的肿瘤在小鼠体内永久存在;（3）将pCCIC维持为球体。 我们发现结肠炎间质成纤维细胞具有与CRC相关的成纤维细胞相似的细胞因子和信号特征。 我们将用三个目标来检验我们的中心假设。 目的1）确定来自UC和CRC的异种移植物和球体获得的基因缺陷是否遵循经典腺瘤到癌途径中的基因缺陷。 我们将测量一种假定的结肠干细胞标记ALDH的扩展分布，并使用该标记来富集进行自我更新的上皮。 我们将使用突变谱来预测UC衍生上皮在异种移植和球体测定中的成功。 目的2）确定来自结肠癌起始细胞的球体是否是模拟人CRC的克隆和转移行为的真正CSC。 目标2关注球体本身。 我们将通过克隆形成、分化测定来检查pCCIC的自我更新。 目的3）研究IL 8/ UC间质在上皮细胞增殖中的作用。 将来自正常、结肠炎和癌组织的成纤维细胞分离物与pCCIC共培养，以确定其对增殖和致瘤性的影响。 创新：我们的新型细胞工具包括pCCIC、炎症基质、ALDH（一种特异性SC/CSC标记物）和癌症模型-球体和异种移植物-源自人体组织（正常、结肠炎和结肠癌患者）。 重要性：我们的研究将（i）确定肿瘤起始结肠上皮细胞在癌前炎症状态，（ii）揭示上皮/间质关系，（iii）阐明炎症和癌症之间的联系，（iv）促进预防CAC的新策略的开发，（v）大大提高我们预测那些将进展为恶性肿瘤的患者的能力。

DESCRIPTION (provided by applicant): Our long-term goal is to clarify the link between inflammation and cancer in general, and to prevent colitis associated cancer (CAC) in particular. As CAC pathogenesis is unclear, our immediate objective is to determine mechanisms. Therefore, for CAC we will determine the roles of: (1) gene defects, (2) cancer stem cells (CSCs), especially their clonality and recapitulation of human colorectal cancer, and (3) stromal fibroblasts, especially in the generation of colon cancer initiating cells (CCICs). For interrogating these relationships, CCICs and the colitic stroma are powerful tools while spheroid cultures (spheres) and murine xenografts are powerful cancer models. Our central hypothesis is that the above three factors are key to CAC development. The proposed research is relevant to the NIH's mission to develop fundamental knowledge that will potentially help reduce the burdens of human disability and disease. Our preliminary data demonstrate our ability to (1) enrich for precursor CCICs (pCCICs) in a fraction of non-dysplastic colon from human ulcerative colitis (UC); (2) perpetuate pCCIC-derived tumors in mice; (3) maintain pCCICs as spheres. We found that colitic stromal fibroblasts possess a cytokine and signaling profile similar to those fibroblasts associated with CRC. We will test our central hypothesis using three aims. Aim 1) To determine if gene defects acquired by the xenografts and spheres from UC and from CRC follow those in the classical adenoma-to-carcinoma pathway. We will measure the expanded distribution of a putative colon stem cell marker, ALDH, and use this marker to enrich for epithelium undergoing self-renewal. We will use the mutational profile to predict the success of UC-derived epithelium in xenograft and sphere assays. Aim 2) To determine if spheres from colon cancer initiating cells are true CSC which mimic the clonal and metastatic behavior of human CRC. Aim 2 focuses on the spheres themselves. We will examine the pCCICs for self-renewal via clonogenic, differentiation assays. Aim 3) To show the key contribution of IL8 / UC stroma to epithelial proliferation. Fibroblast isolates from normal, colitic, and cancer tissues will be placed in co-culture with pCCICs to determine their effects on both proliferation and tumorigenicity. Innovation: Our novel cellular tools including pCCICs, inflamed stroma, ALDH, a specific SC/CSC marker, and cancer models - spheres & xenografts - derived from human tissues (patients with normal, colitic, and cancerous colon). Significance: Our studies will (i) define the tumor initiating colonic epithelial cell in a preneoplastic, inflammatory state, (ii) reveal epithelial/stromal relationships, (iii) clarify the link between inflammation and cancer, (iv) facilitate the development of novel strategies to prevent CAC, and (v) substantially improve our ability to predict those patients who will progress to malignancy.