Colibactin-Producing Escherichia coli as an Environmental Stimulus Shaping Pre-Cancer Progression

散发性结直肠癌（CRC）是一个公共卫生问题，每年影响全球100多万人 在美国，它一直排在男性和女性癌症相关死亡原因的前2-3位 尽管结肠镜检查被广泛采用。异时性癌前病变在息肉切除术后也很高。 然而，只有一小部分结肠癌前病变（常规腺瘤或无蒂锯齿状病变） 进展到CRC，我们缺乏分子标记来识别这些个体，也缺乏对这些个体的理解。 促进癌前病变的机制。CRC的巨大疾病负担，与人口密集的 结肠微生物群，以及认识到许多CRC风险因素（例如，吸烟、肥胖、肉食性饮食） 结肠微生物群促使人们研究微生物群是否以及如何促进CRC发病。 累积的数据现在强烈支持这一假设，即微生物群是结肠癌的关键环境贡献者。 致癌作用尽管如此，关于微生物群“是否以及如何”对结肠癌前病变的影响还知之甚少。 癌的我们的初步数据确定大肠杆菌释放非核糖体代谢物遗传毒素， 大肠杆菌素，称为pks+ E。大肠杆菌（Ecpks），与结肠癌前病变的检测密切相关。此外，本发明还 据报道，大肠杆菌素可诱导结肠上皮细胞DNA中的特异性突变特征，特别是突变 在APC中，CRC驱动基因在腺瘤发展中特别重要。值得注意的是， 与CRC有关。在我们的结肠MAP研究中，我们还发现无蒂锯齿状病变 占间隔CRC的很大一部分，很可能是微生物侵害的结果。但委员会仍 不知道Eppks是否有助于SSL的发展和进步。之前的几项人类研究， 包括我们的初步研究，都是基于横截面设计。为了弥补这一差距， 为了理解时间序列，我们还将包括纵向设计和人类结肠模型， 测试我们的核心假设，即ECPKs的一个子集与人类癌前病变相关并有助于推动人类癌前病变 进展我们将使用大量的迭代方法，并在概念上和实验上整合 这个项目与项目1和3以及这个U 54的核心。我们在这个翻译项目中的具体目标是：1） 评估Eppks定植与人类结直肠癌前病变的相关性， vs.使用现有人类队列、临床预测因子和纵向数据，降低（“惰性”）潜力; 2） 检查与癌前结肠病变相关的机制，按疾病状态（无痛或 进行性）和Ecpks状态，使用全外显子组测序、单细胞RNAseq和多重 免疫荧光（MxIF）方法;和3）为了鉴定与Eppks毒力决定簇相关的Eppks毒力决定簇， 进行性（与惰性）结肠癌前病变和疾病机制。在一起， 我们的研究有可能加速非侵入性、具有成本效益的CRC筛查和监测， 散发性CRC风险增加的个体的关键子集。

Sporadic colorectal cancer (CRC) is a public health problem, affecting over a million people each year globally and, in the United States, consistently ranks in the top 2-3 causes of cancer-related death in men and women despite the wide adoption of colonoscopy. Metachronous pre-cancers are also high following polypectomy. However, only a small percent of colon pre-cancers (conventional adenomas or sessile serrated lesions) progress to CRC, and we lack both molecular markers to identify these individuals and an understanding of the mechanisms fostering pre-cancer. The large disease burden of CRC, co-localized with the densely populated colon microbiota, and the recognition that many CRC risk factors (e.g., smoking, obesity, carnivorous diet) modify the colon microbiota has spurred investigations into ‘if and how’ the microbiota contributes to CRC pathogenesis. Accrued data now strongly support the hypothesis that the microbiota is a key environmental contributor to colon carcinogenesis. Nonetheless, little is yet known about ‘if and how’ the microbiota contributes to colon pre- cancers. Our preliminary data identify Escherichia coli that release the non-ribosomal, metabolite genotoxin, colibactin, known as pks+ E. coli (Ecpks), as strongly associated with detection of colon pre-cancers. Further, colibactin is reported to induce specific mutational signatures in colon epithelial cell DNA, in particular, mutations in APC, a CRC driver gene particularly important in adenoma development. Notably, these Ecpks mutations have been associated with CRC. In our COLON MAP study, we have also identified that sessile serrated lesions which account for a large proportion of interval CRC, are likely a result of microbial insult. However, it remains unknown whether Ecpks contributes to SSL development and progression. The few previous human studies, including our preliminary studies, are based on a cross-sectional design. Thus, to address this gap and understand the temporal sequence, we will also include a longitudinal design and human colonoids models to test our core hypothesis that a subset of Ecpks associate with and contribute to driving human pre-cancer progression. We will use a wide array of iterative approaches and conceptual and experimental integration of this project with Projects 1 and 3 and the cores of this U54. Our specific aims in this translational project are: 1) To evaluate the association of Ecpks colonization with human colorectal pre-cancers with greater progressive vs. lower (“indolent”) potential using in-hand human cohorts, clinical predictors, and longitudinal data; 2) To examine mechanisms associated with pre-cancer colon lesions stratified by disease state (indolent or progressive) and Ecpks status using whole exome sequencing, single cell RNAseq and multiplex immunofluorescence (MxIF) approaches; and 3) To identify Ecpks virulence determinants associated with progressive (vs indolent) colon pre-cancer and disease mechanisms using human colonoid models. Together, our studies have the potential to accelerate noninvasive, cost-effective CRC screening and surveillance for a critical subset of individuals at increased risk for sporadic CRC.