Understanding the Mechanism of a Gut Microbial Genotoxin Involved in Colorectal Carcinogenesis

项目总结 结直肠癌(CRC)是美国第三大最流行的癌症形式，也是导致癌症的第二大原因 癌症死亡。过去几十年的研究表明，肠道微生物区系影响结直肠癌，并且 最近的研究表明，肠道细菌基因毒素是癌症发生和发展的关键效应因子。 与癌症关系最密切的细菌基因毒素之一是结肠素，这是一种由人类产生的代谢物 肠道共生细菌，包括某些大肠杆菌菌株，它们拥有一个生物合成基因簇，称为 PKS岛。大肠癌患者中PKS+大肠埃希菌的流行及PKS+菌株增强能力的研究 在结直肠癌小鼠模型中的肿瘤发生表明，结节蛋白可能在癌症进展中起到因果作用。 然而，从机制上理解Colibactin的遗传毒性和对CRC的贡献一直是 由于无法分离和表征活性的遗传毒性代谢物而受阻(S)。在.期间 在这笔赠款的前一个资助期，我们获得了关于Colibactin的结构和模式的关键信息 通过研究参与其生物合成的酶来发挥作用。最值得注意的是，我们确定了Colibactin是一种 DNA烷化剂，并提出了具有活性的基因毒素的潜在结构。这样做的总体目标是 新的应用是阐明Colibactin活性和作用的其他分子机制。 结直肠癌的发生。建立在核心假设的基础上，即Colibactin的基因毒性活性来自于 DNA链间交联链(ICL)的形成，我们的三个具体目标将：1)表征特异性 和结缔组织肌动蛋白-DNA ICL的结构；2)开发抑制微生物中结缔组织蛋白生物合成的小分子 3)阐明结缔组织肌动蛋白介导的DNA损伤在结直肠癌中的生理位置和时间 发展。我们的多学科方法将使这些进步成为可能，这种方法将知识和 来自化学生物学、结构生物学、微生物学、毒理学和癌症生物学的技术。总体而言，这 努力将填补在基础知识方面的关键空白，这些知识需要阐明产生结肠素的肠道的作用。 细菌在结直肠癌的致癌作用，并最终影响癌症的预防、诊断和治疗。另外， 通过成功地证明研究和操纵与疾病相关的个体微生物途径 可以提供关键的机械洞察，这项工作还将支持和验证未来的努力，以了解如何 其他肠道微生物活动影响结直肠癌的发生和发展。

PROJECT SUMMARY Colorectal cancer (CRC) is the third most prevalent form of cancer in the US and the second leading cause of cancer deaths. Studies over the last several decades have revealed that the gut microbiota influences CRC, and recent work has implicated gut bacterial genotoxins as key effectors in cancer development and progression. One of the bacterial genotoxins most strongly connected to cancer is colibactin, a metabolite produced by human gut commensal bacteria, including certain E. coli strains, that possess a biosynthetic gene cluster termed the pks island. The increased prevalence of pks+ E. coli in CRC patients and the ability of pks+ strains to potentiate tumorigenesis in mouse models of CRC suggest colibactin may play a causal role in cancer progression. However, achieving a mechanistic understanding of colibactin's genotoxicity and contribution to CRC has been impeded by an inability to isolate and structurally characterize the active genotoxic metabolite(s). During the previous funding period of this grant, we gained critical information about colibactin's structure and mode of action by studying the enzymes involved in its biosynthesis. Most notably, we established that colibactin is a DNA alkylating agent and proposed a potential structure for the active genotoxin. The overall objective of this renewal application is to elucidate additional molecular mechanisms underlying colibactin's activity and role in CRC carcinogenesis. Building off of the central hypothesis that the genotoxic activity of colibactin derives from the formation of DNA interstrand cross-links (ICLs), our three specific aims will: 1) characterize the specificity and structure of colibactin-DNA ICLs; 2) develop small molecules that inhibit colibactin biosynthesis in microbial communities; 3) elucidate the physiological location and timing of colibactin-mediated DNA damage in CRC development. These advances will be enabled by our multidisciplinary approach, which merges knowledge and techniques from chemical biology, structural biology, microbiology, toxicology, and cancer biology. Overall, this effort will fill critical gaps in fundamental knowledge needed to elucidate the role of colibactin-producing gut bacteria in CRC carcinogenesis and ultimately impact cancer prevention, diagnosis, and treatment. Additionally, by successfully demonstrating that studying and manipulating individual disease-associated microbial pathways can provide key mechanistic insights, this work will also support and validate future efforts to understand how other gut microbial activities influence CRC initiation and development.