First Department of Medicine, Friedrich-Alexander-Universität, Erlangen-Nürnberg, Universitätsklinikum Erlangen, Erlangen, Germany.
Deutsches Zentrum Immuntherapie (DZI), Erlangen, Germany.
Nat Protoc. 2021 Jan;16(1):61-85. doi: 10.1038/s41596-020-00412-1. Epub 2020 Dec 14.
Despite advances in the detection and therapy of colorectal cancer (CRC) in recent years, CRC has remained a major challenge in clinical practice. Although alternative methods for modeling CRC have been developed, animal models of CRC remain helpful when analyzing molecular aspects of pathogenesis and are often used to perform preclinical in vivo studies of potential therapeutics. This protocol updates our protocol published in 2007, which provided an azoxymethane (AOM)-based setup for investigations into sporadic (Step 5A) and, when combined with dextran sodium sulfate (Step 5B), inflammation-associated tumor growth. This update also extends the applications beyond those of the original protocol by including an option in which AOM is serially applied to mice with p53 deficiency in the intestinal epithelium (Step 5C). In this model, the combination of p53 deficiency and AOM promotes tumor development, including growth of invasive cancers and lymph node metastasis. It also provides details on analysis of colorectal tumor growth and metastasis, including analysis of partial epithelial-to-mesenchymal transition, cell isolation and co-culture studies, high-resolution mini-endoscopy, light-sheet fluorescence microscopy and micro-CT imaging in mice. The target audience for our protocol is researchers who plan in vivo studies to address mechanisms influencing sporadic or inflammation-driven tumor development, including the analysis of local invasiveness and lymph node metastasis. It is suitable for preclinical in vivo testing of novel drugs and other interventional strategies for clinical translation, plus the evaluation of emerging imaging devices/modalities. It can be completed within 24 weeks (using Step 5A/C) or 10 weeks (using Step 5B).
尽管近年来在结直肠癌 (CRC) 的检测和治疗方面取得了进展，但 CRC 在临床实践中仍然是一个主要挑战。尽管已经开发出了替代 CRC 建模的方法，但在分析发病机制的分子方面，动物 CRC 模型仍然很有帮助，并且常用于对潜在治疗方法进行临床前体内研究。本方案更新了我们在 2007 年发表的方案，该方案提供了一种基于偶氮甲烷 (AOM) 的方案，用于研究散发性 (步骤 5A)，以及与葡聚糖硫酸钠 (步骤 5B) 结合时炎症相关的肿瘤生长。这一更新还通过包括在肠上皮中 p53 缺陷的小鼠中连续应用 AOM 的选项（步骤 5C），将应用范围扩展到原始方案之外。在该模型中，p53 缺陷和 AOM 的组合促进了肿瘤的发展，包括侵袭性癌症和淋巴结转移的生长。它还提供了关于结直肠肿瘤生长和转移分析的详细信息，包括对上皮 - 间充质转化、细胞分离和共培养研究、高分辨率迷你内镜、光片荧光显微镜和小鼠微 CT 成像的分析。我们方案的目标受众是计划进行体内研究以解决影响散发性或炎症驱动的肿瘤发展的机制的研究人员，包括对局部侵袭性和淋巴结转移的分析。它适用于临床转化的新型药物和其他干预策略的临床前体内测试，以及新兴成像设备/模式的评估。它可以在 24 周内（使用步骤 5A/C）或 10 周内（使用步骤 5B）完成。
