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Colonoscopy-based colorectal cancer modeling in mice with CRISPR-Cas9 genome editing and organoid transplantation.
Nature Protocols ( IF 14.8 ) Pub Date : 2018-Feb-01 , DOI: 10.1038/nprot.2017.136
Jatin Roper 1, 2, 3 , Tuomas Tammela 4 , Adam Akkad 1 , Mohammad Almeqdadi 1 , Sebastian B Santos 1 , Tyler Jacks 1, 5 , Ömer H Yilmaz 1, 6
Affiliation  

Most genetically engineered mouse models (GEMMs) of colorectal cancer are limited by tumor formation in the small intestine, a high tumor burden that limits metastasis, and the need to generate and cross mutant mice. Cell line or organoid transplantation models generally produce tumors in ectopic locations-such as the subcutaneous space, kidney capsule, or cecal wall-that do not reflect the native stromal environment of the colon mucosa. Here, we describe detailed protocols to rapidly and efficiently induce site-directed tumors in the distal colon of mice that are based on colonoscopy-guided mucosal injection. These techniques can be adapted to deliver viral vectors carrying Cre recombinase, CRISPR-Cas9 components, CRISPR-engineered mouse tumor organoids, or human cancer organoids to mice to model the adenoma-carcinoma-metastasis sequence of tumor progression. The colonoscopy injection procedure takes ∼15 min, including preparation. In our experience, anyone with reasonable hand-eye coordination can become proficient with mouse colonoscopy and mucosal injection with a few hours of practice. These approaches are ideal for a wide range of applications, including assessment of gene function in tumorigenesis, examination of tumor-stroma interactions, studies of cancer metastasis, and translational research with patient-derived cancers.

中文翻译:

使用CRISPR-Cas9基因组编辑和类器官移植在小鼠中基于结肠镜检查的结肠直肠癌建模。

大肠癌的大多数基因工程小鼠模型(GEMM)受小肠肿瘤形成,限制转移的高肿瘤负担以及生成和交叉突变小鼠的限制。细胞系或类器官移植模型通常会在异位部位(例如皮下空间,肾囊或盲肠壁)产生肿瘤,这些肿瘤不能反映结肠粘膜的天然基质环境。在这里,我们描述了基于结肠镜检查指导的粘膜注射快速有效地诱导小鼠远端结肠定点肿瘤的详细方案。这些技术可适用于递送携带Cre重组酶,CRISPR-Cas9组件,CRISPR工程化的小鼠肿瘤类器官,或人类癌症类器官到小鼠,以模拟肿瘤进展的腺瘤-癌-转移序列。结肠镜检查注射过程约需15分钟,包括准备工作。根据我们的经验,只要有几个小时的练习,任何具有合理手眼协调能力的人都可以精通鼠标结肠镜检查和粘膜注射。这些方法是广泛应用的理想选择,包括评估肿瘤发生中的基因功能,检查肿瘤与基质之间的相互作用,研究癌症转移以及患者来源的癌症的转化研究。
更新日期:2018-01-04
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