Genetically engineered mouse models (GEMMs) of cancer have proven to be of great value for basic and translational research. Although CRISPR-based gene disruption offers a fast-track approach for perturbing gene function and circumvents certain limitations of standard GEMM development, it does not provide a flexible platform for recapitulating clinically relevant missense mutations in vivo. To this end, we generated knock-in mice with Cre-conditional expression of a cytidine base editor and tested their utility for precise somatic engineering of missense mutations in key cancer drivers. Upon intraductal delivery of sgRNA-encoding vectors, we could install point mutations with high efficiency in one or multiple endogenous genes in situ and assess the effect of defined allelic variants on mammary tumorigenesis. While the system also produces bystander insertions and deletions that can stochastically be selected for when targeting a tumor suppressor gene, we could effectively recapitulate oncogenic nonsense mutations. We successfully applied this system in a model of triple-negative breast cancer, providing the proof of concept for extending this flexible somatic base editing platform to other tissues and tumor types.

中文翻译：

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原位 CRISPR-Cas9 碱基编辑用于开发乳腺癌基因工程小鼠模型。


癌症基因工程小鼠模型（GEMM）已被证明对于基础和转化研究具有巨大价值。尽管基于 CRISPR 的基因破坏提供了一种干扰基因功能的快速方法，并规避了标准 GEMM 开发的某些限制，但它并没有提供灵活的平台来在体内重现临床相关的错义突变。为此，我们生成了具有胞苷碱基编辑器 Cre 条件表达的敲入小鼠，并测试了它们在关键癌症驱动因素错义突变的精确体细胞工程中的实用性。在导管内递送 sgRNA 编码载体后，我们可以在一个或多个内源基因中高效地原位安装点突变，并评估特定等位基因变异对乳腺肿瘤发生的影响。虽然该系统还产生旁观者插入和删除，当靶向肿瘤抑制基因时可以随机选择这些插入和删除，但我们可以有效地重现致癌无义突变。我们成功地将这个系统应用于三阴性乳腺癌模型，为将这种灵活的体细胞碱基编辑平台扩展到其他组织和肿瘤类型提供了概念证明。