In situ CRISPR-Cas9 base editing for the development of genetically engineered mouse models of breast cancer.

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.