A versatile ES cell-based melanoma mouse modeling platform

The cumbersome and time-consuming process of generating new mouse strains and multi-allelic experimental animals often hinders the use of genetically engineered mouse models (GEMM) in cancer research. Here, we describe the development and validation of a melanoma embryonic stem cell (ESC)-GEMM platform. Our platform incorporates twelve clinically relevant genotypes composed of combinations of four driver alleles (LSL-BrafV600E, LSL-NrasQ61R, PtenFlox, Cdkn2aFlox) and regulatory alleles to spatiotemporally control the perturbation of genes-of-interest. Our ESCs produce high contribution chimeras, which develop melanoma phenotypes that are similar to conventionally bred mice. To validate our melanoma ESC-GEMM platform, we modulated Pten expression on a BrafV600E background for proof-of-principle experiments. We demonstrate how the different efficiency and kinetics of CRISPR-Cas9, RNA interference (RNAi), and Cre/loxP impact melanoma formation, and highlight how inducible CRISPR-Cas9 and RNAi can be utilized to study cancer gene function in vivo. Moreover, we show that chimera-derived melanoma cell lines retain regulatory allele competency and can be used in vitro and in syngeneic grafts in vivo to complement ESC-GEMM chimera experiments. Thus, when combined with modern genetic tools, our ESC-GEMM platform enables rapid, high-throughput, and versatile studies aimed at addressing outstanding questions in melanoma biology.