Transposon mutagenesis identifies cooperating genetic drivers during keratinocyte transformation and cutaneous squamous cell carcinoma progression

The systematic identification of genetic events driving cellular transformation and tumor progression in the absence of a highly recurrent oncogenic driver mutation is a challenge in cutaneous oncology. In cutaneous squamous cell carcinoma (cuSCC), the high UV-induced mutational burden poses a hurdle to achieve a complete molecular landscape of this disease. Here, we utilized the Sleeping Beauty transposon mutagenesis system to statistically define drivers of keratinocyte transformation and cuSCC progression in vivo in the absence of UV-IR, and identified both known tumor suppressor genes and novel oncogenic drivers of cuSCC. Functional analysis confirms an oncogenic role for the ZMIZ genes, and tumor suppressive roles for KMT2C, CREBBP and NCOA2, in the initiation or progression of human cuSCC. Taken together, our in vivo screen demonstrates an extremely heterogeneous genetic landscape of cuSCC initiation and progression, which can be harnessed to better understand skin oncogenic etiology and prioritize therapeutic candidates. Author summary Non-melanoma skin cancers, the most common cancers in the US, are caused by UV skin exposure. Nearly 1 million cases of cutaneous squamous cell carcinoma (cuSCC) are diagnosed in the US each year. While most cuSCCs are highly treatable, more than twice as many individuals die from this disease as from melanoma. The high burden of UV-induced DNA damage in human skin poses a challenge for identifying initiating and cooperating mutations that promote cuSCC development and for defining potential therapeutic targets. Here, we describe a genetic screen in mice using a DNA transposon system to mutagenize the genome of keratinocytes and drive squamous cell carcinoma in the absence of UV. By sequencing where the transposons selectively integrated in the genomes of normal skin, skin with pre-cancerous lesions and skin with fully developed cuSCCs from our mouse model, we were able to identify frequently mutated genes likely important for this disease. Our analysis also defined cooperation between sets of genes not previously appreciated in cuSCC. Our mouse model and ensuing data provide a framework for understanding the genetics of cuSCC and for defining the molecular changes that may lead to the future therapies for patients.