AbstractBackgroundMutations in the receptor tyrosine kinase genefibroblast growth factor receptor 2(FGFR2) occur at a high frequency in endometrial cancer (EC) and have been linked to advanced and recurrent disease. However, little is known about how these mutations drive carcinogenesis.MethodsDifferential transcriptomic analysis and two‐step quantitative real‐time PCR (qRT‐PCR) assays were applied to identify genes differentially expressed in two cohorts of EC patients carrying mutations in theFGFR2gene as well as in EC cells harbouring mutations in theFGFR2. Candidate genes and target signalling pathways were investigated by qRT‐PCR assays, immunohistochemistry and bioinformatics analysis. The functional roles of differently regulated genes were analysed using in vitro and in vivo experiments, including 3D‐orthotypic co‐culture systems, cell proliferation and migration protocols, as well as colony and focus formation assays together with murine xenograft tumour models. The molecular mechanisms were examined using CRISPR/Cas9‐based loss‐of‐function and pharmacological approaches as well as luciferase reporter techniques, cell‐based ectodomain shedding assays and bioinformatics analysis.ResultsWe show that commonFGFR2mutations significantly enhance the sensitivity to FGF7‐mediated activation of a disintegrin and metalloprotease (ADAM)17 and subsequent transactivation of the epidermal growth factor receptor (EGFR). We further show that FGFR2 mutants trigger the activation of ADAM10‐mediated Notch signalling in an ADAM17‐dependent manner, highlighting for the first time an intimate cooperation between EGFR and Notch pathways in EC. Differential transcriptomic analysis in EC cells in a cohort of patients carrying mutations in theFGFR2gene identified a strong association betweenFGFR2mutations and increased expression of members of the Notch pathway and ErbB receptor family. Notably, FGFR2 mutants are not constitutively active but require FGF7 stimulation to reprogram Notch and EGFR pathway components, resulting in ADAM17‐dependent oncogenic growth.ConclusionsThese findings highlight a pivotal role of ADAM17 in the pathogenesis of EC and provide a compelling rationale for targeting ADAM17 protease activity in FGFR2‐driven cancers.