Targetable fibroblast phenotypes and EMT malignant cell states cooperate to promote tumor progression in esophageal adenocarcinoma

Abstract Background: Esophageal adenocarcinoma (EAC) is usually resistant to cytotoxic therapies and immunotherapies have gained little traction. Cancer-associated fibroblasts (CAF) are a major stromal cell population in the EAC tumor microenvironment associated with prognosis and treatment outcomes. Recent evidence suggests that cancer cell phenotypes related to EMT may determine CAF heterogeneity, but the molecular and cellular biology that underlies myofibroblast fate in EAC is not well understood. Methods: To obtain the most comprehensive profile of CAF heterogeneity in EAC, we performed histopathologic, single-cell RNA sequencing and transcriptomic analyses on 28 samples from 26 patients and prognostic validation using two EAC cohorts from genomic consortia. Results: Combining histologic and molecular profiles revealed five CAF phenotypes, including three myofibroblast phenotypes, associated with EMT-related signatures in EAC cells and cellular interactions that promote tumor progression and metastasis. We identified a specific myofibroblast subtype (CAF5) in close proximity to cancer cells and tumor vasculature with exclusive expression of TRPA1, offering a potential therapeutic vulnerability. We reconstructed CAF differentiation trajectories from esophagus-resident universal fibroblasts to identify candidate genes central to the CAF phenotype, and used this knowledge to construct a combined EMT-myofibroblast four gene signature (GSN, ATF1, ZEB2 and POSTN) that was highly prognostic in EAC and several other solid tumors. Conclusions: This study stratifies EAC patients into two prognostic groups and reports important data supporting a cancer – CAF signalling axis, promoting myofibroblastic differentiation, that may inform more effective treatment strategies.