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ObjectiveGATA6 is a key regulator of the classical phenotype in pancreatic ductal adenocarcinoma (PDAC). Low GATA6 expression associates with poor patient outcome.GATA4is the second most expressed GATA factor in the pancreas. We assessed whether, and how, GATA4 contributes to PDAC phenotype and analysed the association of expression with outcome and response to chemotherapy.DesignWe analysed PDAC transcriptomic data, stratifying cases according toGATA4andGATA6expression and identified differentially expressed genes and pathways. The genome-wide distribution of GATA4 was assessed, as well as the effects ofGATA4knockdown. A multicentre tissue microarray study to assess GATA4 and GATA6 expression in samples (n=745) from patients with resectable was performed. GATA4 and GATA6 levels were dichotomised into high/low categorical variables; association with outcome was assessed using univariable and multivariable Cox regression models.ResultsGATA4messenger RNA is enriched in classical, compared with basal-like tumours. We classified samples in 4 groups as high/low forGATA4andGATA6. Reduced expression ofGATA4had a minor transcriptional impact but low expression ofGATA4enhanced the effects ofGATA6low expression. GATA4 and GATA6 display a partially overlapping genome-wide distribution, mainly at promoters. Reduced expression of both proteins in tumours was associated with the worst patient survival.GATA4andGATA6expression significantly decreased in metastases and negatively correlated with basal markers.ConclusionsGATA4andGATA6cooperate to maintain the classical phenotype. Our findings provide compelling rationale to assess their expression as biomarkers of poor prognosis and therapeutic response.

Objective: The aggressive basal-like molecular subtype of pancreatic ductal adenocarcinoma (PDAC) harbours a ΔNp63 (p40) gene expression signature reminiscent of a basal cell type. Distinct from other epithelia with basal tumours, ΔNp63+ basal cells reportedly do not exist in the normal pancreas. Design: We evaluated ΔNp63 expression in human pancreas, chronic pancreatitis (CP) and PDAC. We further studied in depth the non-cancerous tissue and developed a three-dimensional (3D) imaging protocol (FLIP-IT, Fluorescence Light sheet microscopic Imaging of Paraffin-embedded or Intact Tissue) to study formalin-fixed paraffin-embedded samples at single cell resolution. Pertinent mouse models and HPDE cells were analysed. Results: In normal human pancreas, rare ΔNp63+ cells exist in ducts while their prevalence increases in CP and in a subset of PDAC. In non-cancer tissue, ΔNp63+ cells are atypical KRT19+ duct cells that overall lack SOX9 expression while they do express canonical basal markers and pertain to a niche of cells expressing gastrointestinal stem cell markers. 3D views show that the basal cells anchor on the basal membrane of normal medium to large ducts while in CP they exist in multilayer dome-like structures. In mice, ΔNp63 is not found in adult pancreas nor in selected models of CP or PDAC, but it is induced in organoids from larger Sox9low ducts. In HPDE, ΔNp63 supports a basal cell phenotype at the expense of a classical duct cell differentiation programme. Conclusion: In larger human pancreatic ducts, basal cells exist. ΔNp63 suppresses duct cell identity. These cells may play an important role in pancreatic disease, including PDAC ontogeny, but are not present in mouse models., SCOPUS: ar.j, info:eu-repo/semantics/published

ObjectiveMolecular taxonomy of tumors is the foundation of personalized medicine and is becoming of paramount importance for therapeutic purposes. Four transcriptomics-based classification systems of pancreatic ductal adenocarcinoma (PDAC) exist, which consistently identified a subtype of highly aggressive PDACs with basal-like features, including ΔNp63 expression and loss of the epithelial master regulator GATA6. We investigated the precise molecular events driving PDAC progression and the emergence of the basal program.DesignWe combined the analysis of patient-derived transcriptomics datasets and tissue samples with mechanistic experiments using a novel dual-recombinase mouse model for Gata6 deletion at late stages of KRasG12D-driven pancreatic tumorigenesis (Gata6LateKO).ResultsThis comprehensive human-to-mouse approach allowed us to show that GATA6 loss is necessary, but not sufficient, for the expression of a basal program in patients and in mice. The concomitant loss of HNF1A and HNF4A, likely through epigenetic silencing, is required for the full phenotype switch. Moreover, Gata6 deletion in mice dramatically increased the metastatic rate, with a propensity for lung metastases. Through RNA-Seq analysis of primary cells isolated from mouse tumors, we show that Gata6 inhibits tumor cell plasticity and immune evasion, suggesting that it works as a barrier for acquiring the fully developed basal and metastatic phenotype.ConclusionsOur work provides both a mechanistic molecular link between the basal phenotype and metastasis and a valuable preclinical tool to investigate the most aggressive subtype of PDAC. These data, therefore, are important for understanding the pathobiological features underlying the heterogeneity of pancreatic cancer in both mice and human.What is already known about this subject?Multiple transcriptomics-based studies have identified a basal-like subtype of pancreatic ductal adenocarcinoma (PDAC) with especially poor prognosis.Loss of GATA6 in PDAC cells is associated with altered differentiation, including ectopic expression of basal markers such as KRT14.Aberrant expression of the ΔNp63 transcription factor can drive the expression of the basal transcriptional program.What are the new findings?Loss of GATA6 expression is necessary but not sufficient for the expression of ΔNp63 and the basal phenotype.Concomitant silencing of HNF4A and HNF1A, possibly through epigenetic mechanisms, is required for the full-blown phenotype.Gata6 deletion in established murine tumors favors the basal and metastatic phenotype, with a lung tropism, in a next-generation model of KRasG12D-driven PDAC.Loss of GATA6 expression is associated with features of immune escape in mouse and human PDAC cells.How might it impact on clinical practice in the foreseeable future?The combined analysis of GATA6, HNFs, and TP63 expression in patient-derived samples will provide a more precise classification of PDAC.Restoration of the classical PDAC phenotype may not only reduce metastatic potential but also increase immune recognition of tumor cells.