Ectopic expression of HNF4α in Het1A cells induces an invasive phenotype.

Barrett's oesophagus (BO) is a pathological condition in which the squamous epithelium of the distal oesophagus is replaced by an intestinal-like columnar epithelium originating from the gastric cardia. Several somatic mutations contribute to the intestinal-like metaplasia. Once these have occurred in a single cell, it will be unable to expand further unless the altered cell can colonise the surrounding squamous epithelium of the oesophagus. The mechanisms by which this happens are still unknown. Here we have established an in vitro system for examining the competitive behaviour of two epithelia. We find that when an oesophageal epithelium model (Het1A cells) is confronted by an intestinal epithelium model (Caco-2 cells), the intestinal cells expand into the oesophageal domain. In this case the boundary involves overgrowth by the Caco-2 cells and the formation of isolated colonies. Two key transcription factors, normally involved in intestinal development, HNF4α and CDX2, are both expressed in BO. We examined the competitive ability of Het1A cells stably expressing HNF4α or CDX2 and placed in confrontation with unmodified Het1A cells. The key result is that stable expression of HNF4α, but not CDX2, increased the ability of the cells to migrate and push into the unmodified Het1A domain. In this situation the boundary between the cell types is a sharp one, as is normally seen in BO. The experiments were conducted using a variety of extracellular substrates, which all tended to increase the cell migration compared to uncoated plastic. These data provide evidence that HNF4α expression could have a potential role in the competitive spread of BO into the oesophagus as HNF4α increases the ability of cells to invade into the adjacent stratified squamous epithelium, thus enabling a single mutant cell eventually to generate a macroscopic patch of metaplasia.

Extracellular Vesicles Inhibit the Response of Pancreatic Ductal Adenocarcinoma Cells to Gemcitabine and TRAIL Treatment.

Pancreatic ductal adenocarcinoma remains an aggressive cancer with a low 5-year survival rate. Although gemcitabine has been a standard treatment for advanced pancreatic cancer, patients often develop resistance to this therapeutic. We have previously shown that treating pancreatic cancer cells in vitro with a combination of gemcitabine and the cytokine TRAIL significantly reduced both cell viability and survival. The data presented here demonstrate that this response to treatment is inhibited when cells are incubated with a conditioned medium derived from untreated cells. We show that this inhibition is specifically mediated by extracellular vesicles present in the conditioned medium, as seen by a significant decrease in apoptosis. Additionally, we further demonstrate that this effect can be reversed in the presence of GW4869, an inhibitor of exosome biogenesis and release. These results show that pancreatic cancer cell-derived extracellular vesicles can confer resistance to treatment with gemcitabine and TRAIL. The implications of these findings suggest that removal of EVs during treatment can improve the response of cells to gemcitabine and TRAIL treatment in vitro.

Crosstalk with lung fibroblasts shapes the growth and therapeutic response of mesothelioma cells.

Mesothelioma is an aggressive cancer of the mesothelial layer associated with an extensive fibrotic response. The latter is in large part mediated by cancer-associated fibroblasts which mediate tumour progression and poor prognosis. However, understanding of the crosstalk between cancer cells and fibroblasts in this disease is mostly lacking. Here, using co-cultures of patient-derived mesothelioma cell lines and lung fibroblasts, we demonstrate that fibroblast activation is a self-propagated process producing a fibrotic extracellular matrix (ECM) and triggering drug resistance in mesothelioma cells. Following characterisation of mesothelioma cells/fibroblasts signalling crosstalk, we identify several FDA-approved targeted therapies as far more potent than standard-of-care Cisplatin/Pemetrexed in ECM-embedded co-culture spheroid models. In particular, the SRC family kinase inhibitor, Saracatinib, extends overall survival well beyond standard-of-care in a mesothelioma genetically-engineered mouse model. In short, we lay the foundation for the rational design of novel therapeutic strategies targeting mesothelioma/fibroblast communication for the treatment of mesothelioma patients.