Tumour–stroma interactions in an organotypic culture model of pancreatic cancer

Abstract

Pancreatic cancer is characterised by an intense fibrotic, or desmoplastic, stroma, which contributes to tumour progression. Three-dimensional in vitro culture models incorporating this non-tumour component may more closely recapitulate the complex in vivo situation. The aim of my project was to develop a physiologically relevant, three-dimensional organotypic culture model of pancreatic cancer to study the tumour-stroma interactions and its modulation by novel therapeutic agents. Cancer cells cultured on top of collagen/Matrigel gels, embedded with or without stromal cells (hTERT immortalised PS1 stellate cells or MRC5 fibroblasts), differentiated into luminal structures, exhibiting a central apoptotic core with a proliferating peripheral rim and apicobasal polarity. Stromal cells induced a reduction in total tumour cell number, which was associated with a decrease in E-cadherin expression, upregulated β-catenin expression and translocation of ezrin from the apical to the basal aspect of cancer cells, where it was associated with invasive activity. Subsequently, this organotypic model was raised to an air-liquid interface to study the direct and indirect effects of all-trans retinoic acid (ATRA), which rendered stellate cells back to their quiescent phenotype. Indirect effects of quiescent stellate cells on pancreatic cancer cells included changes in proliferation (decrease), apoptosis (increase), invasion (decrease), Wnt/β-catenin signalling (decrease) and an altered morphology. The Wnt/β-catenin signalling 6 perturbations were mediated by restoration of sFRP4 (secreted frizzled-related protein 4) secretion by quiescent stellate cells, resulting in reduced cancer cell invasion (reporter and invasion assays). All such observations could be validated in human pancreatic cancer tissue samples. Taken together, pancreatic organotypic culture offers a reproducible, in vitro three-dimensional culture model, which allows the study of tumour-stroma interactions in a physiologically relevant system. For treatment of pancreatic cancer, a tumour characterised by a poor response to conventional chemotherapeutic drugs, targeting the tumour-stroma cross-talk with agents such as ATRA offers an exciting novel therapeutic strategy.