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Novel organotypic cell culture models and 3D spheroid cultures containing pancreas-derived CAFs have been developed in order to investigate the complex interactions between CAFs and PDAC tumour cells. Traditional 2D monolayer-based assays have played a key role in the drug discovery process for decades and are still in use as they provide a cost-effective tool that is fast and easily adapted for high-throughput screening strategies. The failure rate of drugs tested in phase III clinical trials for PDAC is high, with a large discrepancy between the behaviour of drugs or drug combinations pre-clinically and their performance in randomised control clinical trials. The combination of gemcitabine and nanoparticle albuminвЂ“bound paclitaxel (nab-paclitaxel) is a standard of care option for patients with advanced disease. One such other agent is paclitaxel, a microtubule stabilising molecule which has been used to treat many solid tumours. Gemcitabine was approved for use in metastatic PDAC in 1996 and is still used today for the treatment of patients with resectable or advanced PDAC, either as a single agent or in combination with other agents. They therefore should be considered early in the drug development process in order to maximise the probability of clinical success. Additionally, CAFs have been reported to scavenge gemcitabine, a nucleoside analogue, used to treat PDAC, thus reducing the availability of the drug to cancer cells. For example, the extracellular matrix deposited by CAFs forms a physical barrier to chemotherapy and mediates increased interstitial fluid pressure within the tumour causing impaired vascular function. CAFs also play a significant role in chemoresistance through a variety of mechanisms. These cells drive the fibroinflammatory reaction through the deposition of extracellular matrix and the secretion of growth factors and cytokines. The most abundant cell type in PDAC stroma is CAFs, which were first described in 1998. PDAC-associated fibroinflammatory stroma harbours a complex array of activated cancer-associated fibroblasts (CAFs), immune cells, endothelial cells and extracellular matrix proteins such as collagen and fibronectin. A feature of PDAC, which presents a significant barrier to therapeutic intervention, is its dense tumour microenvironment. Poor survival is due in part to a lack of effective therapies with many clinical trials of potential PDAC treatments failing. Pancreatic cancer is the 4 th leading cause of cancer deaths and has an overall 5-year survival of 7вЂ“9%.

Changes to standard 2D mono-culture-based assays and implementation of 3D co-culture assays lend complexity to established models and could provide tools for identifying therapies that will match clinically the success observed with in vitro models, thereby aiding in the discovery of novel therapies. We also identify that a 3D co-culture model of MIA PaCa-2 or PANC-1 with CAFs showed an increased chemoresistance to gemcitabine when compared to standard 2D mono-cultures a difference to paclitaxel which showed no measurable difference between the 2D and 3D models, suggesting a complex interaction between the drug in study and the cell type used. However, CAFs were less sensitive to pulsing with gemcitabine compared to their response to paclitaxel. Following the use of an in vitro drug pulsing experiment, PDAC cell lines showed sensitivity to gemcitabine and paclitaxel. Here, we reveal that gemcitabine and paclitaxel markedly reduce the viability of pancreatic cell lines, but not CAFs, when cultured in 2D.

The objective of this study was to investigate whether changes in the drug-dosing regimen or the addition of cancer-associated fibroblasts (CAFs) to current existing models can impact the efficacy of chemotherapy drugs used in the clinic. Promising therapies that pass pre-clinical testing often fail in pancreatic cancer clinical trials. However, the models used to test new and emerging therapies for PDAC have not increased in complexity to keep pace with our understanding of the human disease. Pancreatic ductal adenocarcinoma (PDAC) is characterised by a pro-inflammatory stroma and multi-faceted microenvironment that promotes and maintains tumorigenesis.

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