title: Integrated molecular analysis to investigate the role of microRNAs in pancreatic tumour growth and progression.
abstract: BACKGROUND
  MicroRNAs (miRNAs) are small non-coding RNAs involved in the post-transcriptional regulation of mRNAs and are aberrantly expressed in cancer with important roles in tumorigenesis. A broad analysis of the combined effects of altered activities of miRNAs in pancreatic ductal adenocarcinoma (PDAC) has not been done, and how miRNAs might affect tumour progression or patient outcomes is unclear.
  
  
  METHODS
  We combined data from miRNA and mRNA expression profiles from PDAC and normal pancreas samples (each n=9) and used bioinformatic analyses to identify a miRNA-mRNA regulatory network in PDAC. We validated our findings in PDAC cell-lines (PANC-1, MIA PaCa-2, LPc006, and LPc167), subcutaneous PDAC xenografts in mice, and laser capture microdissected PDACs from patients (n=91). We used this information to identify miRNAs that contributed most to tumorigenesis.
  
  
  RESULTS
  We identified three miRNAs (miR-21, miR-23a, and miR-27a) that acted as cooperative repressors of a network of tumour suppressor genes that included PDCD4, BTG2, and NEDD4L. Inhibition of miR-21, miR-23a, and miR-27a had synergistic effects in reducing proliferation of PDAC cells in culture and the growth of xenograft tumours. The level of inhibition was greater than that of silencing oncomiR-21 alone. In PDACs from patients, high levels of the combination of miR-21, miR-23a, and miR-27a was a strong independent predictor of short overall survival after surgical resection (hazard ratio 3·21, 95% CI 1·78-5·78). High expression of this combination was also associated with a more aggressive tumour phenotype: more microscopic tumour infiltration at resection margin and increased perineural invasion.
  
  
  CONCLUSIONS
  In an integrated data analysis, we identified functional miRNA-mRNA interactions that contribute to PDAC growth. These findings indicate that miRNAs act together to promote tumour progression and that future therapeutic strategies might require inhibition of several miRNAs. Furthermore, high tumour expression of the miR-21, miR-23a, and miR-27a combination could have potential use in the future as a prognostic signature for patients with PDAC.
  
  
  BACKGROUND
  Peel Medical Research Trust, Alliance Family Foundation, Action Against Cancer, National Institute for Health Research, Association for International Cancer Research, Jason Boas Fellowship, Imperial Biomedical Research Centre, Rosetrees Trust, Joseph Ettedgui Charitable Foundation.
authors: HPB Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, UK. Electronic address: a.frampton@imperial.ac.uk.
authors: Adam E
authors: AE
authors: Frampton
authors: HPB Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Leandro
authors: L
authors: Castellano
authors: Department of Cellular Biotechnology and Haematology, La Sapienza University, Rome, Italy.
authors: Teresa
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authors: Colombo
authors: Department of Medical Oncology, VU University Medical Center, Amsterdam, Netherlands.
authors: Elisa
authors: E
authors: Giovannetti
authors: Division of Oncology, Department of Surgery and Cancer, Imperial Centre for Translational and Experimental Medicine, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Jonathan
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authors: Krell
authors: Division of Oncology, Department of Surgery and Cancer, Imperial Centre for Translational and Experimental Medicine, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Jimmy
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authors: Jacob
authors: Division of Oncology, Department of Surgery and Cancer, Imperial Centre for Translational and Experimental Medicine, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Loredana
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authors: Pellegrino
authors: Division of Oncology, Department of Surgery and Cancer, Imperial Centre for Translational and Experimental Medicine, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Laura
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authors: Roca-Alonso
authors: Experimental and Molecular Oncology, Department of Surgery, University of Pisa, Pisa, Italy.
authors: Niccola
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authors: Funel
authors: HPB Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Tamara M H
authors: TM
authors: Gall
authors: Centre for Pathology, Imperial College Healthcare NHS Trust, Hammersmith Hospital, London, UK.
authors: Raida
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authors: Ahmad
authors: HPB Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Nagy A
authors: NA
authors: Habib
authors: Institute of Pathology, Ludwig-Maximilians-University, Munich, Germany.
authors: Thomas
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authors: Knösel
authors: Division of Oncology, Department of Surgery and Cancer, Imperial Centre for Translational and Experimental Medicine, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Justin
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authors: Stebbing
authors: HPB Surgical Unit, Department of Surgery and Cancer, Imperial College, Hammersmith Hospital Campus, London, UK.
authors: Long R
authors: LR
authors: Jiao