Adrián Vallejo1, Oihane Erice2, Rodrigo Entrialgo-Cadierno1, Iker Feliu1, Elizabeth Guruceaga3, Maria J Perugorria4, Paula Olaizola5, Alexandra Muggli6, Irati Macaya1, Michael O'Dell7, Borja Ruiz-Fernandez de Cordoba1, Sergio Ortiz-Espinosa1, Aram F Hezel7, Imanol Arozarena8, Fernando Lecanda9, Matias A Avila10, Maite G Fernandez-Barrena10, Matthias Evert6, Mariano Ponz-Sarvise11, Diego F Calvisi6, Jesus M Banales12, Silve Vicent13. 1. University of Navarra, Centre for Applied Medical Research, Program in Solid Tumours, Pamplona, Spain. 2. University of Navarra, Centre for Applied Medical Research, Program in Solid Tumours, Pamplona, Spain; IdiSNA, Navarra Institute for Health Research, Pamplona, Spain. 3. IdiSNA, Navarra Institute for Health Research, Pamplona, Spain; University of Navarra, Centre for Applied Medical Research, Computational Biology Program, Pamplona, Spain; ProteoRed-Instituto de Salud Carlos III (ISCIII), Madrid, Spain. 4. University of the Basque Country, San Sebastian, Spain; Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute - Donostia University Hospital -, University of the Basque Country (UPV/EHU), San Sebastian, Spain; National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd, Instituto de Salud Carlos III), Spain. 5. Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute - Donostia University Hospital -, University of the Basque Country (UPV/EHU), San Sebastian, Spain. 6. Institute of Pathology, University of Regensburg, Regensburg, Germany. 7. University of Rochester Medical Centre, Rochester, NY, USA. 8. IdiSNA, Navarra Institute for Health Research, Pamplona, Spain; Cancer Signalling Unit, Navarrabiomed, Complejo Hospitalario de Navarra (CHN), Universidad Pública de Navarra (UPNA), Pamplona, Spain. 9. University of Navarra, Centre for Applied Medical Research, Program in Solid Tumours, Pamplona, Spain; IdiSNA, Navarra Institute for Health Research, Pamplona, Spain; Centro de Investigación Biomédica en Red de Cáncer (CIBERONC, Instituto de Salud Carlos III), Madrid, Spain; University of Navarra, Department of Pathology, Anatomy and Physiology, Pamplona, Spain. 10. IdiSNA, Navarra Institute for Health Research, Pamplona, Spain; National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd, Instituto de Salud Carlos III), Spain; University of Navarra, Centre for Applied Medical Research, Hepatology Program, Pamplona, Spain. 11. Clinica Universidad de Navarra, Department of Medical Oncology, Pamplona, Spain. 12. Department of Liver and Gastrointestinal Diseases, Biodonostia Health Research Institute - Donostia University Hospital -, University of the Basque Country (UPV/EHU), San Sebastian, Spain; National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd, Instituto de Salud Carlos III), Spain; Ikerbasque, Basque Foundation for Sciences, Bilbao, Spain. 13. University of Navarra, Centre for Applied Medical Research, Program in Solid Tumours, Pamplona, Spain; IdiSNA, Navarra Institute for Health Research, Pamplona, Spain; Centro de Investigación Biomédica en Red de Cáncer (CIBERONC, Instituto de Salud Carlos III), Madrid, Spain; University of Navarra, Department of Pathology, Anatomy and Physiology, Pamplona, Spain. Electronic address: silvevicent@unav.es.
Abstract
BACKGROUND & AIMS: Cholangiocarcinoma (CCA) is a neoplasia of the biliary tract driven by genetic, epigenetic and transcriptional mechanisms. Herein, we investigated the role of the transcription factor FOSL1, as well as its downstream transcriptional effectors, in the development and progression of CCA. METHODS: FOSL1 was investigated in human CCA clinical samples. Genetic inhibition of FOSL1 in human and mouse CCA cell lines was performed in in vitro and in vivo models using constitutive and inducible short-hairpin RNAs. Conditional FOSL1 ablation was done using a genetically engineered mouse (GEM) model of CCA (mutant KRAS and Trp53 knockout). Follow-up RNA and chromatin immunoprecipitation (ChIP) sequencing analyses were carried out and downstream targets were validated using genetic and pharmacological inhibition. RESULTS: An inter-species analysis of FOSL1 in CCA was conducted. First, FOSL1 was found to be highly upregulated in human and mouse CCA, and associated with poor patient survival. Pharmacological inhibition of different signalling pathways in CCA cells converged on the regulation of FOSL1 expression. Functional experiments showed that FOSL1 is required for cell proliferation and cell cycle progression in vitro, and for tumour growth and tumour maintenance in both orthotopic and subcutaneous xenograft models. Likewise, FOSL1 genetic abrogation in a GEM model of CCA extended mouse survival by decreasing the oncogenic potential of transformed cholangiocytes. RNA and ChIP sequencing studies identified direct and indirect transcriptional effectors such as HMGCS1 and AURKA, whose genetic and pharmacological inhibition phenocopied FOSL1 loss. CONCLUSIONS: Our data illustrate the functional and clinical relevance of FOSL1 in CCA and unveil potential targets amenable to pharmacological inhibition that could enable the implementation of novel therapeutic strategies. LAY SUMMARY: Understanding the molecular mechanisms involved in cholangiocarcinoma (bile duct cancer) development and progression stands as a critical step for the development of novel therapies. Through an inter-species approach, this study provides evidence of the clinical and functional role of the transcription factor FOSL1 in cholangiocarcinoma. Moreover, we report that downstream effectors of FOSL1 are susceptible to pharmacological inhibition, thus providing new opportunities for therapeutic intervention.
BACKGROUND & AIMS: Cholangiocarcinoma (CCA) is a neoplasia of the biliary tract driven by genetic, epigenetic and transcriptional mechanisms. Herein, we investigated the role of the transcription factor FOSL1, as well as its downstream transcriptional effectors, in the development and progression of CCA. METHODS: FOSL1 was investigated in human CCA clinical samples. Genetic inhibition of FOSL1 in human and mouse CCA cell lines was performed in in vitro and in vivo models using constitutive and inducible short-hairpin RNAs. Conditional FOSL1 ablation was done using a genetically engineered mouse (GEM) model of CCA (mutant KRAS and Trp53 knockout). Follow-up RNA and chromatin immunoprecipitation (ChIP) sequencing analyses were carried out and downstream targets were validated using genetic and pharmacological inhibition. RESULTS: An inter-species analysis of FOSL1 in CCA was conducted. First, FOSL1 was found to be highly upregulated in human and mouse CCA, and associated with poor patient survival. Pharmacological inhibition of different signalling pathways in CCA cells converged on the regulation of FOSL1 expression. Functional experiments showed that FOSL1 is required for cell proliferation and cell cycle progression in vitro, and for tumour growth and tumour maintenance in both orthotopic and subcutaneous xenograft models. Likewise, FOSL1 genetic abrogation in a GEM model of CCA extended mouse survival by decreasing the oncogenic potential of transformed cholangiocytes. RNA and ChIP sequencing studies identified direct and indirect transcriptional effectors such as HMGCS1 and AURKA, whose genetic and pharmacological inhibition phenocopied FOSL1 loss. CONCLUSIONS: Our data illustrate the functional and clinical relevance of FOSL1 in CCA and unveil potential targets amenable to pharmacological inhibition that could enable the implementation of novel therapeutic strategies. LAY SUMMARY: Understanding the molecular mechanisms involved in cholangiocarcinoma (bile duct cancer) development and progression stands as a critical step for the development of novel therapies. Through an inter-species approach, this study provides evidence of the clinical and functional role of the transcription factor FOSL1 in cholangiocarcinoma. Moreover, we report that downstream effectors of FOSL1 are susceptible to pharmacological inhibition, thus providing new opportunities for therapeutic intervention.