Abstract
Abstract
Background
Cholangiocarcinoma (CCA) is still a deadly tumour. Histological and molecular aspects of thioacetamide (TAA)-induced intrahepatic CCA (iCCA) in rats mimic those of human iCCA. Carcinogenic changes and therapeutic vulnerabilities in CCA may be captured by molecular investigations in bile, where we performed bile proteomic and metabolomic analyses that help discovery yet unknown pathways relevant to human iCCA.
Methods
Cholangiocarcinogenesis was induced in rats (TAA) and mice (JnkΔhepa + CCl4 + DEN model). We performed proteomic and metabolomic analyses in bile from control and CCA-bearing rats. Differential expression was validated in rat and human CCAs. Mechanisms were addressed in human CCA cells, including Huh28-KRASG12D cells. Cell signaling, growth, gene regulation and [U-13C]-D-glucose-serine fluxomics analyses were performed. In vivo studies were performed in the clinically-relevant iCCA mouse model.
Results
Pathways related to inflammation, oxidative stress and glucose metabolism were identified by proteomic analysis. Oxidative stress and high amounts of the oncogenesis-supporting amino acids serine and glycine were discovered by metabolomic studies. Most relevant hits were confirmed in rat and human CCAs (TCGA). Activation of interleukin-6 (IL6) and epidermal growth factor receptor (EGFR) pathways, and key genes in cancer-related glucose metabolic reprogramming, were validated in TAA-CCAs. In TAA-CCAs, G9a, an epigenetic pro-tumorigenic writer, was also increased. We show that EGFR signaling and mutant KRASG12D can both activate IL6 production in CCA cells. Furthermore, phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme in serine-glycine pathway, was upregulated in human iCCA correlating with G9a expression. In a G9a activity-dependent manner, KRASG12D promoted PHGDH expression, glucose flow towards serine synthesis, and increased CCA cell viability. KRASG12D CAA cells were more sensitive to PHGDH and G9a inhibition than controls. In mouse iCCA, G9a pharmacological targeting reduced PHGDH expression.
Conclusions
In CCA, we identified new pro-tumorigenic mechanisms: Activation of EGFR signaling or KRAS mutation drives IL6 expression in tumour cells; Glucose metabolism reprogramming in iCCA includes activation of the serine-glycine pathway; Mutant KRAS drives PHGDH expression in a G9a-dependent manner; PHGDH and G9a emerge as therapeutic targets in iCCA.
Funder
CIBERehd
Instituto de Salud Carlos III
Fundación Científica Asociación Española Contra el Cáncer
Gobierno de Navarra
Interdisziplinäres Zentrum für Klinische Forschung, Universitätsklinikum Aachen
Sonderforschungsbereiche Transregio
Sonderforschungsbereiche (SFB)
Deutsche Forschungsgemeinschaft
Deutsche Krebshilfe
Junta de Castilla y León
Comunidad de Madrid
Ministerio de Ciencia, Innovación y Universidades
MCIU/AEI/FEDER
Eusko Jaurlaritza
Bio-Eusko Fundazioa
Euskadi RIS3
Fundación La Caixa
ERAB: The European Foundation for Alcohol Research
Unión Iberoamericana de Universidades
Fundación Eugenio Rodríguez Pascual
Fundación Echébano
Fundació Mataro TV3
Fundación Mario Losantos
Fundación M Torres
AMMF: The Cholangiocarcinoma Charity
Fundación BBVA
COST Action Euro-cholangio-Net
Miguel Servet
PSC Partners US and PSC Supports UK
Horizon 2020 Research and Innovation Program
Ramón y Cajal Program
Publisher
Springer Science and Business Media LLC
Cited by
8 articles.
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