FGF10/FGFR2 Signaling: Therapeutically Targetable Vulnerability in Ligand-responsive Cholangiocarcinoma Cells.

BACKGROUND/AIM: Increasing evidence has revealed FGFR2 as an attractive therapeutic target for cancer including cholangiocarcinoma (CCA). The present study investigated the oncogenic mechanisms by which FGF10 ligand activates FGFR2 in CCA cells and determined whether FGFR inhibitors could suppress FGF10-mediated migration of CCA cells. MATERIALS AND METHODS: Effects of FGF10 on the proliferation, migration, and invasion of KKU-M213A cells were assessed using clonogenic and transwell assays. Protein expression levels of FGFR2 and pro-angiogenic factors were determined via immunoblotting and antibody array analysis. FGFR2 knockdown using a small interfering RNA was used to validate the role of FGF10 in promoting cell migration via FGFR2. The effects of infigratinib (FGFR inhibitor) on cell viability, were determined in KKU-100, KKU-M213A, KKU-452 cells. Moreover, the efficacy of the FGFR inhibitor in suppressing migration via FGF10/FGFR2 stimulation was assessed in KKU-M213A cells. RESULTS: FGF10 significantly increased the expression of phospho-FGFR/FGFR2 and promoted the proliferation, migration, and invasion of KKU-M213A cells. FGF10 increased the expression levels of p-Akt, p-mTOR, VEGF, Slug, and pro-angiogenic proteins related to metastasis. Cell migration mediated by FGF10 was markedly decreased in FGFR2-knockdown cells. Moreover, FGF10/FGFR2 promoted the migration of cells, which was suppressed by the FGFR inhibitor. CONCLUSION: FGF10/FGFR2 activates the Akt/mTOR and VEGF/Slug pathways, which are associated with the stimulation of migration and invasion in CCA. Moreover, the FGF10/FGFR2 signaling was inhibited by an FGFR inhibitor resulting suppression of cell migration, which warrants further studies on their clinical utility for CCA treatment.

Inhibition of FGFR2 enhances chemosensitivity to gemcitabine in cholangiocarcinoma through the AKT/mTOR and EMT signaling pathways.

AIM: To investigate the oncogenic role of FGFR2 in carcinogenesis in cholangiocarcinoma (CCA) cells. In addition, the feasibility of using FGFR inhibitors in combination with standard chemotherapy was also explored for the chemosensitizing effect in CCA cells. MAIN METHODS: Five CCA cell lines were used to screen FGFR2 expression by Western immunoblotting. Two CCA cell lines, KKU-100 and KKU-213A, were knocked down of the FGFR2 gene using siRNA. Cell viability was assessed by the MTS cell proliferation assay. Reproductive cell death was assessed by clonogenic assay. The effects on cell migration and invasion were analyzed by the Transwell chamber method. Cell cycle analysis was performed by flow cytometry. Cell angiogenesis was assessed by HUVEC tube formation and human angiogenesis antibody array analysis. Proteins associated with proliferative and metastatic properties were evaluated by Western blotting. KEY FINDINGS: Knockdown of FGFR2 suppressed cell growth and colony formation in CCA cells in association with G2/M cell cycle arrest and downregulation of STAT3, cyclin A and cyclin B1. Silencing FGFR2 enhanced the suppressive effect of gemcitabine (Gem) on cell migration and invasion. The combination of infigratinib, an FGFR inhibitor, and Gem, interrupted cell growth, migration, and invasion via downregulation of FGFR/AKT/mTOR pathways and the EMT-associated proteins vimentin and slug. Moreover, the combination also suppressed tube formation together with decreased expression of the proangiogenic factor VEGF. SIGNIFICANCE: Inhibition of FGFRs by infigratinib enhanced the antitumor effect of Gem in CCA cells through downregulation of the FGFR/AKT/mTOR, FGFR/STAT3 and EMT signaling pathways.

Phenformin inhibits proliferation, invasion, and angiogenesis of cholangiocarcinoma cells via AMPK-mTOR and HIF-1A pathways.

Phenformin (Phen), a potent activator of AMPK, is effective against some resistant cancers. This study evaluated the inhibition of proliferation, migration, invasion, and angiogenesis by Phen in aggressive cancer cells and investigated the underlying mechanism of the inhibition. Cholangiocarcinoma (CCA) KKU-156 and KKU-452 cells were used in this study. The results showed that Phen suppressed cell proliferation and induced apoptosis in both cells. Phen suppressed migration and invasion of cancer cells in wound healing and transwell chamber assays, respectively. The effects were associated with depletions of glutathione (GSH) and decreased glutathione redox ratio which represents cellular redox state. The redox stress was linked with the loss of mitochondrial transmembrane potential, as evaluated by JC-1 assay. The effect of Phen on angiogenesis was performed using HUVEC cultured cells. Phen alone did not affect tube formation of HUVEC cells. However, conditioned media from CCA cell cultures treated with Phen suppressed the tube-like structure formation. The antitumor effect of Phen was associated with AMPK activation and suppression of mTOR phosphorylation, HIF-1A, and VEGF protein expression. In conclusion, Phen inhibits cell proliferation, migration, invasion, and angiogenesis probably through AMPK-mTOR and HIF-1A-VEGF pathways. Phen may be repurposed as chemoprevention of cancer.