Radiosensitizing And Its Mechanism Study Of -et Small Molecule Tyrosine Kinase Inhibitor ARQ197 On Lung Adenocarcinoma Cell Line

Objective: To explore whether and how c-Met small molecule tyrosine kinase inhibitor ARQ197 could enhance radiosensitivity of human lung adenocarcinoma cell. The aim is to provide experimental foundation that :ARQ197 as a radiosensitizer could be used for the therapeutic alliance of lung adenocarcinoma.Methods : We used human lung adenocarcinoma cell lines A549 and H1299 in vitro as research obsject. Lung adenocarcinoma cells were treated by different series of concentration of HGF and ARQ197, respectively. Cloning formation assay was used to investigate the proliferation of cells, and the appropriate concentration of HGF and ARQ197 were selected for the next step research of radiosensitization. Cells were divided into four groups as follow:control,HGF-treated, ARQ197-treated, and HGF+ ARQ197-treated group. Cell survival curves after irradiation were drawn by using cloning formation assay, and the radiosensitizing differences among different groups were evaluated. Then cells were treated by HGF alone or in combination with different series of concentration of ARQ197, respectively.The the impact of HGF alone or in combination with ARQ197 on phosphorylation and expression levels of both c-Met and its downstream signaling molecules Akt, and Erk1/2 were assessed by Western blottings. Lung adenocarcinoma cells were divided into two groups as follow: control, ARQ197-treated group. The changing of phosphorylation and expression levels of both c-Met and its downstream signaling molecules Akt, and Erk1/2 before and after irradiation in two groups were assessed by Western blotting.Results:The cloning formation rate increases with increasing HGF concentration, and decreases with increasing ARQ197 concentration, respectively.The cell survival curve showed that the SF2 values of control, HGF-treated, HGF+ARQ197-treated, and ARQ197-treated group,A549 cell were 48.28±2.16%、82±2.62%、41.53±1.71%和 38.91±1.71%,H1299 cell were 58.41.±3.47%、85.23±6.07%、44.80±3.47%和 43.30±2.96%..The radiation dose enhancement ratio(DER) of HGF、HGF+ARQ197、ARQ197 for A549 cells were 0.79、1.11、1.19, for H1299 cells were 0.85、1.20、1.27,respectively.The Western blot showed that the expression of p-c Met, p-Akt and p-Erk1/2 protein increase after HGF stimulation,but combinating with ARQ197, with increasing ARQ197 concentration, the expression of p-c Met, p-Akt, p-Erk1/2 protein decrease.After 2Gy radiation,the expression of p-c Met, p-Akt and p-Erk1/2 protein increase, but decrease significantly in combination with ARQ197,no significant changes in the expression of total c-Met, Akt and Erk1/2 protein.Conclusion:ARQ197 could increase the radiation sensitivity of lung adenocarcinoma cell according to colony-forming assay.ARQ197 has significantly radiosensitive effect on lung adenocarcinoma cell line by inhibiting c-Met phosphorylation and downstream c-Met signaling pathways in vitro according to Western blot.So the study was supposed to provide experimental foundation that :ARQ197 as a radiosensitizer could be used for the therapeutic alliance of lung adenocarcinoma.