Effects of combined inhibition of STAT3 and VEGFR2 pathways on the radiosensitivity of non-small-cell lung cancer cells

Abstract

Purpose: The goals of this study were to determine the effects of combined inhibition of STAT3 and vascular endothelial growth factor receptor 2 (VEGFR2) pathways on the radiosensitivity of non-small-cell lung cancer (NSCLC) cells, and to assess the underlying mechanisms. Methods: The expressions of VEGFR2, STAT3, related signaling molecules, hypoxia-inducible factor 1-alpha (HIF-1α), and cyclin D1 were determined by Western blotting. Radiosensitivity was assessed using the colony-forming assay, and cell cycle and cell death were analyzed by flow cytometry. A nude mouse xenograft tumor model of Calu-1 cells was established. The hepatorenal toxicity of the above-mentioned treatment on tumor-bearing mice was observed by H&E staining. The expression of STAT3, VEGFR2, HIF-1α, and cyclin D1 of the transplanted tumor tissues was detected by immunohistochemistry. Apoptosis of tumor tissues was evaluated by TUNEL staining. Results: In vitro, we selected two cell lines with high expression levels of STAT3, including Calu-1 cells that exhibit high VEGFR2 expression and A549 cells that exhibit low VEGFR2 expression. When apatinib treatment was combined with S3I-201, the expression of VEGFR2, STAT3, and their downstream signaling molecules was significantly decreased (P<0.01). There was an increase in cell death and G2/M phase arrest after treatments, with the most significant changes occurring upon dual inhibition of STAT3 and VEGFR2 (P<0.01). In vivo, combined treatment of radiotherapy and dual inhibition of VEGFR2 and STAT3 was well tolerated and did not deliver additional toxicity. Compared with the control group and the radiation treatment (RT) + apatinib or RT + S3I-201 duplex group, the expression level of STAT3, p-STAT3, VEGFR2, HIF-1α, and cyclin D1 in the triple group (RT + apatinib + S3I-201) was the lowest, and the proportion of apoptotic cells was the highest (P<0.05). Conclusion: The combined inhibition of VEGFR2 and STAT3 is effective in enhancing radiosensitizing effects in NSCLC cells.