Ixazomib inhibited the proliferation of human esophageal squamous carcinoma cells
Ixazomib is an orally available proteasome inhibitor that potently, reversibly, and selectively targets the proteasome. Here, we aimed to evaluate its therapeutic potential toward esophageal squamous carcinoma. Cell proliferation assay and colony formation assay were used to explore the anti-cancer effect of the ixazomib in four commonly used ESCC cell lines. Results showed that ixazomib displays excellent anti-proliferation activity as measured by the CCK-8 assay in ESCC cell lines. Treatment with 1 μM ixazomib inhibited cell proliferation by 40.3%, 27.4% and 13.5% at 24h and 72.5%, 50.1% and 37.4% at 48h for KYSE150, KYSE510, KYSE520, respectively (P < 0.05 for both), but the inhibition of cell proliferation was slightly weaker in Eca109 cell lines than in other cell lines. (Figure 1A). And with an IC50 value at 0.85, 2.63, 10.61 and 42.27 μM for KYSE150, KYSE510, KYSE520 and Eca109 cell line, respectively (Figure 1B). We also found that ixazomib had no effect on proliferation in the esophageal epithelial cell line (HECC). The measurement of cell proliferation demonstrated that Kyse150 and Kyse510 cells were more sensitive to ixazomib treatment, therefore we selected these ESCC cell lines in the following study.
When evaluated by the colony formation assay, high-dose ixazomib (0.5 μM) significantly reduces in the number of colonies by 58.9% and 69.4% for KYSE150 and KYSE510, respectively (P < 0.01 for both) (Figure 1C, D). All these findings demonstrate that ixazomib has anticancer activity toward ESCC.
Ixazomib induced G2/M cell cycle arrest
To understand the underlying mechanisms of antiproliferative activity of ixazomib, we first examined the effect of ixazomib-treated ESCC cells. The Kyse150 and Kyse510 cells were treated with a gradient concentration of 0.25, 0.5 and 1μM of ixazomib for 24 h. The results show that a dose-dependent increase of cells in the G2/M phase indicated the ixazomib induction of a G2/M phase arrest (Figure 2A, B). The results revealed that ixazomib treatment increased the expression of G2/M phase related proteins p21 (Figure 2C). These results suggest that ixazomib has been found to induce cell cycle arrest in G2/M phase in ESCC cells.