Figure 2.

The combination of vincristine and ALK inhibitors enhances the anti-proliferative effect in EML4-ALK V1 cells, but not in V3. ALK-positive lung cancer cells, H3122 (V1) and H2228 (V3), were treated with increasing doses of (A,C) crizotinib −/+ vincristine (0.5 or 1 nM), or (B,D) ceritinib −/+ vincristine (0.5 or 1 nM) for 72 h. Cell viability was determined using CellTiter-Glo assays. The IC₅₀ values were calculated using the Prism 9.0 software. Data represent the mean of three biological replicates in each column; the bars denote ±SD. (E,F) Table summarises the IC₅₀ of the combination of vincristine and ALK inhibitors in H3122 and H2228 cells. (G,H) H3122 and H2228 cells were treated with either vincristine, ceritinib or in combination for 48 h before analysis by annexin V-based flow cytometry. Histograms represent the percentage of cells in apoptosis. Cells were classified as early apoptotic, late apoptotic and dead. Data represent the mean of three biological replicates; the bars denote ±SD. * p < 0.5 in comparison to DMSO by two-way ANOVA. (I) H3122 and H2228 cells were treated with either single agents of vincristine or ceritinib or in combination for 24 h before being fixated and stained with cleaved caspase-3 (green), anti-α-tubulin (red), and DAPI (blue). Scale bars, 20 μm. (J). Crystal violet staining of H3122 and H2228 cells. Cells were treated with the indicated drugs for 72 h before fixed and stained. ns= not significant.