Fig. 2

Effects of refametinib and pictilisib alone and in combination on colonies formation and on EGFR-dependent intracellular signaling in parental SW48 and LIM1215 human colon cancer cell lines and in their HER2-amplified derivatives (SW48-HER2 and LIM1215-HER2). a Both parental and HER2-amplified derivatives cells were treated with different concentrations of refametinib and pictilisib as single agents (range, 0.05 to 10 μM), for 96 h and evaluated byclonogenic assay. Data were represented as cell colony area percentage calculated by ImageJ software after staining with crystal violet, as described in Materials and Methods. b SW48-HER2 and LIM1215-HER2 cell lines were treated with lapatinib, trastuzumab, refametinib and pictilisib at 0.5 μM, as single agent and in their possible combinations (**p < 0.01 compared to other treatments). Colony area percentage was presented in the Table. c SW48-HER2 and LIM1215-HER2 cells were treated with refametinib and pictilisib (0.5 μM) alone and in combination for 24 h. Cell protein extracts were subjected to immunoblotting with the indicate antibodies, as described in Materials and Methods. α-Tubulin was used as the loading control. Results represent the mean of three separate experiments, each performed in duplicate