In basal conditions, Erk is phosphorylated in all cell lines with a low activation in EFO-21 cells, c-Src is phosphorylated in all cell lines except for IGROV-1 and Akt is phosphorylated in all cell lines. the additive effect. A synergistic combination is plotted around the left of the solid collection while an antagonistic combination is usually plotted on the right. Isobolograms were generated with the CompuSyn 1.0 software. (PDF LY 254155 39?kb) 13048_2017_319_MOESM2_ESM.pdf (39K) GUID:?C4C0C498-78FA-4DD4-991A-5CC3CD80DFC8 Additional file 3: In vitro inhibition of HOAC viability by carboplatin alone, or in combination with two kinase inhibitors. HOACs were treated with a dose range of LY 254155 carboplatin alone or in combination with dose range of Crizotinib?+?Dasatinib, Crizotinib?+?Gefitinib, or Dasatinib?+?Gefitinib, based on a ratio of the IC50 of the three drugs. Seventy-two hours after treatment, cell viability was determined by a colorimetric assay using SRB. The unfavorable control (no treatment) of each condition corresponds to the 100% cell viability (Mean +/? SEM, n??3). (PDF 75?kb) 13048_2017_319_MOESM3_ESM.pdf (75K) GUID:?B1B40F59-6C6F-4A52-B611-96B249AA86E3 Additional file 4: In vitro inhibition of HOAC viability kinase inhibitors in tandem. HOACs were treated with a dose range of Crizotinib (Cr), Dasatinib (Da) or Gefitinib (Ge) in tandem, based on a ratio of the IC50 of the two drugs. The IC50 of each drug are plotted around the axes and the circle represents LY 254155 the concentrations of each drug resulting in 50% of cell viability inhibition (Fa?=?0.5). The solid collection represents the additive effect. A synergistic combination is plotted around the left of the solid collection while an antagonistic combination is usually plotted on the right. Isobolograms were generated with the CompuSyn 1.0 software. (PDF 39?kb) 13048_2017_319_MOESM4_ESM.pdf (39K) GUID:?35582D51-F7FF-41EB-AD34-4A25A2421178 Additional file 5: In vitro induction of late apoptosis and necrosis in HOACs by Dasatinib, Gefitinib or LY 254155 a combination of both drugs. HOACs were treated with Dasatinib, Gefitinib (IC50 after 72?h of treatment for each cell collection) or an equieffective combination of both treatments. The unfavorable control corresponds to non-treated cells 48?h after treatment, cells were stained with a FITC-Annexin V/PI apoptosis detection kit. FITC-Annexin staining and PI incorporation were measured in cells with a FACS Canto II circulation cytometer and analyzed with FACS Diva. Late apoptotic and necrotic cells correspond to the Annexin V positive and PI positive populace. (Mean +/? SEM, **?=?test (independent values) for non-parametric data. Each experiment was performed at least three times with independent samples (biological replicates). Results Individual kinase inhibitors induce a moderate cell-specific sensitization of HOAC to carboplatin We aimed to determine if inhibitors of Met, c-Src and EGFR, respectively Crizotinib, Dasatinib or Gefitinib, were able to sensitize HOAC to carboplatin. We decided to work on a panel of carboplatin-sensitive (OVCAR-3, IGROV-1, A2780; IC50 from 13 to 52?M) or carboplatin-resistant (SKOV-3, EFO-21; IC50 from 120 to 935?M) cell lines (Fig.?1a and b). Most of the tested cell lines showed a relative resistance to Crizotinib alone (IC50 from 3.12 to 8.38?M) except for A2780 with a low IC50 of 0.71?M. As for the carboplatin, OVCAR-3, IGROV-1 and A2780 cells were sensitive to Gefitinib alone (IC50 from 4.2 to 7.77?M) whereas SKOV-3 and EFO-21 cells were more resistant (IC50 from 72.66 to 139.87?M). Finally, OVCAR-3 and IGROV-1 cells were sensitive to a treatment with Crizotinib alone with sub-millimolar IC50 RPLP1 (from 0.21 to 0.26?M) contrary to A2780, SKOV-3 and EFO-21 cells (IC50 from 3.29 to 4.37?M). Open in a separate window Fig. 1 In vitro inhibition of HOAC viability by carboplatin or kinase inhibitors in monotherapy. HOACs were treated with a dose range of carboplatin, Crizotinib, Dasatinib or Gefitinib in monotherapy and showed cell-specific sensitivity or resistance. a 72?h after treatment, cell viability was determined by a colorimetric assay using LY 254155 SRB (Mean +/? SEM, n??3). b The IC50 of carboplatin or kinase inhibitors.