Fig. 6

BTZ and PD0166285 has a synergistic effect in suppressing growth of subcutaneous tumor xenografts. a, b BP-Combo showed much stronger effect than BTZ or PD0166285 monotreatment in repressing colony formation of tumor cells. SNU449 and HeLa cells were treated with vehicle, BTZ or PD0166285 alone, or with BP-Combo for 10 days before staining with 0.1% crystal violet. The representative images (a) and colony quantification (b) are shown. Scale bar, 2 mm. Error bars: SEM from at least three independent experiments. c–g BP-Combo showed much stronger effect than BTZ or PD0166285 monotreatment in inhibiting subcutaneous tumor xenograft development. The BALB/c nude mice were subcutaneously injected with HeLa cells, and then intraperitoneally injected with the indicated inhibitors one day after tumor cell implantation (early treatment, c), or when tumor volumes reached ~50 mm³ (late treatment, d, e). Early treatment: n = 5 (vehicle), 6 (BTZ), 6 (PD0166285) and 6 (BP-Combo). Late treatment: n = 3 (vehicle), 3 (BTZ), 3 (PD0166285) and 4 (BP-Combo). For f, g, C57BL/6J mice were subcutaneously injected with Hepa1-6 cells, and then intraperitoneally injected with the indicated inhibitors when tumor volumes reached ~50 mm³. n = 4 for each group. h BP-Combo had much stronger effect than BTZ or PD0166285 monotreatment in increasing pH3-S10 level and inducing GSDME cleavage in tumor xenografts. Mouse xenograft tissues from late treatment groups in Figures d and e were analyzed. i, j Silencing of GSDME diminished the anti-tumor effect of BP-Combo. C57BL/6J mice were subcutaneously injected with Hepa1-6-shNC or Hepa1-6-shGsdme cells, and then intraperitoneally administered with vehicle or BP-Combo treatment when tumor volumes reached ~50 mm³. n = 3 for each group. One-way ANOVA (b, c, e, g) and two-way ANOVA (d, f, i, j) were used. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001; ns not significant