Fig. 4

Targeting IL8 overcomes tamoxifen resistance. A FACS analysis of the proportion of ALDH + BCSCs in the matched MCF-7 cells and tamoxifen-resistant MCF-7 TamR cells. B Immunoblot analysis of IL8 levels in the matched MCF-7 cells and tamoxifen-resistant MCF-7 TamR cells. C The matched MCF-7 or MCF-7 TamR cells transfected with vector or IL8 shRNA were treated with a graded concentration of tamoxifen for 5 days. Cell viability was determined by MTT assay. D The matched MCF-7 or MCF-7 TamR cells were treated with a graded concentration of tamoxifen for 5 days in the presence or absence of 0.1 mM repertaxin. Cell viability was determined by MTT assay. E Colony formation assay of MCF-7 or MCF-7 TamR cells treated with treated with 5 μM tamoxifen and in the presence or absence of 0.1 mM repertaxin. F The growth curves (left) or image (right) of tumors derived from MCF-7 TamR cells for 30 days. The mice were treated with slow-release tamoxifen supplement alone, repertaxin alone, or combined tamoxifen and repertaxin. G The representative image (left) and statistical result (right) of Ki-67 staining and TUNEL labeling of the respective MCF-7 TamR cells-derived tumor sections as indicated. Scale Bar: 100 μm. H RNA-sequencing analysis of the CCND1 levels in MCF-7 cells cultured with CM fom BCSCs or parental cells. I Immunoblot analysis of Cyclin D1, pRb and RB levels in MCF-7 cells cultured with the indicated CM. J MCF-7 cells were cultured with CM derived from parental cells or BCSCs in the presence or absence of 0.1 mM repertaxin and treated with a graded concentration of palbociclib. Cell viability was determined by MTT assay. K Colony formation assay MCF-7 cells cultured with control CM or BCSC CM in the presence or absence of palbociclib. L, M MTT L or colony formation assay M of MCF-7 parental, TamR cells in the presence or absence of 0.1 mM repertaxin and treated with 1 μM fulvestrant and/or palbociclib. N Tumor growth curve of MCF-7 TamR cells treated with fulvestrant/ palbociclib and/or repertaxin. All experiments were repeated at least three times. Results are shown as mean ± S.D. *P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant (Unpaired two-tailed Student’s t-test in A, H, one-way ANOVA followed by Tukey’s multiple comparison in G, L, others Two-way ANOVA test)