Fig. 1

BCSC secretome confers tamoxifen resistance. A Top 10 enriched signature gene sets representing RNA-seq data from MCF-7 cells cultured with control CM or BCSC CM. B Enrichment of upregulated ESR1 target genes upon culture of MCF-7 cells with BCSC CM by GSEA. C Immunoblot showing the phosphorylation of ERα in MCF-7 cells with BCSC CM. D The ERα transcriptional activity of estrogen response element (ERE4) in MCF-7 cells with BCSC CM. E Enrichment of upregulated genes involved in endocrine therapy resistance or tamoxifen resistance upon culture of MCF-7 cells with BCSC CM by GSEA. F MCF-7 or T47D cells were cultured in CM derived from respective mammosphere-enriched BCSCs or parental cells and treated with a graded concentration of tamoxifen for 5 days. Cell viability was determined by MTT assay. G MCF-7 or T47D cells were co-cultured with the respective mammosphere-enriched BCSCs or parental cells and treated with 5 μM tamoxifen or vehicle. Cell viability was determined by cell count assay. (H-I) Colony formation assay H or Apoptosis (Annexin V-FITC + /PI-) I of MCF-7 or T47D cells cultured with CMs derived from respective mammosphere-enriched BCSCs or parental cells and treated with 5 μM tamoxifen or vehicle. J The schematic of co-implantation model. 2 × 10⁶ luciferase-labeled MCF-7 cells were co-implanted with 4 × 10⁵ unlabeled parental cells or mammosphere-enriched BCSCs into the second fat pad of host mice. A slow-release tamoxifen pellet was implanted or repertaxin was injected to the treatment group as indicated. K In vivo bioluminescent imaging was performed of the tumor burden generated by MCF-7-luc cells. 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 (Two-way ANOVA test in F, others one-way ANOVA followed by Tukey’s multiple comparison)