Fig. 5

HDAC inhibitors increase susceptibility of EwS to first line chemotherapeutics. a Heatmaps showing excess over Bliss synergy scores for combination treatment of Doxorubicin with HDACi FK228 (left) or MS-275 (right) in CHLA-10 or SK-N-MC cells. Scores greater than 1 (shown in red) denote synergistic combinations, whereas scores less than 1 (shown in green) denote antagonistic combinations. Data represent 3 technical triplicates. b Western blot analysis of caspase 3 and PARP as a measure of apoptosis susceptibility after treatment with Doxorubicin and HDACi, either in combination or individually. CHLA-10 were treated for 36 h and SK-N-MC for 24 h. Tubulin was used as loading control. c Focused heat map of genes, ≥1.5-fold differentially expressed in EwS line EW7 after combined treatment with DMSO, HDACi and/or first line chemotherapeutics Doxorubicin or Vincristine for 17 h. Treatment with Doxorubicin or Vincristine and MS-275, respectively, are highlighted in red. Each column represents one individual array. Microarray data with their normalized fluorescent signal intensities were used (robust multichip average (RMA); GSE162785). d GSEA enrichment plots of up- and downregulated gene sets after combined treatment with MS-275 and Doxorubicin. NES: normalized enrichment score. GSEA: http://www.broadinstitute.org/gsea/index.jsp. e GSEA leading-edge analysis of identified gene sets (C5_all, GO gene sets) downregulated after combined treatment with MS-275 and Doxorubicin. Set-to-set analysis shows a correlation between combined MS-275 and Doxorubicin treatment and downregulation of gene sets important for chromosomal regulation