Fig. 3

Expression analysis and downstream factors in xenograft tumor samples. A–L DNMT3A staining of OCI-AML2 and KG1 tumor sections. DNMT3A was detected in nuclei of OCI-AML2 tumor sections treated with PBS and αCD33-mAB-P/P-control-siRNA; scr, scrambled (= control) (A–D), but DNMT3A staining was almost completely lost after treatment with αCD33-mAB-P/P-DNMT3A-siRNA (E–F). Reduced DNMT3A staining was also observed in KG1 tumor sections from the αCD33-mAB-P/P-DNMT3A-siRNA treatment group (K–L compared to G–J). Nuclear counterstain was performed using Hoechst33342 (B, D, F, H, J, L). M–Q Relative expression (RT-PCR) of DNMT3A and downstream genes in OCI-AML2 tumors ex vivo upon previous in vivo exposure to PBS, control carriers (αCD33-mAB-P/P-scr-siRNA) and αCD33-mAB-P/P-DNMT3A-siRNA. D3A, DNMT3A siRNA; α, anti. R,S Gene set enrichment analysis (GSEA) of RNA sequencing data in αCD33-mAB-P/P-DNMT3A-siRNA vs. scr-siRNA carrier-treated OCI-AML2 tumors, ex vivo. GSEA analysis of hallmark gene sets from the molecular signature database identified significant enrichment for hallmark_oxidative phosphorylation (R) and hallmark_c-Myc targets (T) in αCD33-mAB-P/P-scr-siRNA carrier-treated OCI-AML2 tumors. Shown are representative gene set enrichment plots. FDR, false discovery rate q-value and NES, normalized enrichment score. S, U Graphical presentation of central genes within the biological networks identified in the GSEA analysis. Top 15 hub genes identified by String/cytoscape/cytoHubba interface were presented as a circular layout (color coded: red and yellow indicating a higher and lower rank, respectively) using degree based algorithm of cytoHubba plug-in. Rank, gene and the scores were presented in the adjacent table. Furthermore, protein–protein interaction network/subnetworks were analyzed using 12 different algorithms of cytoHubba plug-in and hub genes representing intersection of at least nine algorithms (75% overlapping signature) potentially indicate key DNMT3A related hub genes in inducing c-Myc targets and oxidative phosphorylation. Detailed information on these hub genes and their importance in different cancers and therapy outcome is depicted in Additional file 1: Figure S4