Enhancer DNA methylation distinguishes CN-AML from granulocytes and progenitor cells. (A) Volcano plots showing DNA methylation in CN-AML relative to normal bone marrow (NBM). Of 13 448 DMCs, 6607 (49.1%) display hypermethylation. (B) DNA methylation profile of CN-AML relative to NBM at CAGE enhancers, 446 (70%) of 632 DMCs are hypermethylated. Dots represent methylation levels of individual probes in the Illumina 450k array. Y-axis indicates the odds ratio of –log₁₀ false discovery rate (FDR); X-axis shows the fold change of the methylation value (M-value). Statistical significance cutoffs are marked with red dashed lines (FDR <0.05; FC >1.5). (C) Principal component analysis (PCA) of CN-AML, normal progenitor cells, and mature cells at CAGE enhancers. (D) PCA of TSS and promoter-associated probes. Enhancer probes (8202) are annotated to CAGE enhancers defined by the FANTOM Consortium, whereas probes located within 2000 bp upstream of TSS regions are included as promoter probes (117 789). Mature myeloid cells (PMCs, PMN cells) are shown as red dots, progenitors (CMPs, GMPs) are shown as blue dots, and CN-AML patients are shown as green dots. (E) Heat map and hierarchical clustering of enhancer DMCs of CN-AML, progenitors, and mature myeloid cells. Enhancer DMCs have been defined by pairwise comparison and clustered for all tested samples with supervised categories. M-value of each of the DMC probes is color-coded in the heat map from low (blue) to high (red). PC1, principal component 1; PC2, principal component 2.