Increased DNA methylation at relapse. (A) Plot of the first two principal components (PC) of the genome-wide DNA methylation data in 27 diagnostic-first/second relapse paired samples. The diagnostic sample is indicated by a filled yellow circle and the last relapse sample is indicated by an arrowhead. (B) PCA of the 10 diagnostic-first relapse paired samples in the Quebec childhood ALL (QcALL) dataset. (C) Enrichment of DMCs in relation to gene region, CpG island annotation, and chromatin marks. Hypermethylated DMCs are shown in red and hypomethylated DMCs are shown in blue. In each box the fold enrichment for each specific mark is shown. Bold numbers indicate annotations enriched for relapse DMCs compared to the distribution of probes on the 450k array (Bonferroni corrected one sided Fisher’s exact P < 0.001). (D) Distribution of the methylation β-values in the relapse signature at remission (n = 3), diagnosis (n = 27), first relapse (n = 27), and second relapse (n = 5). (E) The mean β-values of the DMCs in the relapse signature in the Nordic Society of Pediatric Hematology and Oncology (NOPHO) and QcALL datasets. The color legend for (A,B,D,E) is to the right of (E). (F) Heatmap of the top ranked relapse DMCs. From the 27 patients in the NOPHO dataset, paired remission BM was available from 3, diagnostic samples were available from 27, first relapse data were available from 27, and second relapse data were available from 5 individuals. From the QcALL dataset, DNA samples from remission BM, diagnosis and relapse were available from each of the 10 patients. Each DMC is plotted as an individual row. Each column represents one DNA sample. The proportion of methylated CpG sites is shown in the bottom panel. The gene annotations of the CpG sites are given on the right vertical axis. The color key for the methylation levels is provided at the bottom right.