Figure 2

The Genome-wide DNA methylation profiles of primary OS. (A) Unsupervised hierarchical clustering analysis using normally unmethylated CGIs (47,759 genomic blocks) of primary OS (n = 28), Ewing sarcoma (EWS) (n = 11), normal whole blood (n = 3), muscle (n = 3), fat (n = 2), and bone (n = 3) samples. Ten OS and one ES were classified into a cluster with larger numbers of methylated CGIs. The remaining 18 OS and 10 EWS clustered together with normal samples with limited numbers of methylated CGIs. Gastric cancer (n = 8) and colorectal cancer (n = 5) (41,452 genomic blocks) were similarly clustered for comparison. (B) Fraction of methylated CGIs in OS with a relatively large number of methylated CGIs, CIMP(+) gastric cancer, and colon cancer. The fraction was much smaller in the OS than in the CIMP(+) gastric cancer. (C) Unsupervised hierarchical clustering analysis of primary OS samples using the 500 most variable genomic blocks. Primary OS samples were classified mainly into two clusters, and one cluster (cluster IV-a) had a larger number of methylated genomic blocks. (D) Unsupervised hierarchical clustering analysis of primary and metastatic OS samples. The 500 most variable genomic blocks used in (C) were utilized, and four of six metastatic OS samples were clustered with primary OS samples in cluster IV-a (clusters IV-b and V). (E) Distribution of somatic mutations in the OS samples in clusters IV-a and cluster III-a. There was no association between the methylation clusters and mutation frequency of a specific gene. Tumors in red are the pairs of primary and metastatic tumors from same patients (the pairs are OS_8 and OS_8M, OS_16 and OS_16M, OS_21 and OS_21M). (F) Kaplan-Meier curves of progression-free survival (PFS) for 28 primary (M0) OS patients according to their methylation clusters. There was no significant difference (P = 0.868).