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. 2022 Jan 5;2(1):1–9. doi: 10.1158/2767-9764.CRC-21-0088

FIGURE 2.

FIGURE 2

DNA methylation changes in recurrent IDH-wt GBM after treatment with LMF. Note: “Folic” is LMF. A, Kaplan–Meier curve of OS in LMF-treated patients (blue line) versus historical control group treated with bevacizumab alone (red line). The 95% CI is represented by the blue and red shaded areas. See Supplementary Table S1 for patient characteristics and n value. CTR, bevacizumab treatment; folic is LMF-treated patients. B, Volcano plot comparing the average methylation change from initial GBM (at diagnosis) to autopsied GBM (after exposure to LMF at recurrence, measurements were taken from distinct paired samples). Colored dots represent CpG sites that show significant hypermethylation, FDR<0.05 and Δβ > 0.2, (orange dots, total count provided in the parentheses) or hypomethylation, FDR<0.05 and Δβ < −0.2, (purple dots, total count provided in the parentheses) at autopsy. The total CpGs that were FDR<0.05 at any level of Δβ are summarized outside the parentheses. The volcano plot was generated using the 850K-EPIC platform. Folic, LMF treated. C, The genomic distribution of the 645 hypermethylated CpGs [red box plot, initial GBM (at diagnosis); blue box plot, autopsied GBM tumor (after exposure to LMF at recurrence)]. Center line, median; box limits, upper and lower quartiles; whiskers, 1.5× interquartile range; points, outliers. Folic, LMF treated. D, Comparison of the DNA MDI between LMF-treated patients (Folic), TCGA paired samples (TCGA), and our institutional paired controls (CTR). Box plot elements as in C. E, Using gene ontology (GO) analysis, we show pathways impacted by the 645 hypermethylated CpGs identified in B. GO terms were summarized using REVIGO (http://revigo.irb.hr/). For each class, terms remaining after the redundancy reduction are represented in a two-dimensional space, which summarizes GO terms’ semantic similarities. Semantically similar terms are close together in the plot, but the semantic space units have no intrinsic meaning. Bubble color indicates the P value (all terms included had a P < 0.05); circle size indicates the frequency of the GO term in the underlying GO database (more general terms have larger plot size). CpG Island: 300 to 2,000 bp in length close to promoters with high GC content; CpG Shores: <2 kb from CpG Island; CpG Shelves: 2–4 kb from CpG Island; Open Sea: >4 kb from CpG Island.