Promoter Methylation–Expression Coupling of Gliogenesis Genes in IDH-Wildtype Glioblastoma: Longitudinal Analysis and Prognostic Value

Glioblastoma (GBM) shows extensive epigenetic heterogeneity. In <i>IDH</i>-wildtype (<i>IDH</i>-WT) GBM, promoter DNA methylation may regulate lineage programs influencing tumor evolution and prognosis; here, we systematically profiled promoter-level methylation dynamics across longitudinal tumors. Genome-wide DNA methylation data were obtained from the publicly available Gene Expression Omnibus (GEO; GSE279073) dataset, comprising a longitudinal cohort of 226 <i>IDH</i>-wildtype glioblastomas profiled on the Illumina Infinium EPIC 850K array across primary and recurrent stages at the University of California, San Francisco. From 333 Gene Ontology gliogenesis-annotated genes (GO:0042063), a 48-gene promoter panel was derived, with ≥2 probes per gene. Promoter methylation was summarized as the median β-value and tested using one-sample Wilcoxon with FDR correction. Functional enrichment, longitudinal variation, and patient-level methylation burden were assessed. Validation analyses were performed using independent <i>IDH</i>-wildtype GBM datasets from The Cancer Genome Atlas (RNA-seq and 450K methylation; <i>n</i> = 347). Promoter hypomethylation predominated across all stages, with 25 genes consistently hypomethylated and 7 hypermethylated. Functional enrichment highlighted gliogenesis, glial cell differentiation, neurogenesis, and Notch-related signaling. In TCGA, promoter methylation inversely correlated with expression for 11 of 33 genes (FDR < 0.05). An Expression Score contrasting hypomethylated and hypermethylated genes was positively associated with improved overall survival, where higher scores predicted better outcome (HR = 0.87, <i>p</i> = 0.016; Q4 vs. Q1 HR = 0.68, <i>p</i> = 0.025), and a complementary Methylation Score showed that higher promoter hypermethylation predicted poorer outcome (HR = 1.73, <i>p</i> < 0.001). <i>CNTN2</i> and <i>TSPAN2</i> were adverse prognostic genes (FDR < 0.05). The Expression Score was highest in Proneural tumors and lowest in Mesenchymal tumors (<i>p</i> < 0.001), reflecting a proneural-like state associated with better prognosis. Promoter methylation within gliogenesis genes defines a stable yet prognostically informative epigenetic signature in <i>IDH</i>-WT GBM. Hypomethylation promotes transcriptional activation and a favorable outcome, whereas hypermethylation represses lineage programs and predicts poorer survival.