IDH-wildtype glioblastoma exhibits substantial biological heterogeneity despite current molecular classification. Although dynamic malignant cell states have been described, molecular features that position tumors along these states remain incompletely defined. We investigated whether ZIC3 expression identifies a transcriptional state associated with clinical outcome in IDH-wildtype glioblastoma.

RNA sequencing and clinical data from The Cancer Genome Atlas glioblastoma cohort were analyzed after restriction to genomically confirmed IDH-wildtype tumors (n = 167). Survival analyses were performed in patients with available survival data (n = 161) using Cox proportional hazards regression and Kaplan-Meier analysis. Associations with MGMT promoter methylation, tumor microenvironment scores, differential expression, gene set enrichment, and single-cell RNA sequencing data were evaluated to characterize transcriptional programs associated with ZIC3 expression.

Higher ZIC3 expression was associated with longer overall and progression-free survival and remained associated with a more favorable survival profile after adjustment for age, sex, radiotherapy, and temozolomide treatment. ZIC3 expression was not associated with MGMT promoter methylation or with ESTIMATE-derived stromal, immune, or composite scores. In the single-cell dataset, ZIC3-positive cells were more frequent in malignant tumor cells than in non-malignant cells. ZIC3-high tumors showed enrichment of neural lineage-associated and blood-brain barrier-related programs, whereas ZIC3-low tumors showed enrichment of hypoxia-related, glycolytic, and metabolic stress programs. Ordered heatmap analysis showed that lineage-associated genes and hypoxia-related genes varied inversely across tumors ordered by ZIC3 expression.

ZIC3 expression appears to mark a developmental transcriptional axis in IDH-wildtype glioblastoma that links neural lineage-associated programs to hypoxia-adaptive tumor states.