Introduction
Glioma is the most common primary intracranial tumor, accounting for ~22.2% of all central nervous system malignancies (1). The World Health Organization (WHO) classifies gliomas into grades I–IV, with grades I–II considered low-grade gliomas (LGG) and grades III–IV considered high-grade gliomas (HGG) (2). Histopathological features are closely correlated with the degree of malignancy (3,4). HGGs are associated with high morbidity, therapeutic resistance, frequent recurrence and poor survival, thereby imposing a substantial burden on patients and society (5–7). Surgical resection remains the primary treatment; however, its efficacy is limited and adjuvant radiotherapy and chemotherapy are often required (8–10). Despite multimodal therapy, recurrence rates remain high (11). The median survival of patients with LGG is ~10 years, whereas that of untreated patients with HGG is <20 months (12), with only a minority surviving beyond 3 years after surgery (13). Therefore, understanding the molecular mechanisms underlying glioma progression is essential for identifying novel diagnostic biomarkers and therapeutic targets.
Primarily localized in the nucleus, the histone methyltransferase EZH2 is integral to the assembly of the Polycomb repressive complex (14,15). This enzyme trimethylates lysine 27 of histone H3 (H3K27me3), leading to gene silencing through both canonical and non-canonical mechanisms, thereby exerting transcriptional repression or activation (16–18). EZH2 plays a crucial role in cell proliferation, differentiation and apoptosis and its functions are closely associated with the regulation of diverse targets and signaling pathways (19,20).
EZH2 has recently emerged as a significant regulator in various cancers (21), influencing cell proliferation, apoptosis, epithelial-mesenchymal transition, invasion and drug resistance in glioma and other malignancies (22–24). Suppression of EZH2 through small-molecule inhibitors or gene knockdown results in reduced growth and tumor formation in rectal cancer cells (25). Moreover, within the tumor microenvironment, EZH2 expression directly influences T-cell responses and is involved in the suppression of chemokine signaling and cytotoxic lymphocyte activity (26). In cancer, elevated EZH2 expression is associated with tumor aggressiveness, unfavorable prognosis and recurrence across multiple tumor types, thereby serving as a diagnostic marker (27–29). The present study investigated the prognostic relevance of EZH2 expression in glioma by integrating public database analyses [TIMER2.0, The Cancer Genome Atlas (TCGA)] with a large Chinese cohort of 147 patients, with detailed stratification by WHO grade, IDH status and 1p/19q co-deletion.
Discussion
Gliomas represent the most prevalent primary tumors of the central nervous system (1,30,31), with glioblastoma accounting for 60–75% of cases (32). Despite the establishment of combined surgical resection, localized fractionated radiotherapy, and adjuvant temozolomide as the standard first-line therapy, the 5-year survival rate for patients with glioblastoma remains strikingly low at 6.8% (33,34). This highlights the critical need to identify novel and reliable prognostic biomarkers to improve survival prediction and optimize therapeutic strategies for patients with glioma.
EZH2, a histone methyltransferase, is frequently overexpressed in multiple cancers and is consistently associated with poor prognosis (35–38). Mechanistically, EZH2 exerts its oncogenic effects through both canonical and non-canonical pathways. Canonically, it catalyzes H3K27 trimethylation (H3K27me3), leading to transcriptional silencing of tumor suppressor genes such as CDKN2A, CDKN1C and RUNX3 (39). Non-canonically, EZH2 can regulate gene expression or methylate non-histone proteins independently of its methyltransferase activity (40). Moreover, crosstalk between EZH2 and several signaling pathways, including PI3K/AKT and Notch, has been implicated in pancreatic neuroendocrine neoplasm cell proliferation, maintenance of stemness, and therapy resistance (41). Previous studies have shown that EZH2 inhibition impairs glioma cell self-renewal in vitro, reduces tumorigenicity in vivo, and enhances chemo-radio-sensitivity, ultimately improving survival (42–44). Despite these findings, the association between EZH2 expression and key clinicopathological features, including glioma grade, IDH mutation status, 1p/19q co-deletion, and prognosis, had not been comprehensively characterized prior to the present study.
In the present study, EZH2 expression in glioma was systematically evaluated using both public databases and the present clinical cohort. Analysis of TIMER2.0 and TCGA data demonstrated that EZH2 mRNA expression was significantly upregulated in glioma tissues compared with normal brain tissue and was highly expressed across multiple cancer types. In the TCGA cohort, high EZH2 mRNA expression was strongly associated with higher WHO grade, aggressive histological subtypes, IDH wild-type status, and the absence of 1p/19q co-deletion. Kaplan-Meier analysis showed that elevated EZH2 mRNA expression was associated with shorter OS, DSS, and PFI. To validate these findings at the protein level, immunohistochemistry was performed on 147 glioma specimens spanning WHO grades I–IV. The results demonstrated that higher WHO grade was associated with increased EZH2 protein expression. Conversely, lower EZH2 expression was associated with more favorable pathological features, lower histological grade, reduced recurrence, and longer survival. Univariate Cox regression analysis identified EZH2 expression level, patient age, sex, and WHO grade as significant prognostic factors. Subsequent multivariate analysis confirmed that EZH2 expression, age, and WHO grade were independent predictors of OS in patients with glioma. Collectively, these findings indicate a strong association between elevated EZH2 expression and higher tumor grade, with increased expression levels predicting poorer prognosis.
Beyond its prognostic value, EZH2 is emerging as a promising therapeutic target. The EZH2 inhibitor tazemetostat has received FDA approval for epithelioid sarcoma and follicular lymphoma, and preclinical studies have evaluated its efficacy in glioma models (45,46). In glioblastoma cell lines and xenograft models, EZH2 inhibition suppresses proliferation, induces differentiation, and sensitizes tumors to temozolomide and radiotherapy (47,48). However, the clinical translation of EZH2 inhibitors for glioma faces considerable challenges, including the blood-brain barrier and intratumoral heterogeneity. Nevertheless, these findings support further exploration of EZH2 not only as a biomarker for patient stratification but also as a candidate for combination therapy with current standard-of-care regimens.
Limitations and future directions: Several limitations of the present study should be acknowledged. First, the molecular mechanisms by which EZH2 promotes glioma progression were not experimentally investigated. In vitro or in vivo functional assays were not performed (for example, knockdown or overexpression of EZH2) to establish causality or dissect the downstream signaling pathways. Second, multi-factor interaction analyses incorporating other clinically relevant molecular markers, such as MGMT promoter methylation and TERT mutations, were not conducted, as these data were not available for all patients in the present retrospective cohort. Consequently, the authors could not assess whether EZH2 expression adds independent predictive value beyond these established markers or evaluate its role in treatment response prediction. Third, the tissue samples were obtained from a single commercial source (Shanghai Outdo Biotech Company), which may introduce selection bias and limit the generalizability of the findings of the present study to other populations or clinical settings. Finally, the retrospective design inherently carries the risk of unmeasured confounding. Future studies should prospectively validate the findings of the present study in larger, multi-center cohorts with comprehensive molecular profiling to enable robust multi-factor interaction analyses. Mechanistically, gain-of-function and loss-of-function approaches are urgently needed to elucidate the canonical and non-canonical pathways of EZH2 in glioma. Additionally, the therapeutic potential of combining EZH2 inhibitors with temozolomide and radiotherapy warrants further investigation, particularly in the context of overcoming chemo-radio-resistance.
In summary, the present study provided robust evidence that EZH2 expression is closely associated with glioma grade, molecular subtype and patient survival. The consistency between mRNA and protein expression data, its independent prognostic value in multivariate analysis, and its biological plausibility based on established mechanisms collectively support EZH2 as a clinically useful prognostic biomarker in glioma. Further independent validation and mechanistic studies are warranted to facilitate translation of these findings into clinical practice.