Abstract

Background: Glioblastoma represents the most common primary malignant brain tumor with dismal prognosis despite multimodal therapies. The immunosuppressive tumor microenvironment and cancer stem cell populations contribute to therapeutic resistance and recurrence. Nitric oxide (NO) metabolism has emerged as a critical regulator of tumor progression, yet its role in coordinating glioma stemness and immune evasion remains poorly understood.
Methods: We performed integrative multi-omics analysis using CGGA, TCGA, and Rembrandt datasets (n=1,500) to investigate NO metabolism patterns across glioma grades and molecular subtypes. Single-cell RNA sequencing data (SCP503, n=65,655 cells) characterized cellular heterogeneity and stemness features. PRMT1 was identified through Cox regression and LASSO modeling as a key NO metabolism-associated prognostic gene. Functional validation used glioma cell lines and patient-derived glioma stem cells through knockdown/overexpression experiments, proliferation assays, invasion studies, and limiting dilution analysis. An orthotopic mouse model evaluated PRMT1's role in tumor growth and stemness maintenance.
Results: NO metabolism activity progressively increased with glioma grade and correlated with poor survival (p<0.001). Single-cell analysis revealed significant positive correlation between NO metabolism and stemness scores (R=0.35, p<0.001). PRMT1 expression was significantly elevated in high-grade gliomas and positively correlated with NOS2 across multiple cohorts (R=0.26-0.46, p<0.001). Spatial transcriptomic analysis showed distinct expression patterns of PRMT1 and NOS2 in different glioma anatomical regions. Functional studies demonstrated that PRMT1 knockdown significantly reduced cell proliferation (76% inhibition), colony formation (75% reduction), and stemness markers (CD133+ cells decreased from 11.74% to 3.11%). NO donor treatment rescued PRMT1 knockdown effects, confirming the NO-dependent mechanism. In vivo experiments showed PRMT1 silencing reduced tumor luminescence to 35% of controls (p<0.001) and decreased expression of stemness markers (SOX2) and immune checkpoint molecules (PD-L1).
Conclusion: PRMT1 functions as a central hub linking NO metabolism to glioma stemness maintenance and immunosuppression. PRMT1 regulates key stemness transcription factors (OCT4, SOX2) and immune checkpoint molecules (PD-L1) through NO-dependent mechanisms. These findings identify PRMT1 as a potential therapeutic target that could simultaneously disrupt tumor stem cell populations and remodel the immunosuppressive microenvironment, providing a novel strategy for precision glioma therapy.

Keywords

PRMT1, glioma, nitric oxide metabolism, cancer stem cells, immunosuppression, tumor microenvironment