Abstract
<title>Abstract</title> <p>Purpose Diffuse gliomas remain lethal because malignant-state plasticity, immune suppression, and treatment resistance are interlocked within the tumor ecosystem. The Mediator complex links enhancer-bound transcription factors to RNA polymerase II, yet the relevance of the Mediator head-module subunit MED8 in glioma remains unclear. This study tested whether MED8 marks and functionally supports a mesenchymal-immune glioma state. Methods We integrated CGGA and TCGA transcriptomic cohorts, clinical and molecular stratification, survival modeling, pathway and network analysis, immune deconvolution, public single-cell and spatial transcriptomic context, DepMap/CRISPR dependency screening, pharmacogenomic prioritization, and replicate-level experimental validation. MED8 perturbation was assessed by shRNA knockdown, qRT-PCR, western blot densitometry, Transwell invasion, and mesenchymal/immune-regulatory marker assays. Results MED8 expression increased with WHO grade and was higher in IDH-wildtype tumors. In the processed CGGA survival summary, MED8-high tumors had shorter median overall survival than MED8-low tumors (18.5 vs 52.3 months; HR = 2.81, 95%CI 2.18–3.62; P = 2.5x10^-18), consistent with the external TCGA summary (HR = 4.34, 95%CI 3.12–6.05). MED8-high tumors were enriched for EMT, TNF/NF-kB, inflammatory response, MYC/E2F, IL6/JAK/STAT3, and PI3K-AKT-mTOR programs. Immune deconvolution, TIDE, CRISPR dependency, and pharmacogenomic prioritization nominated macrophage-associated immune suppression and BET/mTOR pathway vulnerability as testable hypotheses. Experimentally, MED8 knockdown reduced MED8 mRNA (1.00+/-0.02 to 0.28+/-0.01; P = 5.82x10^-5), MED8 protein density (1.00 to 0.28), Transwell invasion (245.0+/-6.7 to 68.0+/-3.2 cells/field; P = 0.0002), mesenchymal markers, NF-kB/STAT3-associated protein signals, and immune-regulatory factors. Conclusions MED8 marks an aggressive mesenchymal-immune glioma state and has functional evidence supporting a role in invasion and mesenchymal/immune-regulatory marker programs. The data nominate MED8 as a mechanistic and translational candidate, while direct chromatin, rescue, macrophage co-culture, clinical protein-validation, and MED8-stratified drug-response experiments remain necessary before clinical translation.</p>