(761) Interrogating immune niches in a murine GBM model under radiation therapy

Introduction/Rationale: Adult glioblastoma (GBM) is the most aggressive primary brain tumor in adults, with a median survival of approximately 14 months. Standard of care, surgical resection followed by radiation therapy (RT) and temozolomide, has remained largely unchanged since 2005. A major barrier to therapeutic progress is limited understanding of how distinct immune populations within the tumor microenvironment (TME), particularly microglia and peripherally derived macrophages, respond to therapy.

Methods: We developed an immunocompetent, autochthonous adult GBM mouse model driven by EGFRvIII expression with loss of PTEN and CDKN2A. Tumor-bearing mice received a single 8 Gy dose of whole-brain radiation. Immune profiling was performed seven days post-treatment using immunohistochemistry, single-cell RNA sequencing, and spatial transcriptomics to assess myeloid composition and treatment-induced changes within the TME.

Results: RT-treated tumors exhibited reduced tumor burden compared to untreated controls, along with increased accumulation of IBA1⁺ myeloid cells. Single-cell RNA sequencing identified brain-resident microglia and peripherally derived macrophages as the dominant immune populations in both treatment groups. RT induced a relative enrichment of CD49d⁺ peripheral macrophages. Transcriptomic analyses revealed ontogeny-specific responses to RT, with peripheral macrophages enriched for inflammatory, angiogenic, and lipid-associated gene programs consistent with a disease-associated phenotype.

Conclusion: Ongoing spatial transcriptomic analysis using 10x Genomics Visium HD will spatially resolve microglia and peripheral macrophages within the TME and adjacent non-tumor brain regions. Together, this integrated approach defines myeloid heterogeneity in GBM and provides insight into how distinct immune populations respond to radiation therapy.