Radiobiology and Radioresistance in High-Dose Radiosurgery for Brain Tumors: A Hypothesis-Generating Study Using an Intracranial Glioma Mouse Model.

Abstract

Introduction: Stereotactic radiosurgery (SRS) is a precise, non-invasive treatment for brain tumors, yet underlying radiobiological mechanisms remain unclear. This study explored long-term dose-dependent tumor response to high-dose SRS in a murine glioma model, focusing on tumor-associated macrophages (TAMs) as key regulators of tumor microenvironment and immune modulation.

Methods: Using the intracranial GL261-glioma mouse model, single-dose SRS was administered at either 20-Gy (clinically prescribed dose) or 40-Gy as a dose-escalation approach (n=24/dose). Tumor response was assessed longitudinally using 7T MRI at predefined intervals (d7, d30, d90, and d180) post-SRS or earlier upon symptom onset. Histological analyses performed at each timepoint evaluated cell proliferation, apoptosis, vascular morphology, blood-brain/tumor-barrier integrity, hypoxia, TAM recruitment, and polarization. Immune cell populations within tumor microenvironment were characterized using flow cytometry. Statistical analyses included a T-test and one-way ANOVA with Bonferroni or Dunnett correction.

Results: SRS efficacy was dose-dependent: 40-Gy suppressed tumor growth, while 20-Gy led to regrowth in 29% of cases between d30-51 post-SRS, necessitating stratification into responders and non-responders. Responders demonstrated reduced cell proliferation, sustained apoptosis, and vascular remodeling indicative of vessel normalization. Non-responders exhibited up to 94.5% increased hypoxia and up to 300-fold increased CXCR4 expression compared to responders. TAM recruitment inversely correlated with tumor volume (r=-0.8619, p=0.0056). The M1/M2 ratio in non-responders was similar to that of matched controls, but 3.8-fold and 4.8-fold lower than in responders at d30 and d90 post-SRS, respectively. FACS analysis confirmed an increased M1/M2 ratio by d30 in responders.

Discussion: This study offers key insights into longitudinal SRS radiobiology, highlighting the dynamic role of TAMs in sustaining long-term tumor control. Our findings support an association between hypoxia, CXCL12/CXCR4 signaling, and treatment resistance, and suggest a potential SRS-induced vascular normalization that may support improved therapeutic outcomes. While not designed to establish causality, the data provide a spatially and temporally resolved framework to guide future mechanistic studies and inform more effective SRS-based combination strategies.