Minibeam radiation therapy remodels tumor microenvironment and suppresses HIF-1α/VEGFR axis to overcome radioresistance in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is resistant to radiotherapy due to tumor hypoxia and abnormal angiogenesis, necessitating strategies to enhance therapeutic outcomes. This study evaluates the use of minibeam radiation therapy (MBRT), delivered through a novel 3D-printed collimator made of polylactic acid (PLA) and tungsten, to modulate the TNBC microenvironment and potentially overcome radioresistance. Three collimator configurations (400, 600, 800 μm beam widths) were tested. Mice received MBRT (150 Gy) or conventional radiotherapy (CRT, 7 or 15 Gy), with tumor responses assessed using histology, RNA sequencing, and immunohistochemistry. The measured beam FWHM values for the MBRT 0.4, 0.6, and 0.8 groups were 419 ± 23 μm, 575 ± 31 μm, and 798 ± 50 μm, respectively, while the CTC distances were 832 ± 25 μm, 1296 ± 21 μm, and 1651 ± 49 μm. MBRT generated stable, spatially fractionated dose distributions with high peak-to-valley ratios. Compared to CRT at equivalent valley doses, MBRT significantly reduced tumor growth, proliferation, and hypoxia while increasing necrosis. Mechanistically, MBRT downregulated HIF-1α/VEGFR signaling, alleviating hypoxia and angiogenesis, and enhanced vascular normalization via increased pericyte coverage. These findings suggest MBRT reprograms the TNBC microenvironment, supporting its potential as a radiosensitizing strategy for clinical translation.