Glioblastoma (GBM) is a devastating cancer of the brain with an extremely poor prognosis. While X-ray radiotherapy and chemotherapy remain the current standard, proton beam therapy is an appealing alternative as protons can damage cancer cells while sparing the surrounding healthy tissue. However, the effects of protons on in vitro GBM models at the cellular level, especially when co-cultured with endothelial cells, the building blocks of brain micro-vessels, are still unexplored. In this work, novel 3D-engineered scaffolds inspired by the geometry of brain microvasculature are designed, where GBM cells cluster and proliferate. The architectures are fabricated by two-photon polymerization (2PP), pre-cultured with endothelial cells (HUVECs), and then cultured with a human GBM cell line (U251). The micro-vessel structures enable GBM in vivo-like morphologies, and the results show a higher DNA double-strand breakage in GBM monoculture samples when compared to the U251/HUVECs co-culture, with cells in 2D featuring a larger number of DNA damage foci when compared to cells in 3D. The discrepancy in terms of proton radiation response indicates a difference in the radioresistance of the GBM cells mediated by the presence of HUVECs and the possible induction of stemness features that contribute to radioresistance and improved DNA repair.