Biophysical enhancement of radiation-activated photodynamic therapy (radioPDT) and translational brain tumor models

Background: Radiation-activated photodynamic therapy (radioPDT) uses X-ray-excited nanoscintillators to activate photosensitizers deep within tissues, generating cytotoxic reactive oxygen species without requiring receptor expression. This biophysical strategy is well suited for heterogeneous tumors such as glioblastoma (GBM), where receptor-targeted approaches often fail. A major translational barrier is the blood-brain barrier (BBB), which restricts nanoparticle entry. Physical modulation via focused ultrasound (FUS) and microbubble cavitation can enhance nanoparticle delivery and potentially improve radioPDT efficacy. To evaluate this approach, we established a multi-model pipeline incorporating flank xenografts, intracranial GBM models, and the chick CAM system, allowing visualization of nanoparticle transport, vascular effects, and treatment response. Methods: SCID mice bearing PC3 flank tumors received control, radiation, FUS, NP+RAD, NP+FUS+RAD, or NP+microbubbles+FUS+RAD. Tumors were analyzed using multiplex immunofluorescence for proliferation/apoptosis (Ki67, cleaved Caspase-3), DNA damage (gamma-H2AX, 53BP1), oxidative injury (4-HNE, TUNEL), vascular structure (CD31, NG2), hypoxia (CA9, HIF-1alpha), and inflammation (CD45, Iba1). For translational studies, U87 and U251 GBM cells expressing LUC-GFP were validated and used to generate intracranial xenografts for bioluminescence imaging and a CAM model enabling rapid assessment of nanoparticle behavior and radioPDT effects. Results: radioPDT alone disrupted endothelial cells, while radioPDT combined with microbubble-enhanced FUS produced both endothelial and pericyte disruption, suggesting potential for BBB modulation. Multiplex tumor analysis is ongoing. LUC-GFP GBM models have been established, and intracranial and CAM tumors provide complementary systems to study FUS-mediated BBB opening and nanoparticle delivery during radioPDT. Conclusions: radioPDT may amplify ultrasound-mediated vascular modulation without requiring receptor-specific targeting. The integrated PC3, intracranial GBM, and CAM platforms establish a translational pathway to evaluate BBB-penetrant, physics-based radioPDT. These studies are designed to define how physical forces-rather than receptor-mediated uptake-shape nanoparticle distribution, vascular response, DNA damage, oxidative injury, and overall radioPDT potency.