Dual Sensitization Enables Synergistic Photodynamic Therapy and Radiotherapy for Breast Cancer

Radiotherapy (RT) and photodynamic therapy (PDT) for breast cancer are limited by tumor hypoxia and suboptimal photosensitizer performance. We developed folate-modified copper-doped carbon dots and loaded them with 5-aminolevulinic acid (ALA) to yield FCA, a nanoplatform that executes cascade nanozyme activities to remodel the tumor microenvironment: decomposing H₂O₂ to relieve hypoxia, generating hydroxyl radicals and singlet oxygen (¹O₂), and depleting glutathione (GSH). This priming enabled efficient ALA-to-protoporphyrin IX conversion, which subsequently amplified reactive oxygen species generation. The elevated oxidative stress then synergized with RT to accumulate DNA double-strand breaks and trigger cell cycle arrest. Consequently, FCA-PDT-RT reduced 4T1 cell viability to 20.09% and induced 83.82% apoptosis outcomes mechanistically linked to nuclear factor erythroid 2-related factor 2 (NRF2)-Kelch-like ECH-associated protein 1 (KEAP1)-heme oxygenase 1 (HMOX1) pathway activation. Despite compensatory upregulation of antioxidant genes (HMOX1 and glutamate-cysteine ligase modifier subunit [GCLM]), intracellular GSH and adenosine triphosphate were severely depleted, establishing a metabolic crisis wherein synthesis could not match consumption. This redox/energy collapse drove the pronounced cytotoxicity observed. In an orthotopic 4T1 model, FCA-PDT-RT achieved superior tumor control at only 12 Gy, which correlated with increased CD3⁺/CD8⁺ T cell infiltration and suppressed angiogenesis, while maintaining favorable safety. FCA thus enables synergistic PDT-RT through sequential microenvironment remodeling, oxidative amplification, and metabolic exhaustion, offering a dose-sparing strategy with translational promise for breast cancer therapy.