A Sequence-Dependent Combination of Photodynamic Therapy and Carboxyamidotriazole Orotate for Enhanced Treatment of Glioblastoma

Glioblastoma (GBM) remains a highly lethal malignancy characterized by profound treatment resistance and metabolic plasticity. While photodynamic therapy (PDT) and the mitochondrial complex I inhibitor carboxyamidotriazole orotate (CTO) have individually shown promise, their combined potential requires further exploration and optimization. This study systematically investigated the interaction between 5-aminolevulinic acid (5-ALA)-mediated PDT and CTO in U87 GBM models. Intriguingly, we discovered a sequence-dependent interaction under the tested treatment schedules: CTO pre-incubation before PDT resulted in attenuated PDT-induced cytotoxicity, possibly due to CTO-mediated suppression of reactive oxygen species (ROS) accumulation. In contrast, a sequential “PDT→CTO” regimen enhanced anti-tumor efficacy both in vitro and in vivo. Mechanistically, the sequential approach was associated with enhanced mitochondrial depolarization and reduced expression of glycolysis-related genes, suggesting a potential metabolic “dual-hit” involving disturbance of mitochondrial function and compensatory glycolytic adaptation. These results highlight treatment sequence as a critical determinant of PDT-CTO interaction and provide a basis for further preclinical investigation of PDT followed by metabolic intervention as a combination strategy with potential translational relevance.