MitoGPS: A Mitochondrial DNA G-Quadruplex-Targeting Strategy for Tumor-Preferential Type I Photodynamic Therapy.

Photodynamic therapy (PDT) is a clinically promising, minimally invasive cancer treatment offering precise spatiotemporal control. However, achieving preferential tumor accumulation remains a critical challenge, which limits therapeutic efficacy and induces off-target toxicity. Here, we introduce the mitoGPS (mitochondrial G-quadruplex-targeting photosensitizer) strategy, a rational design framework for developing photosensitizers (PSs) with prolonged intratumoral retention through targeting mitochondrial DNA G-quadruplexes (mtDNA G4s). This approach leverages the unique structural features and biological attributes of mtDNA G4s, which are increasingly implicated in cancer cell metabolism and survival, to achieve preferential photosensitizer accumulation and action within tumor mitochondria. By integrating molecular generation, G4-specific docking simulations, and evaluation of photosensitizing properties, we established a modular workflow for the discovery of mitoGPS. Using this pipeline, we designed and identified A1, a lead mitoGPS compound. A1 exhibits preferential mitochondrial localization in tumor cells, with its retention influenced by interactions with mtDNA G4s, and generates predominantly Type I reactive oxygen species (ROS) upon light activation. The resulting mitochondrial oxidative stress disrupts mitochondrial integrity and engages apoptosis-associated and ferroptosis-associated responses. Our work supports mtDNA G4-associated recognition as a strategy for improving the subcellular precision of PDT and provides a generalizable framework for developing mitochondria-directed phototherapies.