Tetrahydrobiopterin inhibitor-based antioxidant metabolic strategy for enhanced cancer ferroptosis-immunotherapy

The induction of immunogenic ferroptosis in cancer cell is limited by the complex and delicate antioxidant system in the organism. Synergistic induction of oxidative damage and inhibition of the defensive redox system in tumor cells is critical to promote lethal accumulation of lipid peroxides and activate immunogenic death (ICD). To address this challenge, we present a multifunctional and dual-responsive layered double hydroxide (LDH) nanosheet to enhance immunogenic ferroptosis. The MTX-LDH@MnO₂ nanoplatform is constructed by intercalating methotrexate (MTX) into LDH interlayers and electrostatically absorbing biomineralized ovalbumin (OVA)-MnO₂ onto the LDH surface. Specifically, the released Mn²⁺ from the incorporated MnO₂ triggers a Fenton-like reaction, leading to reactive oxygen species (ROS) accumulation, while the depletion of reduced glutathione (GSH) further intensifies oxidative stress, resulting in the induction of ferroptosis. MTX is released in response to the acidic environment of tumor cells and inhibits the regeneration of tetrahydrobiopterin (BH4), modulating the GTP cyclic hydrolase 1 (GCH1)/BH4 axis. MTX disrupts the antioxidant metabolic activity regulated by GCH1/BH4 axis and inhibits ROS consumption, further boosting the ferroptosis effect, which promoted the release of damage-associated molecular patterns (DAMPs) and triggered ICD in the tumor. This activation subsequently leads to significant antitumor immune reactions, including DCs maturation, infiltration of CD4⁺/CD8⁺ T cells and cytokines release. The redox-controllable nanoplatform demonstrates promising anticancer efficacy in a mouse breast model providing a novel strategy for cancer immunotherapy.