Abstract

Ferroptosis has emerged as a promising strategy for cancer treatment, which damages tumor cells by triggering the lethal accumulation of reactive oxygen species (ROS) and lipid peroxides. However, the development of efficient ferroptosis-inducing medicines is severely impeded by the unidentified key subcellular sites of lipid peroxidation. Given the critical role of endoplasmic reticulum (ER) during ferroptosis, an ER-targeting prodrug nanoassembly (ISSM) is designed to efficiently evoke ferroptotic cell death. The molecular scaffold of ISSM is constructed by conjugating the cyclooxygenase-2 (COX-2) inhibitor indomethacin with an ER-targeting moiety via a disulfide-containing linker. Following cellular internalization and subsequent ER localization, ISSM depletes cellular glutathione (GSH) through the thiol-disulfide exchange reactions, thereby crippling GSH-glutathione peroxidase 4 defense axis. Meanwhile, the released active drug not only elevates the arachidonic acid (AA) levels by inhibiting COX-2, but also induces intense endoplasmic reticulum stress and ROS production. The synergistic effect of these actions drives a robust increase in lipid peroxidation, culminating in potent ferroptotic cell death. This work elucidates an ER-targeting nanotherapeutic strategy that specifically amplifies ferroptosis by concurrently depleting GSH and elevating AA levels, offering a compelling approach for cancer therapy. Statement of Significance Ferroptosis has emerged as a promising strategy for cancer treatment, which damages tumor cells by triggering the lethal accumulation of reactive oxygen species and lipid peroxides. However, the development of efficient ferroptosis-inducing medicines is severely impeded by the unidentified key subcellular sites of lipid peroxidation. In this work, we report an ER-targeting prodrug nanoassembly (ISSM) that synergistically promotes ferrop