Oridonin targets PRDX1 to promote apoptosis by inducing ROS-mediated ER stress and modulating autophagy

, exhibits broad-spectrum anti-cancer activity, with several derivatives currently in clinical trials. However, the molecular mechanism underlying its anticancer effects, especially its direct target proteins, remain to be fully elucidated. Here, we found that Oridonin promoted intracellular reactive oxygen species (ROS) accumulation, which in turn induced endoplasmic reticulum (ER) stress-mediated apoptosis. Moreover, ER stress was instrumental in inducing autophagy after Oridonin treatment, while blockade of autophagy further exacerbated Oridonin-induced cytotoxicity. Notably, using activity-based protein profiling (ABPP), we identified the anti-oxidant enzyme Peroxiredoxin 1 (PRDX1) as a key direct covalent target of Oridonin. By binding to Cysteine 173 of PRDX1, Oridonin increased intracellular ROS levels. Furthermore, PRDX1 over-expression mitigated, whereas PRDX1 knockdown potentiated, Oridonin-induced ROS accumulation, autophagy, and subsequently apoptosis. Overall, our results indicate that PRDX1 is a direct covalent binding target mediating Oridonin-induced apoptosis. These findings not only provide fresh insights into the core mechanism of Oridonin-induced cytotoxicity, but also highlight PRDX1 as a potential therapeutic target for renal cancer drug development.