Multi-stimuli activated PROTAC prodrug for controlled protein degradation with enhanced therapeutic effects.

Proteolysis-targeting chimeras (PROTACs) have emerged as a transformative strategy for targeted protein degradation, yet their clinical translation is hindered by systemic toxicity and poor tumor selectivity, leading to dose-limiting side effects. To overcome these limitations, we designed a multi-stimuli-responsive prodrug that enables tumor-selective activation of PROTACs in response to elevated reactive oxygen species (ROS) and glutathione (GSH) in the tumor microenvironment. By masking the hydroxyl group of the VHL ligand with a ROS/GSH-cleavable thioether-urea linker, we developed a PROTAC prodrug that responds to 1O2, HOCl, H2O2, and GSH-key mediators of oxidative stress in tumors. This proof-of-concept was verified by caging BRD4 and AR PROTAC with a methylene blue fluorophore to yield NZ-BRD and NZ-AR. Upon encountering tumor-associated stimuli, these prodrugs underwent efficient activation, releasing functional PROTACs that selectively degraded BRD4 and AR in prostate cancer cells. Intriguingly, the methylene blue liberated during activation served as a self-amplifying photosensitizer, creating a positive feedback loop that boosted 1O2 generation and further enhanced prodrug cleavage. The synergistic effect between PROTAC-mediated protein degradation and photodynamic therapy led to superior antitumor efficacy of PROTAC prodrugs in vitro and in vivo. Our work establishes a spatiotemporally controlled drug activation paradigm that combines precision protein degradation with ROS-amplified activation, presenting a promising approach to mitigate the systemic toxicity associated with conventional PROTAC therapy.