Cathepsin B‐Cleavable Doxorubicin Prodrug Nanoparticles for Eliciting Cancer‐Specific Immunogenic Cell Death and Minimizing Systemic Toxicity

ABSTRACT Doxorubicin (DOX) is a potent anticancer drug, but its poor tumor selectivity and systemic toxicity limit clinical use and hinder cancer immunotherapy despite its ability to induce immunogenic cell death (ICD). To overcome these limitations, we developed self‐assembling, cathepsin B‐cleavable peptide ( Phe‐Arg‐Arg‐X : FRRX)‐DOX prodrugs forming nanoparticles via hydrophobic and π – π interactions. Among them, the FRRL‐DOX nanoparticles (cathepsin B‐cleavable DOX prodrug nanoparticles; CatB‐NPs) exhibited stable nanoparticle structure with an average diameter of 165.1 ± 24.7 nm and enhanced tumor accumulation through the enhanced permeation and retention (EPR) effect. In the cell culture systems, CatB‐NPs showed the cancer cell‐specific ICD while minimizing non‐specific damage to normal cells and immune cells. In the CT26 tumour‐bearing mice, CatB‐NPs exhibited improved pharmacokinetics, enhanced tumor accumulation, and significantly greater therapeutic efficacy. When combined with anti‐PD‐L1 antibody, CatB‐NPs achieved 100% complete tumor regression, with elevated dendritic cell (DC) activation, cytotoxic T cell infiltration, and durable immunological memory. Furthermore, CatB‐NPs greatly reduced the cardiotoxicity and splenic toxicity associated with free DOX, preserving tissue structure and immune function. This study highlights the clinical potential of self‐assembling and cathepsin B‐cleavable prodrug nanoparticles as a next‐generation anticancer drug with improved safety and immune compatibility.