Bacteria biohybrids integrating anticancer peptide-loaded nanoparticles for tumor immunotherapy through pyroptosis activation

Developing bacterial-based biohybrid systems that effectively integrate tumor-targeting, immune modulation, and advanced drug delivery remains a major challenge in cancer therapy. This is primarily due to the complexities of achieving selective tumor colonization, overcoming immune clearance mechanisms, and ensuring controlled drug release within the tumor microenvironment. Here, we introduce P/L@EcN, a bacteria biohybrid system that conjugates ROS-responsive, ruxotemitide (LTX-315)-loaded poly(ethylene glycol)-<i>block</i>-poly(lactide-<i>co</i>-glycolide) (PEG-PLGA) nanoparticles (P/L-NPs) with the tumor-targeting probiotic <i>Escherichia coli</i> Nissle 1917 (EcN) <i>via</i> copper-free azide-alkyne click chemistry. This hybrid system exhibits enhanced tumor accumulation, improved cellular uptake, and deep tumor penetration, while effectively inducing pyroptosis through caspase-1-dependent pathways. In an <i>in vivo</i> orthotopic breast cancer model, P/L@EcN enhanced anti-tumor immunity by remodeling the tumor microenvironment, promoting the macrophage M1/M2 ratio, and reducing myeloid-derived suppressor cells (MDSCs), ultimately achieving significant tumor growth suppression without systemic toxicity. Together, these findings establish P/L@EcN as a promising biohybrid immunotherapy strategy that integrates bacterial-mediated targeting with immune activation, offering a powerful approach for cancer treatment.