An orally administered biohybrid complex leveraging probiotic guidance to treat colorectal cancer via in situ vaccination.

Oral therapy for colorectal cancer (CRC) holds inherent promise for enhancing patient compliance and enabling gastrointestinal targeting, while its application is hampered by low oral drug utilization efficiency and precise colonic localization. This study developes an orally delivered biohybrid complex, CS@CB-Lipo@5-FU/R837, which leverages the hypoxic tropism of the probiotic Clostridium butyricum (CB) for active tumor targeting. The core of the system consists of nanoliposomes co-loaded with 5-fluorouracil (5-FU) and the TLR7 agonist, R837, which are site-specifically anchored to CB via bioorthogonal conjugation. A chitosan-based outer coating ensures gastrointestinal stability and enables enzyme-responsive drug release in the tumor-colonized environment. Within the tumor, the complex orchestrates a coordinated immunotherapeutic cascade: 5-FU induces immunogenic cell death, releasing tumor antigens and damage-associated molecular patterns, while R837 promotes dendritic cell maturation and antigen presentation. This spatiotemporally coupled "antigen-adjuvant' delivery effectively mimics an in situ vaccination mechanism, stimulating potent antitumor immunity. Evaluation in orthotopic and subcutaneous CRC models shows treatment leads to tumor growth inhibition, remodeling of the immune microenvironment, and extended survival. This work establishes a versatile strategy for oral in situ vaccines based on probiotic-guided delivery and localized immune activation.