Octopus-inspired triple-engineered bacteria for precision cancer immunotherapy

Bacterial cancer therapies face precision and safety challenges, which limit their clinical application. This study presents a triple-engineered Salmonella typhimurium strain (OITE-bacteria), designed for targeted cancer therapy, inspired by octopus biology and leveraging synthetic biology and biomimetic principles. The strain incorporates SpyTag proteins on the outer membrane, allowing covalent attachment of SpyCatcher-fused quadruple RGD peptides to enhance tumor adhesion via integrin avß3 interactions. This promotes bacterial enrichment in various solid tumors and lung metastases. Subsequently, Quorum sensing induces expression of strain HtrA protein within tumors, enhancing extracellular polysaccharide-mediated immunogenicity for activating immune cells. Additionally, the bacteria locally release anti-PD1 nanobodies to anti-immunosuppressive. In murine models, OITE-bacteria achieved a ninefold increase in tumor colonization within 12 hours and sustained melanoma growth suppression for over 40 days, with a 100% response rate. This modular approach, integrating safety and efficacy, provides a new strategy in microbial therapeutics for solid tumors. Overall design: A total of 2×10^5 B16F10 cells were inoculated into C57BL/6 mice (7 weeks old, female) in the right axillary region. Once the tumor volume reached 80-120 mm^3, the mice were randomly divided into groups. Subsequently, the mice were intraperitoneally injected with physiological saline, the respective VNP bacterial strain, or modified strains (1.0×10^6 CFU per mouse). Three days later, tumor tissues were collected. To avoid spatial heterogeneity, the entire tumor was homogenized and thoroughly mixed before RNA extraction from the tissue.The extracted RNA was then subjected to RNA sequencing.