Engineered Salmonella expressing IL-18 as a therapeutic strategy for colorectal cancer

Abstract Colorectal cancer (CRC) remains the second leading cause of cancer-related mortality worldwide, largely due to therapeutic resistance and metastatic progression. Novel strategies capable of targeting the tumor microenvironment and modulating key oncogenic pathways are urgently needed. In this study, we investigated the antitumor effect of attenuated Salmonella Typhimurium SL3261 genetically engineered to express interleukin-18 (IL-18) in both precursor (pro-IL-18) and bioactive (IL-18) forms. Treatment with bacterial supernatants significantly reduced cell viability in colorectal cancer cell lines (CT-26 and HCT-8), mouse organoids and patient-derived organoids. Notably, organoids exhibited pronounced sensitivity, with near-complete loss of viability after prolonged exposure, particularly in response to bioactive IL-18. Co-culture experiments further demonstrated enhanced cytotoxicity, with IL-18–expressing bacteria inducing a rapid and sustained reduction in organoid viability. IL-18 delivery was linked with modulation of tumor-associated inflammatory responses. These findings suggest a shift towards a pro-inflammatory and antitumor microenvironment. The effects observed with recombinant IL-18 alone partially recapitulated the bacterial-mediated responses, indicating a synergistic contribution of bacterial factors and cytokine signalling. Overall, our results demonstrate that Salmonella -mediated delivery of IL-18 exerts potent antitumor effects in CRC models, highlighting its potential as a novel immunotherapy.