Engineered Calprotectin-Responsive Probiotics for Inflammatory Bowel Disease Treatment

Abstract Inflammatory bowel disease (IBD) is a complex intestinal disorder for which effective treatment options remain limited. The modulation of gut microbiota balance using probiotics holds promise as a therapeutic approach to restore intestinal microenvironment homeostasis. However, the clinical translation of conventional probiotics is constrained by the suboptimal therapeutic efficacy. Here, we constructed a recombinant Escherichia coli Nissle 1917 strain (EcN-CR-IL2) based on an optimized calprotectin-responsive ykgMO promoter to achieve inflammation-dependent expression of the immunomodulatory cytokine interleukin-2 (IL-2), which selectively expands regulatory T cells to suppress aberrant intestinal inflammation. The optimized promoter achieved approximately 100-fold greater induction than the native ykgMO promoter and responded robustly to calprotectin both in vitro and in a DSS-induced murine colitis model in vivo. Oral administration of EcN-CR-IL2 significantly alleviated colitis symptoms in both prophylactic and therapeutic settings, suppressed proinflammatory cytokine levels, restored intestinal barrier integrity, and shifted gut microbiota composition toward a more homeostatic profile. Importantly, the therapeutic output of EcN-CR-IL2 correlated with local calprotectin levels, enabling on-demand modulation of anti-inflammatory effects in response to disease progression. This study presents a probiotic delivery system that combines real-time sensing of a clinically validated biomarker with autonomous therapeutic production, providing a new approach for the treatment of chronic intestinal inflammatory diseases.