SCOPE: Inflammatory bowel disease (IBD) involves a range of immune-mediated disorders marked by systemic and local intestinal inflammation. We synthesized a novel compound DJ-X-025 and uncovered its anti-inflammatory properties using lipopolysaccharide (LPS)-induced RAW 264.7 macrophages in vitro and a dextran sodium sulfate (DSS)-induced model of colitis. METHODS AND RESULTS: We evaluated the alteration in cell morphology, cytoskeletal proteins, and inflammatory markers of DJ-X-025 treated LPS-stimulated RAW 264.7 macrophages. We administered DJ-X-025 by oral gavage in DSS-induced colitis, examined colon histology, and alterations of immune cells by flow cytometry, and performed molecular studies using RT-qPCR and western blot analysis. DJ-X-025 treatment markedly altered the morphology of LPS-treated RAW 264.7 macrophages from elongated to round shapes, modulated actin and tubulin, and reduced the level of inflammatory markers like TNF-alpha, IL-1beta, IL-6, and iNOS. Further, we observed that DJ-X-025 steered to improve colon length, muscularis mucosa thickness, and colon inflammatory score compared to the DSS group alone. DJ-X-025 effectively inverted the increased population of activated T cells, Th17, and macrophages in lamina propria by DSS treatment, leading to a substantial reduction in the inflammatory response in the colon. Strikingly, DJ-X-025 treatment enhanced the expression of occludin and diminished the expression of NF-kappaB and phosphorylation of STAT3 in the colon of DSS-treated mice compared to DSS-alone. Additionally, DJ-X-025 induced the expression of Foxp3 in the colon and, reduced systemic inflammatory cytokine/chemokine levels further supporting its immunomodulatory effects. These results suggest that DJ-X-025 is linked to the induction of occludin expression and decreased expression of p-STAT3/NF-kappaB and Th17 response in the colon, which together suppresses systemic and colon inflammatory cytokines for effective amelioration of experimental colitis. CONCLUSION: These findings suggest that DJ-X-025 might be a promising therapeutic agent for the treatment of IBD.