Spatial interaction between Gli1 high tumor cells and M2 macrophages predicts survival and recurrence in gastric cancer

Purpose: This study investigated the prognostic significance of GLI1 expression in gastric cancer by analyzing its association with the infiltration and spatial proximity of tumor-associated M2 macrophages identified by CD163 and CD206. We evaluated how the combined GLI1-M2 macrophage spatial phenotype influences clinical outcomes, including survival and recurrence patterns. Methods: GLI1 expression and M2 macrophage infiltration were assessed using immunohistochemistry and spatial biology approaches, including multiplex immunofluorescence and machine-learning-based digital pathology. CD163 and CD206 defined M2 macrophage populations. Spatial metrics included GLI1 expression intensity, M2 macrophage density, and the intercellular distance between GLI1-positive tumor cells and CD163/CD206-positive macrophages. Associations with clinical outcomes were analyzed using Kaplan-Meier survival curves and multivariate Cox regression. Results: High GLI1 expression demonstrated a strong correlation with increased infiltration of CD163/CD206-positive M2 macrophages (p=0.001) and reduced spatial distance between tumor cells and M2 macrophages (p=0.01). The GLI1-high and M2-close phenotype was associated with markedly reduced overall survival, higher recurrence rates including early relapse, and a microenvironment enriched with immunosuppressive and tumor-promoting signals. In contrast, tumors with lower GLI1 expression and greater separation from M2 macrophages showed improved survival outcomes and reduced recurrence. Conclusions: GLI1 expression and the spatial proximity of CD163/CD206-positive M2 macrophages serve as strong prognostic indicators in gastric cancer. The GLI1-high and M2-close phenotype identifies a clinically vulnerable subgroup with significantly poorer outcomes. These findings highlight the biological relevance of the GLI1-M2 macrophage axis and suggest that targeting GLI1 signaling or reprogramming M2 macrophages may provide therapeutic benefit and reduce recurrence risk in gastric cancer.