Spatial proximity of M2 macrophages to glioma cells identifies an immune niche associated with unfavorable prognosis

Background The spatial organization of immune cells may provide prognostic information beyond cell abundance in glioma. We examined whether Toll-like receptor 2 (TLR2) identifies a prognostic tumor-associated macrophage (TAM)–tumor spatial niche. Methods TCGA, CGGA, GTEx, and single-cell RNA-sequencing datasets were analyzed to define TLR2 expression, prognostic and immune associations, and its cellular source. Multiplex immunohistochemistry of a 141-case glioma tissue microarray confirmed macrophage localization and quantified TAM infiltration and 20-µm proximity to S100⁺ or TLR2⁺S100⁺ tumor cells in relation to survival. TLR2 silencing or activation in M2-like macrophages assessed polarization; co-culture models evaluated effects of silencing on glioma-cell proliferation, migration, and invasion. Results TLR2 was upregulated, associated with malignancy and poor prognosis, and expressed mainly by macrophages. TLR2⁺ tumors showed higher infiltration densities of M0 and M2-like TAMs. Spatially, TLR2⁺ M2-like TAMs were closest to glioma cells (mean distance, 2.82 µm), and greater proximity was associated with shorter overall and disease-free survival. Comparison with TLR2⁺ tumor-cell centers suggested that TLR2 expression in macrophages was the more informative spatial feature. Co-culture models showed that TLR2 silencing in M2-like macrophages significantly reduced glioma-cell migration and invasion but did not affect proliferation. Silencing also reduced M2 markers, whereas activation produced opposite changes. Conclusion Close positioning of TLR2⁺ M2-like TAMs around glioma cells defines an unfavorable spatial immune feature whose prognostic pattern varies by index tumor cell. These findings support further evaluation of TLR2 expression in macrophages and associated M2-like polarization as spatial prognostic markers and candidate targets for TAM-directed intervention.