METTL3-mediated m6A modification of CXCR4 drives M2 macrophage polarization and suppresses anti-tumor immunity in esophageal cancer.

Dense tumor-associated macrophage infiltration correlates with poor prognosis in esophageal cancer (EC), yet the underlying molecular mechanisms remain undefined. An EC cell-macrophage co-culture system was established. M2 polarization was evaluated by flow cytometry and ELISA. qPCR and Western blot were employed to detect mRNA and protein levels. MeRIP-PCR assessed CXCR4 m6A methylation. Colony formation, wound-healing, and apoptosis assays measured EC cell proliferation, migration, and apoptosis. Compared with macrophages co-cultured with normal esophageal epithelial cells, those co-cultured with EC cells exhibited increased IL-10 and TGF-β secretion, accompanied by up-regulation of CD206, CD163, CXCR4, and METTL3. METTL3 up-regulated CXCR4 by mediating CXCR4 m6A methylation. METTL3-overexpressing macrophages elevated CD206/CD163 levels and IL-10/TGF-β secretion, whereas CXCR4 knockdown reversed these effects. Moreover, METTL3-high macrophages promoted EC cell proliferation and migration, up-regulated PD-L1, and inhibited apoptosis. METTL3 is up-regulated in EC-associated macrophages, promotes M2 polarization via m6A modification of CXCR4, and thereby accelerates EC malignant progression.