High TNFAIP3 expression and its induced M2-like macrophage polarization serve as prognostic markers for postsurgical distant metastasis in breast cancer

Abstract

Postsurgical distant metastasis remains the leading cause of breast cancer-related mortality, yet reliable biomarkers for identifying patients at high risk of recurrence are lacking. Tumor necrosis factor alpha-induced protein 3 (TNFAIP3) and tumor-associated macrophages (TAMs) contribute to an immunosuppressive tumor microenvironment; however, their combined prognostic value in breast cancer metastasis has not been defined. TNFAIP3 and M2 macrophage markers (CD163 and CD206) were assessed by immunohistochemistry in 96 breast cancer surgical specimens, including 48 metastatic and 48 non-metastatic cases. Expression patterns were validated using 1,085 breast cancer samples from the GEPIA2 database. Functional studies were performed using TNFAIP3-overexpression and TNFAIP3-knockdown breast cancer cell models co-cultured with macrophages. Macrophage polarization, cytokine production, and tumor cell migration and invasion were analyzed by flow cytometry, qPCR, ELISA, and Transwell assays. Predictive performance was evaluated using multivariate logistic regression and receiver operating characteristic (ROC) analyses. TNFAIP3 expression was positively correlated with CD163 ( r  = 0.322, P  < 0.001) and CD206 ( r  = 0.370, P  < 0.001) in breast tumors developing postoperative distant metastasis, with stronger correlations observed in the GEPIA2 cohort. Mechanistically, TNFAIP3 overexpression promoted CCL2 secretion, activated CCR2 signaling, and induced M2-like macrophage polarization, resulting in a 1.5- to 2.3-fold increase in tumor cell migration and invasion. Elevated TNFAIP3, CD163, and CD206 were independently associated with metastasis. TNFAIP3 alone showed good predictive performance (AUC = 0.84), while the combination of TNFAIP3 and CD163 achieved the highest accuracy (AUC = 0.90), followed by TNFAIP3 and CD206 (AUC = 0.87). TNFAIP3 promotes macrophage reprogramming through the CCL2/CCR2 axis, fostering a pro-metastatic tumor microenvironment in breast cancer. Combined assessment of TNFAIP3 and M2 macrophage markers provides a novel biomarker panel for postsurgical risk stratification and prognosis of metastatic relapse, supporting further prospective clinical validation.