Mistletoe Lectin Overcomes Macrophage-Mediated Chemo-Resistance in 3D Co-Culture Models of Triple-Negative Breast CancerOA

Objective Tumor-associated macrophages(TAMs)contribute to chemoresistance in triple-negative breast cancer(TNBC),yet strategies to reprogram TAMs while enhancing chemotherapy efficacy remain limited.This study investigated whether Viscum album L.var.coloratum agglutinin(VCA)could sensitize TNBC cells to doxorubicin(DOX)and modulate TAM-mediated chemoresistance in three-dimensional(3D)co-culture models.Methods MDA-MB-231 TNBC cells were co-cultured with RAW264.7 macrophages in collagen-embedded 3D spheroids.Spheroid viability was assessed using an ATP-based luminescent assay.Cytokine secretion and epithelial-mesenchymal transition(EMT)markers were measured using ELISA and Western blotting.Drug synergy was evaluated using combination index(CI)calculations.Results VCA-DOX combination demonstrated synergistic cytotoxicity exclusively in co-culture spheroids(CI=0.72),reducing viability to 25.9%(p<0.001),while showing no synergy in monoculture(CI=1.52).Combination treatment decreased VEGF secretion by 49%and IL-6 by 74%,while elevating TNF-α2.7-fold,suggesting macrophage reprogramming.VCA enhanced E-cadherin expression while suppressing mesenchymal markers in co-culture spheroids and reduced Matrigel invasion by 60%(p<0.001).Conclusion VCA-DOX combination demonstrates synergistic anticancer effects through TAM reprogramming and enhanced chemosensitization specifically in 3D co-culture models,warranting further investigation for overcoming macrophage-mediated chemoresistance in TNBC.