Tumor microenvironment acidity and breast cancer cell secretome modulate macrophage acid-base homeostasis and phagocytosis

Tumor-associated macrophages (TAMs) shape the tumor microenvironment in triple-negative breast cancer (TNBC), where they engage in sustained crosstalk with cancer cells and are influenced by the acidic extracellular pH (pHe) typical of solid tumors. Acidic pHe suppresses anti-tumor immunity and promotes tumor aggressiveness, yet how macrophages maintain acid–base homeostasis under these conditions, and how cancer-cell secretome affects this process, remains unclear. This study examined the contribution of four major pH-regulatory transporters to macrophage homeostasis across distinct polarization states and assessed changes in their localization after exposure to secretomes from wild-type (WT) or acid-adapted (AA) TNBC cells. Our findings show that macrophage phenotypes differ in their reliance on pH-regulatory transporters, with M2-like macrophages displaying the highest metabolic resilience. AA conditioned medium increased plasma membrane localization of NBCn1, NHE1, and MCT4 in M0 macrophages, suggesting an acid-adaptive response. Proteomic analysis revealed an enrichment of metabolic and immunomodulatory components – predominantly exosomal – in the AA secretome. Functionally, M1-like macrophages showed reduced phagocytosis of AA compared with WT cancer cells. These results identify mechanisms through which acid-adapted cancer cells influence macrophage behavior and point to potential targets for reprogramming TAM function in TNBC.