The net acid extruders NHE1, NBCn1 and MCT4 promote mammary tumor growth through distinct but overlapping mechanisms.

High metabolic and proliferative rates in cancer cells lead to production of large amounts of H⁺ and CO₂, and as a result, net acid extruding transporters are essential for the function and survival of cancer cells. We assessed protein expression of the Na⁺/H⁺ exchanger NHE1, the Na⁺‐ HCO 3 − cotransporter NBCn1, and the lactate‐H⁺ cotransporters MCT1 and −4 by immunohistochemical analysis of a large cohort of breast cancer samples. We found robust expression of these transporters in 20, 10, 4 and 11% of samples, respectively. NHE1 and NBCn1 expression both correlated positively with progesterone receptor status, NHE1 correlated negatively and NBCn1 positively with HER2 status, whereas MCT4 expression correlated with lymph node status. Stable shRNA‐mediated knockdown (KD) of either NHE1 or NBCn1 in the MDA‐MB‐231 triple‐negative breast cancer (TNBC) cell line significantly reduced steady‐state intracellular pH (pH_i) and capacity for pH_i recovery after an acid load. Importantly, KD of any of the three transporters reduced <italic>in vivo</italic> primary tumor growth of MDA‐MB‐231 xenografts. However, whereas KD of NBCn1 or MCT4 increased tumor‐free survival and decreased <italic>in vitro</italic> proliferation rate and colony growth in soft agar, KD of NHE1 did not have these effects. Moreover, only MCT4 KD reduced Akt kinase activity, PARP and CD147 expression and cell motility. This work reveals that different types of net acid extruding transporters, NHE1, NBCn1 and MCT4, are frequently expressed in patient mammary tumor tissue and demonstrates for the first time that they promote growth of TNBC human mammary tumors <italic>in vivo</italic> via distinct but overlapping mechanisms. [ABSTRACT FROM AUTHOR]