pH and drug resistance. I. Functional expression of plasmalemmal V-type H⁺-ATPase in drug-resistant human breast carcinoma cell lines

A major obstacle for the effective treatment of cancer is the phenomenon of multidrug resistance (MDR) exhibited by many tumor cells. Many, but not all, MDR cells exhibit membrane-associated P-glycoprotein (P-gp), a drug efflux pump. However, most mechanisms of MDR are complex, employing P-gp in combination with other, ill-defined activities. Altered cytosolic pH (pH(i)) has been implicated to play a role in drug resistance. In the current study, we investigated mechanisms of pH(i) regulation in drug-sensitive (MCF-7/S) and drug-resistant human breast cancer cells. Of the drug-resistant lines, one contained P-gp (MCF-7/DOX; also referred to as MCF-7/D40) and one did not (MCF-7/MITOX). The resting steady-state pH(i) was similar in the three cell lines. In addition, in all the cell lines, HCO₃^- slightly acidified pH(i) and increased the rates of pH(i) recovery after an acid load, indicating the presence of anion exchanger (AE) activity. These data indicate that neither Na⁺/H⁺ exchange nor AE is differentially expressed in these cell lines. The presence of plasma membrane vacuolar-type H⁺-ATPase (pmV-ATPase) activity in these cell lines was then investigated. In the absence of Na⁺ and HCO₃^-, MCF-7/S cells did not recover from acid loads, whereas MCF-7/MITOX and MCF-7/DOX cells did. Furthermore, recovery of pH(i) was inhibited by bafilomycin A₁ and NBD-Cl, potent V-ATPase inhibitors. Attempts to localize V-ATPase immunocytochemically at the plasma membranes of these cells were unsuccessful, indicating that V-ATPase is not statically resident at the plasma membrane. Consistent with this was the observation that release of endosomally trapped dextran was more rapid in the drug-resistant, compared with the drug-sensitive cells. Furthermore, the drug-resistant cells entrapped doxorubicin into intracellular vesicles whereas the drug-sensitive cells did not. Hence, it is hypothesized that the measured pmV-ATPase activity in the drug-resistant cells is a consequence of rapid endomembrane turnover. The potential impact of this behavior on drug resistance is examined in a companion manuscript. Copyright (C) 1999 Elsevier Science Inc.