Altered pHi regulation in 3T3/CFTR clones and their chemotherapeutic drug-selected derivatives

Recently (L. Y. Wei, M. J. Stutts, M. M. Hoffman, and P. D. Roepe. Biophys. J. 69: 883-895, 1996), 3T3 cells overexpressing the cystic fibrosis transmembrane conductance regulator (CFTR) were found to exhibit chemotherapeutic drug resistance and other traits of multidrug resistant (MDR) cells. In the present work, NIH 3T3/CFTR clones were selected with either doxorubicin or vincristine in incremental fashion to generate series of stable MDR cell lines that exhibit increasing levels of drug resistance. Thus C3D6 (grown in the presence of 600 nM doxorubicin) was selected from C3D4 (grown in the presence of 400 nM doxorubicin), which was selected from C3D1 (grown in the presence of 100 nM doxorubicin), which was in turn selected from the original 3T3/CFTR clone C3 (M. J. Stutts, S. E. Gabriel, J. C. Olsen, J. T. Gatzy, T. L. O'Connell, E. M. Price, and R. C. Boucher. J. Biol. Chem. 268: 20653-20658, 1993), which was not grown in the presence of chemotherapeutic drug. A similar series was generated via selection with vincristine. In both series, as well as series derived from a different CFTR clone, initial low-level drug selection increases CFTR expression without promoting MDR 1 or multidrug resistance-associated protein expression. On continued selection at higher drug concentrations, CFTR mRNA levels decrease while MDR 1 mRNA levels concomitantly increase. At each incremental step of selection, intracellular pH (pHi) increases (e.g., pHi of C3D6 > C3D4 > C3D1 > C3). Cl-/HCO3- exchange activity is significantly reduced in the drug-selected derivatives overexpressing MDR 1 but not the parental CFTR clones. The apparent set point of Na+/H+ exchange activity is significantly lower for the non-drug-selected 3T3/CFTR clones, relative to controls, but it increases on initial selection with chemotherapeutic drug. Overexpression of MDR 1 in the higher-level selectants does not appear to further perturb apparent Na+/H+ exchange. These data further describe how CFTR and MDR proteins may affect pHi regulation.