ALTERED PH(I) 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. Bio phys. J. 69: 883-895, 1995), 3T3 cells overexpressing the cystic fibro sis transmembrane conductance regulator (CFTR) were found to exhibit c hemotherapeutic drug resistance and other traits of multidrug resistan t (MDR) cells. In the present work, NIH 3T3/CFTR clones were selected with either doxorubicin or vincristine in incremental fashion to gener ate series of stable MDR cell lines that exhibit increasing levels of drug resistance. Thus C3D6 (grown in the presence of 600 nM doxorubici n) was selected from C3D4 (grown in the presence of 400 nM doxorubicin ), which was selected from C3D1 (grown in the presence of 100 nM doxor ubicin), which was in turn selected from the original 3T3/CFTR clone C 3 (M. J. Stutts, S. E. Gabriel, J. C. Olsen, J. T. Gatzy, T. L. O'Conn ell, 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 contin ued selection at higher drug concentrations, CFTR mRNA levels decrease while MDR 1 mRNA levels concomitantly increase. At each incremental s tep of selection, intracellular pH (pH(i)) increases (e.g., pH(i) of C 3D6 > 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 ac tivity is significantly lower for the non-drug-selected 3T3/CFTR clone s, relative to controls, but it increases on initial selection with ch emotherapeutic drug. Overexpression of MDR 1 in the higher-level selec tants does not appear to further perturb apparent Na+/H+ exchange. The se data further describe how CFTR and MDR proteins may affect pH(i) re gulation.