REDUCED DRUG ACCUMULATION AND MULTIDRUG-RESISTANCE IN HUMAN BREAST-CANCER CELLS WITHOUT ASSOCIATED P-GLYCOPROTEIN OR MRP OVEREXPRESSION

MCF-7 human breast cancer cells selected in Adriamycin in the presence of verapamil developed a multidrug resistant phenotype, which was cha racterized by as much as 100,000-fold resistance to mitoxantrone, 667- fold resistance to daunorubicin, and 600-fold resistance to doxorubici n. Immunoblot and PCR analyses demonstrated no increase in MDR-1 for M RP expression in resistant cells, relative to parental cells. This phe notype is similar to one previously described in mitoxantrone-selected cells. The cells, designated MCF-7 AdVp, displayed a slower growth ra te without alteration in topoisomerase ll alpha level or activity. inc reased efflux and reduced accumulation of daunomycin and rhodamine wer e observed when compared to parental cells. Depletion of ATP resulted in complete abrogation of efflux of both daunomycin and rhodamine. No apparent alterations in subcellular daunorubicin distribution were obs erved by confocal microscopy. No differences were noted in intracellul ar pH. Molecular cloning studies using DNA differential display identi fied increased expression oi the alpha subunit of the amiloridc-sensit ive sodium channel in resistant cells. Quantitative PCR studies demons trated an eightfold overexpression of the alpha subunit of the Na+ cha nnel in the resistant subline. This channel may be linked to the mecha nism of drug resistance in the AdVp cells. The results presented here support the hypothesis that a novel energy-dependent protein is respon sible for the efflux in the AdVp cells. Further identification awaits molecular cloning studies. J. Cell. Biochem. 65:513-526. (C) 1997 Wile y-Liss, Inc.