Reduced drug accumulation and multidrug resistance in human breast cancer cells without associated P-glycoprotein or MRP overexpression

• Published in: Journal of cellular biochemistry 1997-06, Vol.65 (4), p.513-526
• Main Authors: Lee, Jong Seok, Scala, Stefania, Matsumoto, Yoshihito, Dickstein, Bruce, Robey, Rob, Zhan, Zhirong, Altenberg, Guillermo, Bates, Susan E.
• Format: Article
• Language: English
• Subjects: Antibiotics, Antineoplastic - metabolism; Antigens, Neoplasm; ATP-Binding Cassette Transporters - physiology; ATP-Binding Cassette, Sub-Family B, Member 1 - physiology; Breast Neoplasms - drug therapy; Breast Neoplasms - metabolism; Cell Division; Cloning, Molecular; Daunorubicin - metabolism; DNA Topoisomerases, Type II - metabolism; DNA-Binding Proteins; drug resistance; Drug Resistance, Multiple; Drug Resistance, Neoplasm; Female; Fluorescent Dyes; Humans; Isoenzymes - metabolism; mitoxantrone; Multidrug Resistance-Associated Proteins; Neoplasm Proteins - physiology; non-Pgp MDR; Phenotype; Polymerase Chain Reaction; rhodamine; Rhodamines - metabolism; Sodium Channels - genetics; Subcellular Fractions - metabolism; Tumor Cells, Cultured
• ISSN: 0730-2312; 1097-4644
• DOI: 10.1002/(SICI)1097-4644(19970615)65:4<513::AID-JCB7>3.0.CO;2-R

MCF‐7 human breast cancer cells selected in Adriamycin in the presence of verapamil developed a multidrug resistant phenotype, which was characterized by as much as 100,000‐fold resistance to mitoxantrone, 667‐fold resistance to daunorubicin, and 600‐fold resistance to doxorubicin. Immunoblot and PCR analyses demonstrated no increase in MDR‐1 or MRP expression in resistant cells, relative to parental cells. This phenotype is similar to one previously described in mitoxantrone‐selected cells. The cells, designated MCF‐7 AdVp, displayed a slower growth rate without alteration in topoisomerase IIα level or activity. Increased efflux and reduced accumulation of daunomycin and rhodamine were 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 observed by confocal microscopy. No differences were noted in intracellular pH. Molecular cloning studies using DNA differential display identified increased expression of the alpha subunit of the amiloride‐sensitive sodium channel in resistant cells. Quantitative PCR studies demonstrated an eightfold overexpression of the alpha subunit of the Na+ channel in the resistant subline. This channel may be linked to the mechanism of drug resistance in the AdVp cells. The results presented here support the hypothesis that a novel energy‐dependent protein is responsible for the efflux in the AdVp cells. Further identification awaits molecular cloning studies. J. Cell. Biochem. 65:513–526. © 1997 Wiley‐Liss Inc.