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It is increasingly recognized that drug-resistant cells undergo transitions not directly linked to "classical" drug resistance. We examined the expression of growth factors, growth factor receptors, and the estrogen receptor in 17 drug-resistant and 2 revertant human breast cancer sublines to provide an understanding of the phenotypic changes that occur and how these changes could affect the biology of the cell. These sublines were derived from five parental human breast cancer cell lines (MCF-7, ZR75B, T47D, MDA-MB-231, and MDA-MB-453). The expression of estrogen receptor was absent or decreased in 6 of the 15 resistant MCF-7, ZR75B, and T47D sublines. Increases of as much as 49-fold compared to parental levels were observed in transforming growth factor alpha, epidermal growth factor receptor, c-erbB2, and/or c-erbB3 mRNA expression in 14 of the 17 resistant sublines. Altered amphiregulin and insulin-like growth factor-I receptor expression was observed in nine and four drug-resistant sublines, respectively. No major alterations were observed in epidermal growth factor and c-erbB4 expression. Few alterations were observed in two sublines derived from estrogen receptor-negative cells. Higher levels of phosphotyrosine residues were detected in a subset of the resistant sublines, indicating an increased tyrosine kinase activity in these cells. Interestingly, decreased growth rates were observed in all of the sublines, despite up-regulated growth factor-related gene expression. Taken together, these data suggest that loss of estrogen receptor, increased expression of growth factor pathway genes, and decreased growth rate regularly occur in drug-resistant breast cancer cells. Although we do not know whether the altered expression of growth factor pathway genes is linked as a cause or a consequence of the reduced growth rate, it is well established that decreased growth rate confers drug resistance. These phenotypic changes in drug-resistant human breast cancer cells could serve to initiate, support, or extend the drug resistance.

Human breast cancer cells selected for multidrug resistance frequently overexpress ligands and receptors in the epidermal growth factor (EGF) receptor family. To determine whether this overexpression contributes to the drug resistant phenotype, EGF receptor transfected ZR75B human breast cancer cells were examined. Two EGF receptor overexpressing clones were evaluated: clone 11 with > 1 x 10(6) sites, and clone 13 with 310,000 receptor sites/cell. These were compared with clone 2-neo, which was transfected with the neomycin gene only and contained 43,000 receptor sites/cell. The EGF receptor overexpressing clones and the neo transfected control clone displayed comparable growth rates. Cytotoxicity analyses were performed with doxorubicin, vinblastine, cisplatin and 5-fluorouracil to determine the sensitivity of the clones to antineoplastic drugs. The EGF receptor overexpressing clones were found to be 1.5-5.6 times more resistant to the four drugs tested. This increase in the IC50 conferred a selective advantage when grown in the presence of 2, 3 and 6 ng/ml doxorubicin. Clone 13 cells overtook a mixed population which began with clone 2-neo comprising 95% of the cells. Clone 2-neo remained the dominant clone in the absence of drug. Finally, after long-term selection of the clones with 6 ng/ml doxorubicin, clone 2-neo became fourfold more resistant than the unselected clone 2-neo, a level which was comparable to that found in the EGF receptor overexpressing clones 11 and 13. No additional increase in resistance was observed for these clones, suggesting that clone 2-neo had developed additional resistance mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)