Differentiation of cultured human breast cancer cells (AU‐565 and MCF‐7) associated with loss of cell surface HER‐2/neu antigen

Abstract

The relationship between terminal cell differentiation and changes in the subcellular levels of the HER-2/neu antigen was investigated in cultured human breast cancer cells: AU-565 cells (which overexpress the HER-2/neu gene) and MCF-7 cells (which do not overexpress this gene). Differentiation was achieved by treating the cells with mycophenolic acid (MPA), phorbol 12-myristate 13-acetate (PMA), retinoic acid (RA), or the TA-1 monoclonal antibody to the extracellular domain of the HER-2/neu protein. Ten to twenty percent of the cells in untreated, sparsely growing AU-565 cultures exhibited morphological maturation characterized by large lacy nuclei surrounded by sizable flat cytoplasms. A fraction of these cells harbored milk factors such as casein and large lipid droplets. Treatment of the AU-565 cells for 4 d with 9 microM MPA, 1.6 nM PMA, 2.5 microM RA, or 1 microgram/mL TA-1 antibody resulted in cell growth inhibition and an increase in the percentage of cells (48-97%) that exhibit a mature phenotype. MCF-7 cells were also susceptible to differentiation by MPA and RA, but to a lesser degree than the AU-565 cells. Differentiation in the AU-565 and MCF-7 cells was associated with reduced immunostaining for the HER-2/neu protein at the cell surface membrane and with a transient increased diffuse immunostaining for this protein in the cytoplasm.

Effects of retinoic acid and fenretinide on the c-erbB-2 expression, growth and cisplatin sensitivity of breast cancer cells

“…Here, we have demonstated that ATRA and 4-HPR inhibited c- Bacus et al (1990) and Pellegrini et al (1995). No retinoic acid response element has been identified so far, whereas AP-1, AP-2 and SP-l sites have been found in the regulatory region of c-erbB-2.…”

c-erbB3 and c-erbB4 expression is a feature of the endocrine responsive phenotype in clinical breast cancer

“…Elevated c-erbB3 expression in cancer cells particularly in ER positive tumours could therefore represent a di erentiated feature. In this light, it is noteworthy that aheregulin, a ligand for c-erbB3/c-erbB4 receptors, is also more likely to be elevated in ER positive disease (Knowlden and Nicholson, in preparation) and has been shown to initiate cellular di erentiation in breast cancer cells in vitro (Bacus et al, 1992(Bacus et al, , 1993. Indeed, high concentrations of c-erbB3/c-erbB4 ligands, may even result in a parallel inhibition of cellular proliferation (Bacus et al, 1992;Grunt et al, 1995).…”

“…In this light, it is noteworthy that aheregulin, a ligand for c-erbB3/c-erbB4 receptors, is also more likely to be elevated in ER positive disease (Knowlden and Nicholson, in preparation) and has been shown to initiate cellular di erentiation in breast cancer cells in vitro (Bacus et al, 1992(Bacus et al, , 1993. Indeed, high concentrations of c-erbB3/c-erbB4 ligands, may even result in a parallel inhibition of cellular proliferation (Bacus et al, 1992;Grunt et al, 1995). Although there is clearly substantial overlap of the expression of individual erbB family members within the breast cancer population, the di erential e ects of c-erbB3/cerbB4 and the EGFR may result from the ability of cerbB3 to e ciently recruit PI3 Kinase (Solto et al, 1994;Fedi et al, 1994;Prigent and Gullick, 1994) and hence pro®ciently activate additional signal transduction pathways to those employed by EGFR.…”

Increased expression of c-erbB-2 in hormone-dependent breast cancer cells inhibits cell growth and induces differentiation

“…However, while estrogen supplementation strongly increased thymidine incorporation by mock cells, the p185-overexpressing ME#1 clone maintained the same thymidine incorporation both in presence or absence of estrogens, which was much lower than in the estrogen stimulated mock cells ( Figure 4B). To determine whether inhibition of cell growth in complete medium was related to induction of di erentiation, we tested for the presence of lipid droplets, which are indicative of a mature phenotype (Bacus et al, 1990). As shown in Figure 5, c-erbB-2-transfected ME#106 cells (panel a) produced droplets containing neutral lipids that were not detectable in mock cells (panel b).…”

“…Neutral lipids were visualized using a modi®cation of the Oil Red O in propylene glycol method (Bacus et al, 1990). Brie¯y, cells grown on Lab-Tek slides were ®xed by a quick dip in 7208C methanol/acetate (v/v) and incubated at room temperature in absolute propylene glycol for 5 min.…”