DIFFERENCES IN SUSCEPTIBILITY TO TUMOR-NECROSIS-FACTOR ALPHA-INDUCED APOPTOSIS AMONG MCF-7 BREAST-CANCER CELL VARIANTS

Widespread use of MCF-7 human breast carcinoma cells as a model system for breast cancer has Led to variations in these cells between differ ent laboratories. Although several reports have addressed these differ ences in terms of proliferation and estrogenic response, variations in sensitivity to apoptosis have not yet been described. Tumor necrosis factor alpha (TNF-alpha) has been shown to both induce apoptosis and i nhibit proliferation in MCF-7 cells. We observed that TNF-alpha inhibi ted proliferation in MCF-7 cell variants From three different laborato ries (designated M, L, and N). MCF-7 M cells were resistant to TNF-alp ha-induced apoptosis, whereas MCF-7 L cells were moderately resistant to the effect of TNF-alpha. A third variant, MCF-7 N, underwent apopto sis when exposed to TNF-cu. Analysis of the p55 TNF-alpha receptor (TN FR) 1 expression revealed the greatest expression in MCF-7 N cells, wh ereas the MCF-7 L and M cells expressed 89 and 67% of MCF-7 N cell TNF R1 levels, respectively. Ceramide generation occurred in all three var iants in response to TNF-alpha treatment, with MCF-7 N cells expressin g the greatest increase. Cleavage of the CPP32/caspase 3 substrate pol y(ADP-ribose) was observed in MCF-7 N and L cells as early as 3 and 6 h, respectively, but poly(ADP-ribose) cleavage was not observed in MCF -7 M cells. The delayed protease activation in the L variant may repre sent the mechanism by which these cells display delayed sensitivity to TNF-alpha-induced apoptosis. Expression of the Bcl-2, Mcl-1, Bcl-X, B ax, and Bak proteins was analyzed to determine whether the differences in MCF-7 cell sensitivity to apoptosis could be correlated to the dif ferential expression of these proteins. Whereas Bak, BcI-X, and Mcl-1 levels were identical between variants, the levels of Bcl-2 were 3.5-3 .8-fold higher and the levels of Bax were 1.5-1.7-fold lower in the re sistant variants (M and L) as compared with those of the sensitive var iant (N). Taken together, these results suggest that differences in su sceptibility to TNF-alpha-induced apoptosis among MCF-7 breast cancer cell variants may be explained by differences in TNFR expression, cera mide generation, differential expression of the Bcl-2 family of protei ns, and protease activation.