ROLE OF C-MYC IN TAMOXIFEN-INDUCED APOPTOSIS IN ESTROGEN-INDEPENDENT BREAST-CANCER CELLS

Background: The antiestrogen tamoxifen (TAM) is effective in the treat ment of estrogen receptor (ER)-positive as well as some ER-negative br east cancers. However, the precise mechanism of action of TAM, especia lly in estrogen-independent cells, remains unclear. Previous work by o ur laboratory has demonstrated that TAM induces the morphologic and bi ochemical changes that are characteristic of apoptosis in both PR-posi tive and PR-negative cells. Purpose: We compared the effect of TAM at a clinically achievable concentration on cell growth and apoptosis wit h the effect of TAM on c-myc (also known as C-MYC) messenger RNA (mRNA ) and protein expression in PR-negative MDA-231 cells. Methods: MDA-23 1 cells were treated for up to 72 hours with 1.0 mu M TAM alone or in the presence of 50 mu M c-myc antisense or nonsense oligonucleotides, c-myc mRNA expression was determined by northern blot analysis, protei n expression by western blot analysis, cell growth inhibition by cell counts, and DNA cleavage by agarose gel electrophoretic analysis. Diff erences between the mean values from different treatment groups were c ompared with the use of the two-sided Wilcoxon rank-sum test. Results: TAM treatment for 72 hours increased c-myc mRNA fivefold (from a rela tive radiolabeled hybridization signal intensity of 17 +/- 4 up to 93 +/- 10; P<.05) and c-Myc protein threefold (from a relative immunofluo rescence signal intensity of 28 +/- 7 up to 83 +/- 21; P<.05). The ind uction of c-myc by TAM was accompanied by internucleosomal DNA cleavag e characteristic of apoptotic cell death, Addition of c-myc antisense oligonucleotide (5'-CACGTTGAGGGGCAT-3') to MDA-231 cells resulted in a nearly twofold decrease of basal c-myc mRNA (P<.05) and a sevenfold d ecrease of basal c-Myc protein (P<.05) expression. Addition of c-myc a ntisense oligomer also antagonized the TAM-induced increase in c-myc m RNA (P<.05) and protein expression (P<.05) and inhibited TAM-induced c ytostasis (P<.01) and apoptosis. In parallel experiments, addition of the nonsense oligomer had no effect on any of the measured parameters. Conclusions: These results indicate that the effects of TAM on PR-neg ative MDA-231 cells may be mediated through c-myc overexpression, c-my c may play a critical role in the growth and progression of MDA-231 br east cancer cells.