The up regulation of ER by 4OHT contributes to anti oestrogen resistance in ER negative breast cancer cells ER positive breast cancers generally have a better prog nosis and are often responsive to anti oestrogen different therapy, which was the first example of a successful therapy targeting a specific protein, the ER. Unfortunately, ER negative breast cancers are more aggressive and un responsive to anti oestrogens. Although the transform ation of ER negative into ER positive cells by gene therapy or ER gene re expression are common strat egies to restore anti oestrogen responsiveness, we observed in this study that MDA MB 231 ER negative cells were intrinsically resistant to 4OHT despite the overexpression of ER.

Proliferation assays to determine the concentration and time dependent effects of 4OHT on MDA MB 231, showed that this drug stimulated cancer cell proliferation at concentrations as low as 1 nM. Stimulation of cancer cell proliferation, in the presence of 4OHT, was also observed in hormone deprived conditions, which indicated that this effect was independent of oestrogens in the culture medium. This class of resistance to tamoxifen was firstly discovered by Gottardis and Jordan in MCF7 cells and the results agree with additional observation in which long term tam oxifen treatment of MDA MB 231 cells increased their growth and their aggressiveness in animal tumours. ER positive tumours are more differentiated and have lower metastatic potential than ER negative tumours.

Because this suggests a protective role of the oestrogen receptor in tumour progression and metastasis, we next examined whether the 4OHT induced expression of ER resulted in a decreased metastatic potential of MDA MB 231. Using a wound healing assay, we observed that the treatment of MDA MB 231 with 4OHT greatly increased the migratory ability of these cells. These results indicated that the re expression of ER in ER negative breast cancer cells may promote cell growth and the resistance of these cells to anti oestrogen therapy. To determine whether ER plays a role in 4OHT resist ance, we examined the effect of a decreased level of ER on the growth of MDA MB 231 cells. Because the treat ment of these cells with U0126 clearly inhibited the ER expression induced by 4OHT, we first ana lysed whether this drug also inhibited 4OHT induced cell proliferation. As shown in this Figure, U0126 reduced the growth of cells that were treated with 4OHT and untreated cells to the same extent. These results indi cated that the MAPK cascade controls the proliferation of this breast cancer cell line. It is well known that mem brane ER plays a role in the temporal coordination of phosphorylation/dephosphorylation Brefeldin_A events for the ERKs in breast cancer cells.