Specific Inhibition of Estrogen Receptor Alpha Function by Antisense Oligodeoxyribonucleotides

We have tested the effect of a range of antisense oligodeoxyribonucleotides (ODN) directed against the human estrogen receptor alpha (ERα) on ERα protein expression and function. Antisense ERα ODN transfected into the ERα-positive human breast carcinoma cell line MCF7-K2 showed variable responses dependent on the oligo used. The most active antisense ODN (oligo 7) decreased the levels of ERα protein by 61% as measured by Western blot analysis. Exogenous 17β-estradiol (17β-E2), but not 17α-E2, augmented this effect, with a threshold effect at 10-8 M 17β-E2. The inhibitory effect of antisense ERα oligo 7 was confirmed by measurement of functional ERα protein. 3H-17β-E2 binding to MCF7 cell extracts was inhibited to approximately 40% of control values in the presence of oligo 7. Antisense-transfected MCF7-K2 cell cultures produced a further 30% binding reduction in the presence of exogenous 17β-E2. An inhibitory effect on 17β-E2-dependent cell function was confirmed by the demonstration that ERα oligo 7-transfected MCF7-K2 cells failed to exhibit 17β-E2-stimulated cell proliferation. Exogenous 17β-E2 enhanced the inhibitory effect of the antisense ODN by increasing ODN transfection efficiency but without ERα catabolism via the proteosomal pathway, suggesting an effect of 17β-E2 on the plasma membrane and the existence of different ERα degradation pathways in the MCF7-K2 cell subclone. As 17β-E2 had no effect on ER α protein degradation, we conclude that the observed reduction of ER α protein levels is due solely to the presence of the antisense ERα ODN. Antisense ERα ODN molecules, therefore, may form the basis of effective therapies against ERα-dependent malignancies.