Figure 4. Ligand-dependent proteasomal degradation of ERα.
(a) Steroid-deprived MCF-7 cells were treated with 10 μM OBHS-N ligands, OBHS or E2 and 10 nM fulvestrant (Fulv) for 6 hr. Western blot was performed to detect ERα protein levels.
(b) Contributions of the OBHS-N R1 and R2 groups to ERα degradation. Each data point represents the normalized average value for an OBHS-N compound relative to tubulin (n = 2 experiments).
(c) Dose-dependent degradation of ERα protein in MCF-7 cells treated with fulvestrant or OBHS-N compound 12. Cells were treated with indicated doses of compounds for 48 hr and western blot performed as in panel a.
(d) ERα protein expression in the uterus of ovariectomized female mice treated with vehicle, fulvestrant, OBHS-N compounds 12 and 13 as described in Figure 3f. ERα+ cells were quantified by immunohistochemistry (1-way ANOVA, Tukey’s Multiple Comparison Test; *, P < 0.05; **, P < 0.01) (e) Steroid-deprived MCF-7 cells were treated with 100 nM fulvestrant, 10 μM 4-OHT, 10 μM 12, 10 μM 13, 10 μM 4-OHT + 10 nM fulvestrant, 10 μM 4-OHT + 10 μM 12 or 10 μM 4-OHT + 10 μM 13 for 6 hr. Western blot was performed to detect ERα protein levels.