Fig. 2

SCR-6852 is a potent SERD to induce ERα degradation and anti-proliferation of ER + breast cancer cell lines. a The comparison of in-house compounds potency to fulvestrant or 4-OH-tamoxifen by In-Cell Western Assay (ICW). MCF7 cells were seeded in a 384-well plate and linear-dilution compounds were administrated in duplicate for each treatment. After incubation for 24 h, Cells in the assay pate were treated as described in methods. ERα levels were quantified by immunofluorescence assay. 100% normalized to fulvestrant activity at 100 nM. Data are given as mean ± SEM. b The maximal ERα degradation across an ER-positive cell panel was evaluated in an In-cell Western assay. Cells were dispended into 96-well plates and incubated for 24 h with compounds (100 nM) treatment. Cellular ERα levels in each treated well were quantified by immunofluorescence assay.100% normalized to fulvestrant activity at 100 nM. c The comparison of cell viability between SCR-6852, AZD-9496, fulvestrant, or 4-OH-tamoxifen activity, respectively, across an ER-positive cell panel. Cells were seeded in 384-well plates and treated with linear-dilution compounds for 7 days of incubation. Cell viability was assessed using CellTiter-Glo. Cell growth inhibition is presented as a percentage of CellTiterGlo activity relative to the vehicle control. 100% normalized to maximal fulvestrant activity. d The comparison of cell viability between SCR-6852, fulvestrant or RAD-1901 activity, respectively, in MCF7 cells with ER WT, or mutant ESR1 Y537S, or mutant ESR1 D538G with the same operation as above. e–f, The evaluating of ERα level, as well as RB phosphorylation and cyclin D1 as ER targets by western blot, in MCF7 ER. WT (e) and ER. Y537S cells (f). Cells were seeded in 6-well plates and treated with linear-titration Fulvestrant or SCR-6852 for 5 days of incubation. Then cells in each well were collected and the targeting proteins in lysate supernatant were detected by WB