Abstract Approximately 50% of luminal breast cancer patients become resistant to first-line endocrine therapies after an average of 5 years. Acquired activating mutations to the ESR1 ligand binding domain enable hormone therapy resistance in approximately 25-40% of these patients. Competitive estrogen antagonists function by maintaining the receptor in a transcriptionally inactive state by forcing estrogen receptor alpha ligand binding domain (ERα LBD) helix 12 into the AF-2 cleft to prevent the docking of LXXLL-containing transcriptional coactivators. Multiple next-generation ERα antagonists that function as molecular degraders of the receptor have been described and shown activity in breast cancers harboring the activating ESR1 mutations. To understand whether ERα antagonism correlates with receptor stability, we examined a comprehensive panel of structurally diverse antagonists in live breast cancer cell assays using WT, Y537S, and D538G-mutant receptors. We found that potent antagonists are able to both stabilize and degrade the receptor and further demonstrate that the D538G mutation reduces turnover with ICI due to specific defects in receptor ubiquitination and SUMOylation. High resolution x-ray crystal structures of WT and Y537S ERα LBD in complex with stabilizing and degrading antiestrogens suggest that modulators that enforce the classical antagonist-like helix 12 conformation in the mutant receptor achieve the greatest activity in transcriptional assays, regardless of their effects on receptor stability Citation Format: David Hosfield, Nan-Sheng Li, Ross Han, Muriel Laine, Sandra Weber, Madaline Sauvage, Sylvie Madar, Geoffrey Greene, Sean Fanning. Effective ER antagonists resist conformational restrictions imposed by somatic mutation but do not correlate with effects on receptor stability in live cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2472.