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Estrogen receptor alpha somatic mutations Y537S and D538G confer breast cancer endocrine resistance by stabilizing the activating function-2 binding conformation.

ABSTRACT: Somatic mutations in the estrogen receptor alpha (ER?) gene (ESR1), especially Y537S and D538G, have been linked to acquired resistance to endocrine therapies. Cell-based studies demonstrated that these mutants confer ER? constitutive activity and antiestrogen resistance and suggest that ligand-binding domain dysfunction leads to endocrine therapy resistance. Here, we integrate biophysical and structural biology data to reveal how these mutations lead to a constitutively active and antiestrogen-resistant ER?. We show that these mutant ERs recruit coactivator in the absence of hormone while their affinities for estrogen agonist (estradiol) and antagonist (4-hydroxytamoxifen) are reduced. Further, they confer antiestrogen resistance by altering the conformational dynamics of the loop connecting Helix 11 and Helix 12 in the ligand-binding domain of ER?, which leads to a stabilized agonist state and an altered antagonist state that resists inhibition.

SUBMITTER: Fanning SW

PROVIDER: S-EPMC4821807 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Estrogen receptor alpha somatic mutations Y537S and D538G confer breast cancer endocrine resistance by stabilizing the activating function-2 binding conformation.

Fanning Sean W SW Mayne Christopher G CG Dharmarajan Venkatasubramanian V Carlson Kathryn E KE Martin Teresa A TA Novick Scott J SJ Toy Weiyi W Green Bradley B Panchamukhi Srinivas S Katzenellenbogen Benita S BS Tajkhorshid Emad E Griffin Patrick R PR Shen Yang Y Chandarlapaty Sarat S Katzenellenbogen John A JA Greene Geoffrey L GL

eLife 20160202

Somatic mutations in the estrogen receptor alpha (ERα) gene (ESR1), especially Y537S and D538G, have been linked to acquired resistance to endocrine therapies. Cell-based studies demonstrated that these mutants confer ERα constitutive activity and antiestrogen resistance and suggest that ligand-binding domain dysfunction leads to endocrine therapy resistance. Here, we integrate biophysical and structural biology data to reveal how these mutations lead to a constitutively active and antiestrogen- ...[more]

PMID: 26836308

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Project description:The purpose of this study was to determine the effect of estrogen receptor-α gene (ESR1) mutations at the tyrosine (Y) 537 amino acid residue within the ligand binding domain on 18F-fluoroestradiol (18F-FES) binding and in vivo tumor uptake compared with wild-type (WT)-estrogen receptor α (ER). Methods: ER-negative MDA-MB-231 breast cancer cells were used to generate stable cell lines that express WT-ER, Y537S, or Y537C mutant ER. Receptor expression and localization were confirmed by Western blot and immunofluorescence, respectively. ER transcriptional function was measured using an estrogen response element-luciferase reporter gene assay and quantitative polymerase chain reaction analysis of ER-regulated endogenous target genes. Saturation binding and competition assays were performed to determine equilibrium dissociation constant (Kd) and half maximal inhibitory concentration (IC50) values. 18F-FES uptake was measured in tumor xenografts grown in female athymic nude mice by small-animal PET/CT imaging and tissue biodistribution using 5.55 MBq (150 μCi) of 18F-FES. A 10-fold-lower injected dose of 0.555 MBq (15 μCi) of 18F-FES was also used for tissue biodistribution. Statistical significance was determined using ANOVA. Results: Y537S and Y537C mutations resulted in increased ER transcriptional activity in the absence of estrogen compared with WT-ER (11.48 ± 2.42 fold; P = 0.0002, and 5.89 ± 0.94 fold; P = 0.04, respectively). Constitutive ER activation of two target genes (PGR and TFF1) in the absence of estrogen was also observed in Y537S- and Y537C-ER cells compared with WT-ER. Kd values for 18F-FES were 0.98 ± 0.54 nM for Y537S-ER (P = 0.27) and 0.24 ± 0.03 nM for Y537C-ER (P = 0.95) compared with 0.07 ± 0.03 nM for WT-ER. IC50 values were 0.22 ± 0.09 nM for Y537S-ER (P = 0.97), 0.18 ± 0.09 nM for Y537C-ER (P = 0.99), and 0.19 ± 0.11 nM for WT-ER. Tumor xenografts expressing Y537S-ER (mean percentage injected dose per gram, 1.45 ± 0.06; P = 0.77) and Y537C-ER (2.09 ± 0.20; P = 0.21) had similar 18F-FES uptake compared with WT-ER (1.68 ± 0.12). Comparable 18F-FES uptake between Y537S-, Y537C-, and WT-ER xenografts was also observed using a 10-fold-lower injected dose with the tissue biodistribution assay. Conclusion: Since tumoral uptake of 18F-FES is not significantly impacted by Y537S-ER or Y537C-ER mutations, the potential diagnostic utility of 18F-FES PET imaging is expected to be equally valid for patients with or without these activating ESR1 mutations.

Dataset Information

Estrogen receptor alpha somatic mutations Y537S and D538G confer breast cancer endocrine resistance by stabilizing the activating function-2 binding conformation.

Publications

Estrogen receptor alpha somatic mutations Y537S and D538G confer breast cancer endocrine resistance by stabilizing the activating function-2 binding conformation.

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