title: Classification of anti-estrogens according to intramolecular FRET effects on phospho-mutants of estrogen receptor alpha.
abstract: Anti-estrogen resistance is a major clinical problem in the treatment of breast cancer. In this study, fluorescence resonance energy transfer (FRET) analysis, a rapid and direct way to monitor conformational changes of estrogen receptor alpha (ERalpha) upon anti-estrogen binding, was used to characterize resistance to anti-estrogens. Nine different anti-estrogens all induced a rapid FRET response within minutes after the compounds have liganded to ERalpha in live cells, corresponding to an inactive conformation of the ERalpha. Phosphorylation of Ser(305) and/or Ser(236) of ERalpha by protein kinase A (PKA) and of Ser(118) by mitogen-activated protein kinase (MAPK) influenced the FRET response differently for the various anti-estrogens. PKA and MAPK are both associated with resistance to anti-estrogens in breast cancer patients. Their respective actions can result in seven different combinations of phospho-modifications in ERalpha where the FRET effects of particular anti-estrogen(s) are nullified. The FRET response provided information on the activity of ERalpha under the various anti-estrogen conditions as measured in a traditional reporter assay. Tamoxifen and EM-652 were the most sensitive to kinase activities, whereas ICI-182,780 (Fulvestrant) and ICI-164,384 were the most stringent. The different responses of anti-estrogens to the various combinations of phospho-modifications in ERalpha elucidate why certain anti-estrogens are more prone than others to develop resistance. These data provide new insights into the mechanism of action of anti-hormones and are critical for selection of the correct individual patient-based endocrine therapy in breast cancer.
authors.affiliation: Division of Tumor Biology, the Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, the Netherlands. r.michalides@nki.nl.
authors.forename: Wilbert
authors.initials: W
authors.lastname: Zwart
authors.forename: Alexander
authors.initials: A
authors.lastname: Griekspoor
authors.forename: Mariska
authors.initials: M
authors.lastname: Rondaij
authors.forename: Desiree
authors.initials: D
authors.lastname: Verwoerd
authors.forename: Jacques
authors.initials: J
authors.lastname: Neefjes
authors.forename: Rob
authors.initials: R
authors.lastname: Michalides
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D004958:Estradiol / Q000031:analogs & derivatives / Q000494:pharmacology; D004965:Estrogen Antagonists / Q000737:chemistry / Q000494:pharmacology*; D047628:Estrogen Receptor alpha / Q000235:genetics* / Q000502:physiology*; D004967:Estrogens / Q000737:chemistry*; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D000077267:Fulvestrant; D006801:Humans; D008164:Luminescent Proteins / Q000378:metabolism; D008954:Models, Biological; D008956:Models, Chemical; D009154:Mutation; D010880:Piperidines / Q000494:pharmacology; D013629:Tamoxifen / Q000494:pharmacology; D015533:Transcriptional Activation
chemical_list: D001426:Bacterial Proteins; D004965:Estrogen Antagonists; D047628:Estrogen Receptor alpha; D004967:Estrogens; D008164:Luminescent Proteins; D010880:Piperidines; C054562:yellow fluorescent protein, Bacteria; D013629:Tamoxifen; D000077267:Fulvestrant; C012121:ritetronium; D004958:Estradiol