title: Genomic modelling of the ESR1 Y537S mutation for evaluating function and new therapeutic approaches for metastatic breast cancer.
abstract: Drugs that inhibit estrogen receptor-α (ER) activity have been highly successful in treating and reducing breast cancer progression in ER-positive disease. However, resistance to these therapies presents a major clinical problem. Recent genetic studies have shown that mutations in the ER gene are found in >20% of tumours that progress on endocrine therapies. Remarkably, the great majority of these mutations localize to just a few amino acids within or near the critical helix 12 region of the ER hormone binding domain, where they are likely to be single allele mutations. Understanding how these mutations impact on ER function is a prerequisite for identifying methods to treat breast cancer patients featuring such mutations. Towards this end, we used CRISPR-Cas9 genome editing to make a single allele knock-in of the most commonly mutated amino acid residue, tyrosine 537, in the estrogen-responsive MCF7 breast cancer cell line. Genomic analyses using RNA-seq and ER ChIP-seq demonstrated that the Y537S mutation promotes constitutive ER activity globally, resulting in estrogen-independent growth. MCF7-Y537S cells were resistant to the anti-estrogen tamoxifen and fulvestrant. Further, we show that the basal transcription factor TFIIH is constitutively recruited by ER-Y537S, resulting in ligand-independent phosphorylation of Serine 118 (Ser118) by the TFIIH kinase, cyclin-dependent kinase (CDK)7. The CDK7 inhibitor, THZ1 prevented Ser118 phosphorylation and inhibited growth of MCF7-Y537S cells. These studies confirm the functional importance of ER mutations in endocrine resistance, demonstrate the utility of knock-in mutational models for investigating alternative therapeutic approaches and highlight CDK7 inhibition as a potential therapy for endocrine-resistant breast cancer mediated by ER mutations.
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: A
authors.initials: A
authors.lastname: Harrod
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: J
authors.initials: J
authors.lastname: Fulton
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: V T M
authors.initials: VTM
authors.lastname: Nguyen
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: M
authors.initials: M
authors.lastname: Periyasamy
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: L
authors.initials: L
authors.lastname: Ramos-Garcia
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: C-F
authors.initials: CF
authors.lastname: Lai
authors.affiliation: School of Biological Sciences, University of Essex, Wivenhoe Park, Colchester, UK.
authors.forename: G
authors.initials: G
authors.lastname: Metodieva
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: A
authors.initials: A
authors.lastname: de Giorgio
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: R L
authors.initials: RL
authors.lastname: Williams
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: D B
authors.initials: DB
authors.lastname: Santos
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: P J
authors.initials: PJ
authors.lastname: Gomez
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: M-L
authors.initials: ML
authors.lastname: Lin
authors.affiliation: School of Biological Sciences, University of Essex, Wivenhoe Park, Colchester, UK.
authors.forename: M V
authors.initials: MV
authors.lastname: Metodiev
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: J
authors.initials: J
authors.lastname: Stebbing
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: L
authors.identifier: 0000-0002-3059-4829
authors.initials: L
authors.lastname: Castellano
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: L
authors.identifier: 0000-0002-7534-0785
authors.initials: L
authors.lastname: Magnani
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: R C
authors.initials: RC
authors.lastname: Coombes
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: L
authors.initials: L
authors.lastname: Buluwela
authors.affiliation: Department of Surgery & Cancer, Imperial College London, Hammersmith Hospital Campus, London, UK.
authors.forename: S
authors.identifier: 0000-0002-1320-0816
authors.initials: S
authors.lastname: Ali
mesh_terms: D000970:Antineoplastic Agents / Q000627:therapeutic use; D001943:Breast Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000473:pathology*; D064113:CRISPR-Cas Systems; D049109:Cell Proliferation / Q000187:drug effects; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors; D004965:Estrogen Antagonists / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000235:genetics*; D004967:Estrogens / Q000494:pharmacology; D005260:Female; D055879:Gene Knock-In Techniques; D006657:Histones / Q000378:metabolism; D006801:Humans; D061986:MCF-7 Cells; D009154:Mutation; D010766:Phosphorylation; D012694:Serine / Q000378:metabolism; D013629:Tamoxifen / Q000031:analogs & derivatives / Q000494:pharmacology; D000095043:Cyclin-Dependent Kinase-Activating Kinase
chemical_list: D000970:Antineoplastic Agents; C506487:ESR1 protein, human; D004965:Estrogen Antagonists; D047628:Estrogen Receptor alpha; D004967:Estrogens; D006657:Histones; D013629:Tamoxifen; C016601:afimoxifene; D012694:Serine; D018844:Cyclin-Dependent Kinases; D000095043:Cyclin-Dependent Kinase-Activating Kinase; C000722416:CDK7 protein, human