Mutation site and context dependent effects of ESR1 mutation in genome-edited breast cancer cell models

Amir Bahreini, Zheqi Li, Peilu Wang, Kevin M. Levine, Nilgun Tasdemir, Lan Cao, Hazel M. Weir, Shannon L. Puhalla, Nancy E. Davidson, Andrew M. Stern, David Chu, Ben Ho Park, Adrian V. Lee, Steffi Oesterreich

Research output: Contribution to journalArticlepeer-review

Abstract

Background: Mutations in the estrogen receptor alpha (ERα) 1 gene (ESR1) are frequently detected in ER+ metastatic breast cancer, and there is increasing evidence that these mutations confer endocrine resistance in breast cancer patients with advanced disease. However, their functional role is not well-understood, at least in part due to a lack of ESR1 mutant models. Here, we describe the generation and characterization of genome-edited T47D and MCF7 breast cancer cell lines with the two most common ESR1 mutations, Y537S and D538G. Methods: Genome editing was performed using CRISPR and adeno-associated virus (AAV) technologies to knock-in ESR1 mutations into T47D and MCF7 cell lines, respectively. Various techniques were utilized to assess the activity of mutant ER, including transactivation, growth and chromatin-immunoprecipitation (ChIP) assays. The level of endocrine resistance was tested in mutant cells using a number of selective estrogen receptor modulators (SERMs) and degraders (SERDs). RNA sequencing (RNA-seq) was employed to study gene targets of mutant ER. Results: Cells with ESR1 mutations displayed ligand-independent ER activity, and were resistant to several SERMs and SERDs, with cell line and mutation-specific differences with respect to magnitude of effect. The SERD AZ9496 showed increased efficacy compared to other drugs tested. Wild-type and mutant cell co-cultures demonstrated a unique evolution of mutant cells under estrogen deprivation and tamoxifen treatment. Transcriptome analysis confirmed ligand-independent regulation of ERα target genes by mutant ERα, but also identified novel target genes, some of which are involved in metastasis-associated phenotypes. Despite significant overlap in the ligand-independent genes between Y537S and D538G, the number of mutant ERα-target genes shared between the two cell lines was limited, suggesting context-dependent activity of the mutant receptor. Some genes and phenotypes were unique to one mutation within a given cell line, suggesting a mutation-specific effect. Conclusions: Taken together, ESR1 mutations in genome-edited breast cancer cell lines confer ligand-independent growth and endocrine resistance. These biologically relevant models can be used for further mechanistic and translational studies, including context-specific and mutation site-specific analysis of the ESR1 mutations.

Original languageEnglish (US)
Article number60
JournalBreast Cancer Research
Volume19
Issue number1
DOIs
StatePublished - May 23 2017

Keywords

  • Endocrine resistance
  • ESR1 mutations
  • Genome-edited cells
  • Metastatic breast cancer
  • RNA-seq

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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