Background: Endocrine therapy (ET) is an effective treatment of estrogen receptor positive (ER+) breast cancer (BC). However, not all ER+ cancers respond to ET and many eventually acquire resistance. Genomic aberrations in ESR1 have been reported to play a role in resistance to treatment. ESR1 mutations (ESRMs), reported in 10-50% of metastatic or recurrent BCs treated with aromatase inhibitors (AIs), can lead to constitutive activation and reduced sensitivity to ET. The incidence and clinical implications of ESR1 amplification (ESRA) is not well-established. A variety of structural rearrangements involving ESR1 have been reported in primary BCs, with some more strongly associated with tamoxifen and AI resistance. This study aimed to establish a rapid, reliable and cost-effective method to screen and monitor ESR1 genomic aberrations in clinical BC tissue samples and relate these to the 1st line and subsequent ETs patients received.


• Cohort A - 20 post-menopausal women (PMW) with ER+ BC who had acquired resistance to AIs and received subsequent lines of ET. Previous NGS data were available for these patients.

• Cohort (B) - 425 ER+ BC patients, with paired matched tissue samples from the primary and progressive/recurrent cancer on 1st line ET; sites included local recurrence (25%), nodal recurrence (29%), distant recurrence (3%) and primary progression on neoadjuvant ET (44%). Median follow-up 10 years. All patients received 2nd line ET, 14% developed a further recurrence on 2nd line ET.

Methods: ESRMs were assessed by allele-specific real-time quantitative (rt-qPCR) and digital droplet PCR (ddPCR) assays, a novel fluorometric in situ mutation detection (ISMD) approaches and AmpliSeq targeted sequencing. ESRA and ESR1 fusions were detected by targeted sequencing and validated using FISH and custom ligation assays for commonly-reported fusion proteins (currently ESR1-e6>YAP1 and ESR1-e6>PCDH11X), respectively.

Results: Results from ddPCR and ISMD were consistent with NGS findings in cohort A. There was expansion of D538G mutant clones with acquired resistance in 5/20 patients (25%). ESR1 copy number gain was seen in 11/20 patients (55%) in resistant samples. Gene amplification was confirmed by FISH in 6, corresponding to those with the highest gain from the NGS data. In cohort B, recurrent/resistant samples (including 2nd recurrences) with matched primaries are currently being screened for ESR1 genomic aberrations using all methods allowing for a comprehensive comparison. This will allow full characterisation of mutations, copy number changes and gene fusions in the largest cohort of ET resistant cancers to date. Results will be interpreted in the context of 1st line ET (51% Tamoxifen, 34% non-steroidal AI, 8% exemestane, 7% other ET) and 2nd line ET in the 14% of patients who developed a 2nd recurrence. ESR1 genomic aberrations have been identified in 38% of samples to date, with specific aberrations associated with particular ETs.


• A reliable, robust and cost-effective methodology for the detection and quantitation of ESR1 aberrations in clinical BC samples has been developed and compared with NGS and targeted sequencing approaches.

• This method would allow rapid screening for key aberrations with the potential to inform selection of 2nd line therapy.

• Multiplexing of fluorometric assays may enable in situ clonality analysis that allows visualisation of multiple genomic driver aberrations simultaneously.

• In the largest cohort of patients with resistance to ET to date, there is a high incidence of ESR1 genomic aberrations. These are associated with specific ETs. Analysis between these changes and response, disease-free and overall breast cancer-specific survival on 2nd line ET is currently ongoing.

Citation Format: Carlos Martinez-Perez, Charlene Kay, James Meehan, Mark Gray, Rebecca Swan, Lorna Renshaw, Jane Keys, Andrew H Sims, Olga Oikonomidou, J Michael Dixon, Arran K Turnbull. Assessment of ESR1 genomic aberrations and their role in endocrine therapy resistance in breast cancer [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P2-11-06.