Abstract P2-01-08: Esr1 Y537 mutations are associated with increased baseline circulating tumor cells enumeration for patients with estrogen receptor positive metastatic breast cancer
Authors: Zhang Qiang , Jiao Jianhua , D'Amico Paolo , Davis Andrew A. , Qin Weijun , Gerratana Lorenzo , Jacob Saya L. , Zhang Youbin , Donahue Jeannine , Qiang Wenan , Shah Ami N. , Flaum Lisa , Gradishar William , Platanias Leonidas C. , Cristofanilli Massimo 
Published: March 2022

Introduction: Estrogen Receptor (ER)-positive breast cancer is the most frequent breast cancer subtype. Endocrine therapy (ET) targeting the ER pathway including CDK4/6 inhibitors represents the main initial therapeutic approach. However, clinical resistance associated with progression of disease caused by ESR1 mutations is a recognized an important mechanism of ET resistance. ESR1 mutations, most often detected from liquid biopsies, have been consistently associated with a worse outcome and are being currently evaluated as a potential biomarker to guide therapeutic decisions. Here we reported a new finding on the association between ESR1 amino acid Y537 mutations in ctDNA and circulating tumor cells (CTC) in metastatic breast cancer (MBC). Methods: The study included 158 ER positive MBC patients enrolled under an IRB-approved trial (NU16B06) at Lurie Cancer Center, Northwestern University. The patients received systemic treatments in 2016-2020. Whole blood samples (7.5ml/each) were collected in EDTA tubes from patients who were longitudinally characterized for CTCs before therapy (baseline). CTCs enumeration were performed in FDA approved CELLTRACKS ANALYZERII® System (Menarini) by using CXC Kit contains antibodies targeting EpCAM antigen for capturing CTCs, anti-CK-PE which is specific for the intracellular protein cytokeratin in epithelial cells, DAPI for staining the cell nucleus, anti-CD45-APC is specific for leukocytes. The CTCs were classified based on morphology and correct phenotype as CK+, DAPI+ and CD45-. Plasma ctDNA was isolated using a Qiagen circulating nucleic acid kit, and then was analyzed using the Guardant360 next-generation sequencing (NGS)-based assay. In this study, only patients with ESR1 mutations were included for statistical analyses of correlation between the hotspot mutation with the endocrine therapy resistance by using Causal Inference approach. All statistical analyses were conducted Mann-Whitney U test by IBM SPSS version 23.0. Results: ESR1 mutations were detected in 40 out of 158 patients at baseline. ESR1 Y537(N/C/S) mutations were detected in 13 patients, among which there were 9 patients who only had the Y537(N/C/S) mutations and no other ESR1 hotspot mutations (Group 1) and there were 4 patients who had polyclonal ESR1 mutations. In the latter cohort, there were 17 ESR1 amino acid mutations detected in 31 patients (Group 2) including Q314, T347T, N359I, K362N, E380Q, V392I, G442R, F461I, S463P, S464, I487M, K520K, M528V, L536H, D538G, D545D and Q565. There were no any ESR1 mutations detected in the other 118 patients (Group 3). The CTC&gt;5/7.5mL (Stage IV aggressive) were found in 66.7%, 38.7% and 29.3% patients in Group 1, Group 2 and Group 3 respectively. The median total CTCs was 26.0/7.5mL in MBC patients with ESR1 Y537(N/C/S) mutations (Group 1). The median of total CTCs was 2.0/7.5mL in the MBC patients with polyclonal ESR1 mutations but not on Y537 (N/C/S) (Group 2), and it was 1.0/7.5mL in the group of metastatic breast cancer without ESR1 mutations (Group 3). The median total CTCs in Group 1 was significantly higher than Group 2 (Mann-Whitney U, P&lt;0.05) and Group 3 (P=0.009), which indicated that Y537(N/C/S) mutations were correlated with higher baseline CTCs. Conclusions: In this study, we demonstrated a correlation between presence of ESR1 Y537 mutations and increased CTCs (higher Stage IV aggressive cases) in patients with HR+ MBC, These preliminary data may suggest a critical role of ESR1 Y537 mutations in the metastatic process, also indicating that different ESR1 amino acids mutations may play different roles on disease progress. Larger validation studies are needed to confirm that evaluation of ESR1 Y537 variant together with CTCs numeration is an accurate tool to identify endocrine-refractory disease.

Citation Format: Qiang Zhang, Jianhua Jiao, Paolo D'Amico, Andrew A. Davis, Weijun Qin, Lorenzo Gerratana, Saya L. Jacob, Youbin Zhang, Jeannine Donahue, Wenan Qiang, Ami N. Shah, Lisa Flaum, William Gradishar, Leonidas C. Platanias, Massimo Cristofanilli. Esr1 Y537 mutations are associated with increased baseline circulating tumor cells enumeration for patients with estrogen receptor positive metastatic breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-01-08.