Abstract
Abstract Background: Breast cancer (BC) is the second leading cause of cancer related death in American women. There is a critical need for new biomarkers to evaluate treatment response and emerging mechanisms of resistance. The identification and characterization of new biomarkers such as Circulating Tumor Cells (CTCs) may be helpful tools for these purposes as well as understanding the biology of metastatic BC. Methods: Blood samples were processed with a Ficoll gradient to isolate nucleated cells. CTCs were isolated from these samples using an exclusion-based sample preparation technology known as the VERSA (Versatile Exclusion-based Rare Sample Analysis) platform. EpCAM positive CTCs were captured in the VERSA followed by mRNA extraction for gene expression profiling for the estrogen receptor (ER) and other breast cancer relevant targets. CTCs were defined as Hoechst positive, CD45/CD11b/CD14 negative, and cytokeratin positive. Captured CTCs were evaluated for ER protein staining intensity and localization. Results: The VERSA platform was validated to capture and enumerate CTCs from patients with breast cancer using EpCAM capture and immunofluorescent staining for intact nucleus, ER, and cytokeratin while negative for multiple immune markers (CD45, CD11b, CD14). The VERSA can capture cells with >90% capture efficiency with EpCAM antibody in MCF7 and T47D cell lines. ER specific transcriptome analysis in BC patient CTCs and cell lines was performed. We evaluated expression of ER signaling pathway, prognostic, metastatic/EMT; stemness; epithelial markers with concordant expression of RPLPO and HPRT1. We identified expression of ER in 6 out of 7 BC samples. Expression of PR and HER2 in a subset of samples confirms capture of breast cancer cells for further molecular interrogation. Conclusion: In conclusion, we report the expression profiles of ER signaling genes in EpCAM captured CTCs from BC patients. These tools will be used to compare subpopulations of CTCs from patients with BC as predictive and pharmacodynamic biomarkers of response to targeted therapies or chemotherapies. Citation Format: Ashok Singh, Jamie M. Sperger, Jennifer Schehr, Tessa Witkowski, Beth A. Weaver, Mark E. Burkard, Joshua M. Lang. Expression of estrogen receptor specific signaling transcriptome in epithelial cell adhesion molecule (EpCAM) captured circulating tumor cells from patients with breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3784. doi:10.1158/1538-7445.AM2017-3784
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