Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients

作者：Maryam B. Lustberg, Priya Balasubramanian, Brandon Miller, Alejandra Garcia-Villa, Clayton Deighan, Yongqi Wu, Sarah Carothers, Michael Berger, Bhuvaneswari Ramaswamy, Erin Macrae, Robert Wesolowski, Rachel M. Layman, Ewa Mrózek, Xueliang Pan, Thomas A. Summers, Charles L. Shapiro, Jeffrey J. Chalmers · 发表于：Breast Cancer Research · 年份：2014 · DOI：10.1186/bcr3622 · 被引用次数：123 · 研究领域：Cancer Cells and Metastasis、Microfluidic and Bio-sensing Technologies、3D Printing in Biomedical Research

INTRODUCTION: Circulating tumor cells (CTCs) are commonly isolated from the blood by targeting the epithelial cell adhesion molecule (EpCAM) through positive selection. However, EpCAM can be downregulated during metastatic progression, or it can be initially not present. We designed the present prospective trial to characterize CTCs as well as other circulating cell populations in blood samples from women with metastatic breast cancer without EpCAM-dependent enrichment and/or isolation technology. METHODS: A total of 32 patients with metastatic breast cancer were enrolled, and blood samples were processed using a previously described negative depletion immunomagnetic methodology. Samples from healthy volunteers were run as controls (n = 5). Multistep sequential labeling was performed to label and fix cell-surface markers followed by permeabilization for cytokeratins (CK) 8, 18 and 19. Multiparametric flow cytometry (FCM) analysis was conducted using a BD LSR II flow cytometer or a BD FACSAria II or FACSAria III cell sorter. Immunocytochemical staining on postenrichment specimens for DAPI, EpCAM, CD45, CK, epidermal growth factor receptor and vimentin was performed. Expression of these markers was visualized using confocal microscopy (CM). RESULTS: CD45-negative/CK-positive (CD45- CK+) populations with EpCAM + and EpCAM - expression were identified with both FCM and CM from the negatively enriched patient samples. In addition, EpCAM + and EpCAM - populations that were CK + and...