Abstract
Circulating tumor cells (CTCs) are considered as surrogate markers for prognosticating and evaluating patient treatment responses. Here, 226 blood samples from 92 patients with breast cancer, including patients with newly diagnosed or metastatic refractory cancer, and 16 blood samples from healthy subjects were cultured in laser-ablated microwells. Clusters containing an increasing number of cytokeratin-positive (CK+) cells appeared after 2 weeks, while most blood cells disappeared with time. Cultures were heterogeneous and exhibited two distinct sub-populations of cells: 'Small' (≤ 25 μm; high nuclear/cytoplasmic ratio; CD45-) cells, comprising CTCs, and 'Large' (> 25 μm; low nuclear/cytoplasmic ratio; CD68+ or CD56+) cells, corresponding to macrophage and natural killer-like cells. The Small cell fraction also showed copy number increases in six target genes (FGFR1, Myc, CCND1, HER2, TOP2A and ZNF217) associated with breast cancer. These expanded CTCs exhibited different proportions of epithelial-mesenchymal phenotypes and were transferable for further expansion as spheroids in serum-free suspension or 3D cultures. Cluster formation was affected by the presence and duration of systemic therapy, and its persistence may reflect therapeutic resistance. This novel and advanced method estimates CTC clonal heterogeneity and can predict, within a relatively short time frame, patient responses to therapy.
Highlights
Circulating tumor cells (CTCs), derived from either primary or metastatic tumors, have raised considerable interest within the translational oncology community [1, 2]
A laser-ablated, microwell-based assay (Figure 1A– 1D) for CTC culture was established with nucleated cells from patient blood samples (Supplementary Tables 1A–D) after red blood cell (RBC) lysis (Figure 1D)
The macrophage-like behavior of these cells was confirmed with 1-μm fluorescein-labeled polystyrene microbeads that were phagocytosed within a 24-h time frame (Supplementary Figure 4D)
Summary
Circulating tumor cells (CTCs), derived from either primary or metastatic tumors, have raised considerable interest within the translational oncology community [1, 2]. CTCs appear during the early stages of tumor progression [3, 4] as single cells or cell clusters and exhibit a partial or complete epithelial–mesenchymal transitioned (‘EMTed’) phenotype [5, 6]. These cells may later colonize distant organs and develop into clinically detectable metastases. There is an urgent need for techniques that can successfully expand CTCs. Recent studies have reported the establishment of cell lines derived from CTCs of breast [12, 13], colon [14] and prostate [15] cancer patients, obtained following pre-enrichment with affinity binding [12], fluorescenceactivated cell sorting (FACS) [13], or negative selection [15]. There is still a need for a method with improved efficiency and without the need for prior enrichment for practical use in the clinic
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
