Abstract
Circulating tumour cells (CTCs) spread through the body and extravasation into distant organs, where tumour cells with stem signs are more likely to form metastasis.For successful metastases, tumour cells must penetrate the blood vessel wall.Epithelial-mesenchymal transition (EMT) is a critical to tumour cells intravasation. EMT enables them to acquire migratory and invasive potential. In this study, we assessed intravasation potential of tumour cells with different combinations in EMT and stem signs. Eleven patients with invasive breast carcinoma of no special type (age 49±11.2, T1-2N0-3M0, without neoadjuvant chemotherapy) were studied by multiplex immunofluorescence and flow cytometry for the detection tumour cells with stem and EMT features in primary tissue and blood. The following antibodies were used: multiplex IHC panel – Epcam (1:1000, Abcam, UK), CD45 (1:10, DAKO, USA), CD44 (1:100, Thermo, USA), CD24 (1:100, Thermo, USA), N-cadherin (1:500, Abcam, UK) and flow cytometry panel - BV650-anti-EpCam (Sony Biotechnology), BV570-anti-CD45 (Sony Biotechnology), BV510-anti-CD44 (BD Horizont), PerCP-Cy5.5-anti-CD24 (Sony Biotechnology), PE-Cy7-anti-N-cadherin (Sony Biotechnology). We evaluated frequency of tumour cells subtypes in primary tumour and blood simultaneously. Epcam+/CD45-/CD44+/CD24-/N-cadherin+ and Epcam+/CD45-/CD44+/CD24-/N-cadherin- cells were not detected simultaneously in primary tumour and blood (0/11). However, this cells in 23% (3/11) and 18% (2/11) cases were detected in primary tumour. Only in 18% (2/11) cases Epcam+/CD45-/CD44-/CD24-/N-cadherin- cells was detected simultaneously in primary tumour and blood. Epcam+/CD45-/CD44-/CD24-/N-cadherin+ cells in the highest simultaneous frequency in primary tumour and blood (45.45% (5/11)) was found. The high intravasation potency was found in non stem-like tumour cells with EMT sign. Future study of EMT in intravasation processes is key to prevention of distant metastases. Study was supported by RSF (#19-75-30016).
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