Characteristics of stem sings of EpCAM-negative and EpCAM-positive tumor cells in primary tumor and 2D and 3D cultures in breast cancer

One of the predictors of adverse prognosis in breast cancer is the overexpression of the EpCAM protein. However, a signifcant part of tumor cells have low or no EpCAM expression. The molecular features and the ability to grow in 2D and 3D cultures, indirectly refecting metastatic potential of tumor cells without EpCAM expression, have not been sufficiently studied. The aim of the study was to compare phenotypic variants of stem cells and EMT depending on the EpCAM expression in the primary tumor of breast cancer and 2D and 3D cultures. Material and Methods. The study included 7 patients with invasive breast carcinoma of no special type. Luminal A subtype was found in 2/7 patients (29 %), and luminal B HER2-negative molecular subtype was found in 5/7 patients (71 %). The patients didn’t have neoadjuvant chemotherapy. Cells from the primary tumor were cultured in 2D and 3D. The primary tumor cells, 2D and 3D cultures were phenotyped using fow cytometry with antibodies targeting: CD45, EpCAM (CD326), CD44, CD24, N-cadherin (CD325), and CD133, CK7/8, EpCAM (CD326). Results . No significant differences were found in the frequency of detection and the number of cells exhibiting stem cell features among EpCAM- and EpCAM+ tumor cells, 2D and 3D cultures. All phenotypic variants of co-expression of stemness markers CD44, CD24, CD133, and ALDH were present both in the primary tumor and 2D/3D cultures. Stem cells in mammospheres had features of both epithelial and hybrid EMT phenotypes. In primary tumors and 2D/3D cultures, the smallest proportion consisted of stem cells with the CD44+CD24- phenotype and cells co-expressing CD44+CD24- with other stemness markers, while the largest proportion comprised stem cells with ALDH+ and ALDH+CD133+ phenotypes. Marked intra- and inter-tumor heterogeneity in the phenotypic composition of primary tumors, 2D, and 3D cultures was noted. Conclusion . The results indicate that EpCAM-negative and EpCAM-positive tumor cells of luminal molecular subtype of breast cancer are capable of exhibiting various phenotypic variants of stemness and EMT in the primary tumor and 2D and 3D cultures. In most cases, individual tumor cells show co-expression of several stemness markers. The phenotypic composition of primary tumors, 2D and 3D cultures is characterized by pronounced intra- and inter-tumor heterogeneity.