PMC8264184_01
Patient's timeline. At the top, we reported the milestones of her clinical course, while on the bottom the treatment regimens administered to the patient. PET-CT, positron emission tomography-computed tomography; AC, adryamicin-cyclophosphamide; DTX, docetaxel; PTX, paclitaxel; CIS, cisplatin; VRL, vinorelbine; CAP, capecitabine; ERI, eribulin; DXR, doxorubicin; EVE, everolimus; BEV, bevacizumab; CTC, circulating tumor cell.
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F1
Here we report the case of a 32-year-old patient (Figure 1) who presented, on December 20, 2018, during breastfeeding, a clinical onset of a right breast lump. To set up the auto-fluorescence signal detected in FITC channel, we used MCF7 cell line (CD45-) and leukocytes (CD45+) (Supplementary Figure 1).
Microscopic hematoxylin and eosin photograph of metaplastic breast carcinoma (MpBC) with coexisting spindled (green arrows) and oval (black arrows) cells. Scale bar: 50 mum.
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F2
The microscopic photograph (10 x magnification) of hematoxylin and eosin staining of the resected tumor is reported in Figure 2. CTCs with a round-shaped morphology (n = 184). CTC clusters (n = 5) were present as well (Supplementary Figure 2).
(A) DEPArray images of the most representative single circulating tumor cells (CTCs) of the patient based on their shape (round- and spindle-shaped). The DAPI channel was used for nuclear staining using Hoechst 33342, PE channel for epithelial tag [anti-EPCAM, anti-cytokeratins (CKs) and anti-E-cadherin antibodies], and APC channel for mesenchymal tag (anti-N-cadherin, anti-CD44v6, anti-ABCG2, and anti-CD133 antibodies). Anti-CD45 (FITC channel) was used for CTC negative selection (Supplementary Figure 1). Scale bar: 30 mum. (B) Profiling of the CTC ID: 793 reveals the presence of aberrant regions, consistent with tumor nature. Chromosomes (Chr) and number of copies are reported along the x- and y-axis, respectively. Black dots in the figure represent chromosome regions with a normal diploid copy number. Conversely, red dots and blue dots indicate, respectively, significant copy number gains (copies > 2) and losses (copies <2), called by Control-FREEC.
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F3
CTC negative selection. CTC identification and analysis were performed by DEPArray NxT platform (Figure 3A). Through this technique, we were able to unequivocally establish the tumor nature of the unconventional spindle-shaped CTC (ID: 793), since it was characterized by an aneuploid genome with an altered CNA profile (Figure 3B). Through intersection bioinformatic analysis, we observed that the greatest part of the entire genome was not comparable among the three samples, suggesting high heterogeneity levels.