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(1) Background: The aim of the study was to evaluate the prognostic significance of EMT-associated ( TWIST1) and stem-cell (SC) transcript ( CD24 , CD44 , ALDH1 ) quantification in EpCAM+ circulating tumor cells (CTCs) of early breast cancer patients. (2) Methods: 100 early stage breast cancer patients and 19 healthy donors were enrolled in the study. CD24 , CD44 , and ALDH1 transcripts of EpCAM + cells were quantified using a novel highly sensitive and specific quadraplex RT-qPCR, while TWIST1 transcripts were quantified by single RT-qPCR. All patients were followed up for more than 5 years. (3) Results: A significant positive correlation between overexpression of TWIST1 and CD24 -/low /CD44 high  profile was found. Kaplan-Meier analysis revealed that the ER/PR-negative (HR-) patients and those patients with more than 3 positive lymph nodes that overexpressed TWIST1 in EpCAM + cells had a significant lower DFI (log rank test; p &lt; 0.001, p &lt; 0.001) and OS (log rank test; p = 0.006, p &lt; 0.001). Univariate and multivariate analysis also revealed the prognostic value of TWIST1 overexpression and CD24 -/low /CD44 high  and CD24 -/low /ALDH1 high  profile for both DFI and OS. (4) Conclusions: Detection of TWIST1 overexpression and stem-cell ( CD24, CD44, ALDH1 ) transcripts in EpCAM + CTCs provides prognostic information in early stage breast cancer patients.

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Prognostic Significance of TWIST1, CD24, CD44, and ALDH1 Transcript Quantification in EpCAM-Positive Circulating Tumor Cells from Early Stage Breast Cancer Patients