EpCAM-independent isolation of circulating tumor cells with epithelial-to-mesenchymal transition and cancer stem cell phenotypes using ApoStream® in patients with breast cancer treated with primary systemic therapy.

Fanny Le Du,James M Reuben,Minjeong Park,Darren W Davis,Vicente Valero,Naoto T Ueno,Diane D Liu,Carlos H Barcenas,Kumiko Kida,Takeo Fujii,Debu Tripathy,Weiguo Wu,Mariana Chavez-Macgregor,William B Coleman

doi:10.1371/journal.pone.0229903

Abstract

BackgroundTumor cells with a mesenchymal phenotype and/or cancer stem-like cells (CSCs) are known to contribute to metastasis and drug resistance. Circulating tumor cells (CTCs) undergoing epithelial-mesenchymal transition (EMT) and CTCs reflecting a dedifferentiated CSC phenotype may not be detected using only an anti-EpCAM antibody to capture them. We used an antibody-independent CTC enrichment platform, ApoStream®, which does not rely on any antibody, including anti-EpCAM, to capture EMT- and CSC-CTCs in breast cancer patients who received neoadjuvant chemotherapy and correlated them to pathological complete response (pCR).MethodsBlood samples from newly diagnosed breast cancer patients were prospectively collected before neoadjuvant chemotherapy (T0), after chemotherapy but before surgery (T1), and after surgery (T2) and processed using ApoStream. CTCs detected were stained with additional markers to define 3 CTC subsets with the following phenotypes: epithelial CTCs (CK+, EpCAM+ or E-cadherin+), EMT-CTCs (β-catenin+ or vimentin+), and CSC-CTCs (CD44+ and CD24low).ResultsWe enrolled 55 patients, 47 of which had data for analysis. EMT-CTCs were detected in 57%, 62%, and 72% and CSC-CTCs in 9%, 22%, and 19% at the T0, T1, and T2 time points, respectively. Counts of epithelial (P = 0.225) and EMT (P = 0.522) phenotypes of CTCs at T0 did not significantly predict pCR. Moreover, no correlation between CTC count change and pCR was demonstrated.ConclusionsApoStream was successful in detecting EMT-CTCs among patients after neoadjuvant chemotherapy. However, EMT-/CSC-CTC counts did not correlate with pCR. Due to the small sample size and heterogeneity of this patient population, further study in a larger cohort of molecularly homogeneous patients is warranted.

Highlights

Circulating tumor cells (CTCs) are found in human blood; their detection might be used as a marker of early relapse [1,2]
Isolation of CTCs with epithelial-mesenchymal transition (EMT) and cancer stem-like cells (CSCs) phenotypes using ApoStream® in breast cancer patients treated with NACT
ApoStream was successful in detecting EMT-CTCs among patients after neoadjuvant chemotherapy

Summary

Introduction

Circulating tumor cells (CTCs) are found in human blood; their detection might be used as a marker of early relapse [1,2]. The only U.S Food and Drug Administration (FDA)-approved platform for CTC detection is the CellSearch system, which targets expression of the cell-surface epithelial cell adhesion molecule (EpCAM) for CTC enrichment. This technique may not capture other CTC subsets in which epithelial markers are downregulated. Circulating tumor cells (CTCs) undergoing epithelial-mesenchymal transition (EMT) and CTCs reflecting a dedifferentiated CSC phenotype may not be detected using only an anti-EpCAM antibody to capture them. We used an antibody-independent CTC enrichment platform, ApoStream®, which does not rely on any antibody, including anti-EpCAM, to capture EMT- and CSC-CTCs in breast cancer patients who received neoadjuvant chemotherapy and correlated them to pathological complete response (pCR)

Objectives

Methods

Conclusion