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Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients
INTRODUCTION: Circulating tumor cells (CTCs) are commonly isolated from the blood by targeting the epithelial cell adhesion molecule (EpCAM) through positive selection. However, EpCAM can be downregulated during metastatic progression, or it can be initially not present. We designed the present pros...
Autores principales: | , , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4053256/ https://www.ncbi.nlm.nih.gov/pubmed/24602188 http://dx.doi.org/10.1186/bcr3622 |
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author | Lustberg, Maryam B Balasubramanian, Priya Miller, Brandon Garcia-Villa, Alejandra Deighan, Clayton Wu, Yongqi Carothers, Sarah Berger, Michael Ramaswamy, Bhuvaneswari Macrae, Erin R Wesolowski, Robert Layman, Rachel M Mrozek, Ewa Pan, Xueliang Summers, Thomas A Shapiro, Charles L Chalmers, Jeffrey J |
author_facet | Lustberg, Maryam B Balasubramanian, Priya Miller, Brandon Garcia-Villa, Alejandra Deighan, Clayton Wu, Yongqi Carothers, Sarah Berger, Michael Ramaswamy, Bhuvaneswari Macrae, Erin R Wesolowski, Robert Layman, Rachel M Mrozek, Ewa Pan, Xueliang Summers, Thomas A Shapiro, Charles L Chalmers, Jeffrey J |
author_sort | Lustberg, Maryam B |
collection | PubMed |
description | INTRODUCTION: Circulating tumor cells (CTCs) are commonly isolated from the blood by targeting the epithelial cell adhesion molecule (EpCAM) through positive selection. However, EpCAM can be downregulated during metastatic progression, or it can be initially not present. We designed the present prospective trial to characterize CTCs as well as other circulating cell populations in blood samples from women with metastatic breast cancer without EpCAM-dependent enrichment and/or isolation technology. METHODS: A total of 32 patients with metastatic breast cancer were enrolled, and blood samples were processed using a previously described negative depletion immunomagnetic methodology. Samples from healthy volunteers were run as controls (n = 5). Multistep sequential labeling was performed to label and fix cell-surface markers followed by permeabilization for cytokeratins (CK) 8, 18 and 19. Multiparametric flow cytometry (FCM) analysis was conducted using a BD LSR II flow cytometer or a BD FACSAria II or FACSAria III cell sorter. Immunocytochemical staining on postenrichment specimens for DAPI, EpCAM, CD45, CK, epidermal growth factor receptor and vimentin was performed. Expression of these markers was visualized using confocal microscopy (CM). RESULTS: CD45-negative/CK-positive (CD45− CK+) populations with EpCAM + and EpCAM − expression were identified with both FCM and CM from the negatively enriched patient samples. In addition, EpCAM + and EpCAM − populations that were CK + and coexpressing the pan-hematopoietic marker CD45 were also noted. There were more CK + EpCAM − events/ml than CK + EpCAM + events/ml in both the CD45− and CD45+ fractions (both statistically significant at P ≤ 0.0005). The number of CK + CD45− and CK + CD45+ events per milliliter in blood samples (regardless of EpCAM status) was higher in patient samples than in normal control samples (P ≤ 0.0005 and P ≤ 0.026, respectively). Further, a significant fraction of the CK + CD45+ events also expressed CD68, a marker associated with tumor-associated macrophages. Higher levels of CD45-CK + EpCAM − were associated with worse overall survival (P = 0.0292). CONCLUSIONS: Metastatic breast cancer patients have atypical cells that are CK + EpCAM − circulating in their blood. Because a substantial number of these patients do not have EpCAM + CTCs, additional studies are needed to evaluate the role of EpCAM − circulating cells as a prognostic and predictive marker. |
format | Online Article Text |
id | pubmed-4053256 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-40532562014-06-12 Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients Lustberg, Maryam B Balasubramanian, Priya Miller, Brandon Garcia-Villa, Alejandra Deighan, Clayton Wu, Yongqi Carothers, Sarah Berger, Michael Ramaswamy, Bhuvaneswari Macrae, Erin R Wesolowski, Robert Layman, Rachel M Mrozek, Ewa Pan, Xueliang Summers, Thomas A Shapiro, Charles L Chalmers, Jeffrey J Breast Cancer Res Research Article INTRODUCTION: Circulating tumor cells (CTCs) are commonly isolated from the blood by targeting the epithelial cell adhesion molecule (EpCAM) through positive selection. However, EpCAM can be downregulated during metastatic progression, or it can be initially not present. We designed the present prospective trial to characterize CTCs as well as other circulating cell populations in blood samples from women with metastatic breast cancer without EpCAM-dependent enrichment and/or isolation technology. METHODS: A total of 32 patients with metastatic breast cancer were enrolled, and blood samples were processed using a previously described negative depletion immunomagnetic methodology. Samples from healthy volunteers were run as controls (n = 5). Multistep sequential labeling was performed to label and fix cell-surface markers followed by permeabilization for cytokeratins (CK) 8, 18 and 19. Multiparametric flow cytometry (FCM) analysis was conducted using a BD LSR II flow cytometer or a BD FACSAria II or FACSAria III cell sorter. Immunocytochemical staining on postenrichment specimens for DAPI, EpCAM, CD45, CK, epidermal growth factor receptor and vimentin was performed. Expression of these markers was visualized using confocal microscopy (CM). RESULTS: CD45-negative/CK-positive (CD45− CK+) populations with EpCAM + and EpCAM − expression were identified with both FCM and CM from the negatively enriched patient samples. In addition, EpCAM + and EpCAM − populations that were CK + and coexpressing the pan-hematopoietic marker CD45 were also noted. There were more CK + EpCAM − events/ml than CK + EpCAM + events/ml in both the CD45− and CD45+ fractions (both statistically significant at P ≤ 0.0005). The number of CK + CD45− and CK + CD45+ events per milliliter in blood samples (regardless of EpCAM status) was higher in patient samples than in normal control samples (P ≤ 0.0005 and P ≤ 0.026, respectively). Further, a significant fraction of the CK + CD45+ events also expressed CD68, a marker associated with tumor-associated macrophages. Higher levels of CD45-CK + EpCAM − were associated with worse overall survival (P = 0.0292). CONCLUSIONS: Metastatic breast cancer patients have atypical cells that are CK + EpCAM − circulating in their blood. Because a substantial number of these patients do not have EpCAM + CTCs, additional studies are needed to evaluate the role of EpCAM − circulating cells as a prognostic and predictive marker. BioMed Central 2014 2014-03-06 /pmc/articles/PMC4053256/ /pubmed/24602188 http://dx.doi.org/10.1186/bcr3622 Text en Copyright © 2014 Lustberg et al.; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. |
spellingShingle | Research Article Lustberg, Maryam B Balasubramanian, Priya Miller, Brandon Garcia-Villa, Alejandra Deighan, Clayton Wu, Yongqi Carothers, Sarah Berger, Michael Ramaswamy, Bhuvaneswari Macrae, Erin R Wesolowski, Robert Layman, Rachel M Mrozek, Ewa Pan, Xueliang Summers, Thomas A Shapiro, Charles L Chalmers, Jeffrey J Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
title | Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
title_full | Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
title_fullStr | Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
title_full_unstemmed | Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
title_short | Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
title_sort | heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4053256/ https://www.ncbi.nlm.nih.gov/pubmed/24602188 http://dx.doi.org/10.1186/bcr3622 |
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