Special Issue "Circulating Tumor Cells (CTCs)"

A special issue of Cancers (ISSN 2072-6694).

Deadline for manuscript submissions: 30 November 2018

Special Issue Editor

Guest Editor
Prof. Dr. Dario Marchetti

Director, Biomarker Research Program, Houston Methodist Research Institute, Professor, Institute of Academic Medicine, Visiting Professor, Baylor College of Medicine, MS: R7-414, 6670 Bertner Avenue, Houston, TX 77030, USA
Website | E-Mail
Interests: the biology and therapeutic utility of circulating tumor cells (CTCs); liquid biopsies; mechanisms of brain metastasis and dormancy in breast and melanoma cancers; roles of heparanase in development and disease

Special Issue Information

Dear Colleagues,

The major cause of patient mortality in cancer is metastasis; however, understanding of the metastatic cascade and mechanisms underlying this complex set of events remain poorly understood. Cancer cells invade the surrounding tissue of primary or metastatic tumors, intravasate into lymphatic and blood vessels, disseminate to distant tissues, extravasate by adapting to the new microenvironment, and colonize these tissues. Because dissemination mostly occurs through the blood, circulating tumor cells (CTCs) are of paramount importance and one of the most promising areas of oncology research. CTCs might serve as “liquid biopsy” as a better way to diagnose cancer patients compared to painful biopsies of the primary tumor, and can represent a clinically useful tool to better reflect cancer progression and monitoring in the patient. The isolation, characterization, and interrogation of CTCs are hampered by their rarity and heterogeneity, either inherited from respective primary or metastatic tumors or as a result of CTC evolution in blood by genetic/epigenetic progression that may or may not lead to a fully metastatic-competent CTC. Significant technical challenges in the field also persist in regard to identifying and interrogating CTC heterogeneity, assessing CTC biomarkers of clinical utility, and comprehensively capturing these cells (usually, a ratio of one CTC per 106 leukocytes and 109 erythrocytes in one milliliter of blood). Finally, many studies have reported the clinical impact of enumerating CTCs, considering that CTC testing is being employed in over 300 clinical trials worldwide. However, much of the CTC biology needs to be discovered and many scientific/technical challenges must be overcome before their clinical promise as biomarkers and targets for improved therapies can be fulfilled. The objective of this Special Issue is to publish the latest findings in CTC research and clinical implementation. Contributions outlining CTC discoveries in biological and clinical settings, CTC theoretical and pre-clinical models, CTC technologies and methods for their detection, and clinical findings applying CTC concepts, are welcome.

Prof. Dr. Dario Marchetti
Guest Editor

Manuscript Submission Information

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Published Papers (4 papers)

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Research

Open AccessArticle Scanning Electron Microscopy of Circulating Tumor Cells and Tumor-Derived Extracellular Vesicles
Cancers 2018, 10(11), 416; https://doi.org/10.3390/cancers10110416
Received: 13 August 2018 / Revised: 24 October 2018 / Accepted: 30 October 2018 / Published: 31 October 2018
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Abstract
To explore morphological features of circulating tumor cells (CTCs) and tumor-derived extracellular vesicles (tdEVs), we developed a protocol for scanning electron microscopy (SEM) of CTCs and tdEVs. CTCs and tdEVs were isolated by immunomagnetic enrichment based on their Epithelial Cell Adhesion Molecule (EpCAM)
[...] Read more.
To explore morphological features of circulating tumor cells (CTCs) and tumor-derived extracellular vesicles (tdEVs), we developed a protocol for scanning electron microscopy (SEM) of CTCs and tdEVs. CTCs and tdEVs were isolated by immunomagnetic enrichment based on their Epithelial Cell Adhesion Molecule (EpCAM) expression or by physical separation through 5 μm microsieves from 7.5 mL of blood from Castration-Resistant Prostate Cancer (CRPC) patients. Protocols were optimized using blood samples of healthy donors spiked with PC3 and LNCaP cell lines. CTCs and tdEVs were identified among the enriched cells by fluorescence microscopy. The positions of DNA+, CK+, CD45− CTCs and DNA−, CK+, CD45− tdEVs on the CellSearch cartridges and microsieves were recorded. After gradual dehydration and chemical drying, the regions of interest were imaged by SEM. CellSearch CTCs retained their morphology revealing various shapes, some of which were clearly associated with CTCs undergoing apoptosis. The ferrofluid was clearly distinguishable, shielding major portions of all isolated objects. CTCs and leukocytes on microsieves were clearly visible, but revealed physical damage attributed to the physical forces that cells exhibit while entering one or multiple pores. tdEVs could not be identified on the microsieves as they passed through the pores. Insights on the underlying mechanism of each isolation technique could be obtained. Complete detailed morphological characteristics of CTCs are, however, masked by both techniques. Full article
(This article belongs to the Special Issue Circulating Tumor Cells (CTCs))
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Figure 1

Open AccessArticle The Value of Monitoring the Behavior of Circulating Tumor Cells at the End of Endocrine Therapy in Breast Cancer Patients
Cancers 2018, 10(11), 407; https://doi.org/10.3390/cancers10110407
Received: 9 October 2018 / Accepted: 26 October 2018 / Published: 29 October 2018
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Abstract
After five years of endocrine therapy, patients with ER+ (estrogen receptor positive) breast cancer face the question of the benefit of further treatment. Ten years of endocrine therapy has been demonstrated to improve survival compared to five years. However, the individual benefit of
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After five years of endocrine therapy, patients with ER+ (estrogen receptor positive) breast cancer face the question of the benefit of further treatment. Ten years of endocrine therapy has been demonstrated to improve survival compared to five years. However, the individual benefit of continuation remains unclear. Therefore, markers for predicting benefit from endocrine treatment and extended endocrine treatment are desperately needed. In this study the dynamics over time of the tumor cells circulating in peripheral blood of patients, circulating tumor cells/ circulating epithelial tumor cells (CTC/CETC), as the systemic part of the tumor were investigated in 36 patients with ER+ primary breast cancer. CTC/CETCs were monitored serially during and after endocrine therapy. After termination of endocrine therapy 12 patients showed an increase in CTC/CETCs, with 8 of 12 suffering relapse. No change or a reduction was observed in 24 patients, with 2 of 24 suffering relapse. Initial tumor size was marginally prognostic (p = 0.053) but not nodal status nor the mere number of CTC/CETCs. Only the trajectory of CTC/CETCs was a statistically significant predictor of relapse free survival (increasing cell numbers: mean = 940 days vs. stable/decreasing cell numbers mean not reached). Individual cases demonstrated that an increase of CTC/CETCs after discontinuation of tamoxifen therapy could be stopped by resuming the endocrine therapy. Full article
(This article belongs to the Special Issue Circulating Tumor Cells (CTCs))
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Figure 1

Open AccessArticle Classification of Cells in CTC-Enriched Samples by Advanced Image Analysis
Cancers 2018, 10(10), 377; https://doi.org/10.3390/cancers10100377
Received: 21 August 2018 / Revised: 5 October 2018 / Accepted: 8 October 2018 / Published: 10 October 2018
PDF Full-text (2657 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
In the CellSearch® system, blood is immunomagnetically enriched for epithelial cell adhesion molecule (EpCAM) expression and cells are stained with the nucleic acid dye 4′6-diamidino-2-phenylindole (DAPI), Cytokeratin-PE (CK), and CD45-APC. Only DAPI+/CK+ objects are presented to the operator to identify circulating tumor
[...] Read more.
In the CellSearch® system, blood is immunomagnetically enriched for epithelial cell adhesion molecule (EpCAM) expression and cells are stained with the nucleic acid dye 4′6-diamidino-2-phenylindole (DAPI), Cytokeratin-PE (CK), and CD45-APC. Only DAPI+/CK+ objects are presented to the operator to identify circulating tumor cells (CTC) and the identity of all other cells and potential undetected CTC remains unrevealed. Here, we used the open source imaging program Automatic CTC Classification, Enumeration and PhenoTyping (ACCEPT) to analyze all DAPI+ nuclei in EpCAM-enriched blood samples obtained from 192 metastatic non-small cell lung cancer (NSCLC) patients and 162 controls. Significantly larger numbers of nuclei were detected in 300 patient samples with an average and standard deviation of 73,570 ± 74,948, as compared to 359 control samples with an average and standard deviation of 4191 ± 4463 (p < 0.001). In patients, only 18% ± 21% and in controls 23% ± 15% of the nuclei were identified as leukocytes or CTC. Adding CD16-PerCP for granulocyte staining, the use of an LED as the light source for CD45-APC excitation and plasma membrane staining obtained with wheat germ agglutinin significantly improved the classification of EpCAM-enriched cells, resulting in the identification of 94% ± 5% of the cells. However, especially in patients, the origin of the unidentified cells remains unknown. Further studies are needed to determine if undetected EpCAM+/DAPI+/CK-/CD45- CTC is present among these cells. Full article
(This article belongs to the Special Issue Circulating Tumor Cells (CTCs))
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Figure 1

Open AccessArticle Assessment of the Mutational Status of NSCLC Using Hypermetabolic Circulating Tumor Cells
Cancers 2018, 10(8), 270; https://doi.org/10.3390/cancers10080270
Received: 28 May 2018 / Revised: 1 August 2018 / Accepted: 10 August 2018 / Published: 14 August 2018
PDF Full-text (3225 KB) | HTML Full-text | XML Full-text | Supplementary Files
Abstract
Molecular characterization is currently a key step in NSCLC therapy selection. Circulating tumor cells (CTC) are excellent candidates for downstream analysis, but technology is still lagging behind. In this work, we show that the mutational status of NSCLC can be assessed on hypermetabolic
[...] Read more.
Molecular characterization is currently a key step in NSCLC therapy selection. Circulating tumor cells (CTC) are excellent candidates for downstream analysis, but technology is still lagging behind. In this work, we show that the mutational status of NSCLC can be assessed on hypermetabolic CTC, detected by their increased glucose uptake. We validated the method in 30 Stage IV NSCLC patients: peripheral blood samples were incubated with a fluorescent glucose analog (2-NBDG) and analyzed by flow cytometry. Cells with the highest glucose uptake were sorted out. EGFR and KRAS mutations were detected by ddPCR. In sorted cells, mutated DNA was found in 85% of patients, finding an exact match with primary tumor in 70% of cases. Interestingly, in two patients multiple KRAS mutations were detected. Two patients displayed different mutations with respect to the primary tumor, and in two out of the four patients with a wild type primary tumor, new mutations were highlighted: EGFR p.746_750del and KRAS p.G12V. Hypermetabolic CTC can be enriched without the need of dedicated equipment and their mutational status can successfully be assessed by ddPCR. Finally, the finding of new mutations supports the possibility of probing tumor heterogeneity. Full article
(This article belongs to the Special Issue Circulating Tumor Cells (CTCs))
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