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13 pages, 1122 KiB

Open AccessArticle

Single-Cell Analysis of Circulating Tumor Cells from Patients with Colorectal Cancer Captured with a Dielectrophoresis-Based Micropore System

by Masatoshi Nomura, Yuichiro Miyake, Akira Inoue, Yuhki Yokoyama, Nanaka Noda, Shihori Kouda, Tsuyoshi Hata, Takayuki Ogino, Norikatsu Miyoshi, Hidekazu Takahashi, Mamoru Uemura, Tsunekazu Mizushima, Yuichiro Doki, Hidetoshi Eguchi and Hirofumi Yamamoto

Biomedicines 2023, 11(1), 203; https://doi.org/10.3390/biomedicines11010203 - 13 Jan 2023

Cited by 3 | Viewed by 1775

Abstract

This study aimed to analyze circulating tumor cells (CTCs) from patients with colorectal cancer (CRC). We designed a dielectrophoresis-based micropore system and tested its cell capture with HT29 colon cancer cells. Then, blood samples were drawn from 24 patients with stages II-IV CRC. [...] Read more.

This study aimed to analyze circulating tumor cells (CTCs) from patients with colorectal cancer (CRC). We designed a dielectrophoresis-based micropore system and tested its cell capture with HT29 colon cancer cells. Then, blood samples were drawn from 24 patients with stages II-IV CRC. Mononuclear cells were isolated and loaded into the micropore system. Single cells were positioned into small pores with dielectrophoresis. After labeling the cells with the appropriate antibodies, tumor-like cells were collected with an automated micromanipulator. We collected 43 CTCs from 15 out of 24 patient samples. The presence of CTC was significantly associated with ling metastasis. We performed whole genome amplification, followed by PCR and Sanger sequencing, to examine the point mutations in the KRAS, BRAF, and PIK3CA genes. This mutation analysis was successfully performed in 35 cells. Among the 14 cytokeratin (CK)-positive cells, we found PIK3CA mutations in three cells (21%) from two patients. Among the 21 CK-negative cells, we found a KRAS mutation in one cell (5%) from one patient and a PIK3CA mutation in one cell (5%) from one patient. It is noteworthy that these mutations were not detected in the corresponding primary tumors. In conclusion, dielectrophoresis-based capture in a micropore system was useful for detecting both CK-positive and CK-negative CTCs. This simple method could be applied to various tumor types. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

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25 pages, 5918 KiB

Open AccessArticle

ALCAM: A Novel Surface Marker on EpCAM^low Circulating Tumor Cells

by Rossana Signorelli, Teresa Maidana Giret, Oliver Umland, Marco Hadisurya, Shweta Lavania, John Lalith Charles Richard, Ashley Middleton, Melinda Minucci Boone, Ayse Burcu Ergonul, Weiguo Andy Tao, Haleh Amirian, Anton Iliuk, Aliya Khan, Robert Diaz, Daniel Bilbao Cortes, Monica Garcia-Buitrago and Harrys Kishore Charles Jacob

Biomedicines 2022, 10(8), 1983; https://doi.org/10.3390/biomedicines10081983 - 16 Aug 2022

Viewed by 2585

Abstract

Background: Current strategies in circulating tumor cell (CTC) isolation in pancreatic cancer heavily rely on the EpCAM and cytokeratin cell status. EpCAM is generally not considered a good marker given its transitory change during Epithelial to Mesenchymal Transition (EMT) or reverse EMT. There [...] Read more.

Background: Current strategies in circulating tumor cell (CTC) isolation in pancreatic cancer heavily rely on the EpCAM and cytokeratin cell status. EpCAM is generally not considered a good marker given its transitory change during Epithelial to Mesenchymal Transition (EMT) or reverse EMT. There is a need to identify other surface markers to capture the complete repertoire of PDAC CTCs. The primary objective of the study is to characterize alternate surface biomarkers to EpCAM on CTCs that express low or negligible levels of surface EpCAM in pancreatic cancer patients. Methods: Flow cytometry and surface mass spectrometry were used to identify proteins expressed on the surface of PDAC CTCs in culture. CTCs were grown under conditions of attachment and in co-culture with naïve neutrophils. Putative biomarkers were then validated in GEMMs and patient samples. Results: Surface proteomic profiling of CTCs identified several novel protein biomarkers. ALCAM was identified as a novel robust marker in GEMM models and in patient samples. Conclusions: We identified several novel surface biomarkers on CTCs expressed under differing conditions of culture. ALCAM was validated and identified as a novel alternate surface marker on EpCAM^low CTCs. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

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14 pages, 1310 KiB

Open AccessArticle

PD-L1/pS6 in Circulating Tumor Cells (CTCs) during Osimertinib Treatment in Patients with Non-Small Cell Lung Cancer (NSCLC)

by Evangelia Pantazaka, Aliki Ntzifa, Argyro Roumeliotou, Evi Lianidou, Vassilis Georgoulias, Athanasios Kotsakis and Galatea Kallergi

Biomedicines 2022, 10(8), 1893; https://doi.org/10.3390/biomedicines10081893 - 05 Aug 2022

Cited by 6 | Viewed by 1808

Abstract

The PD-1/PD-L1 axis provides CTCs an escape route from the immune system. Phosphorylation of the ribosomal protein S6 is implicated in the same pathway, following mTOR activation. The aim of the study was to investigate the expression of PD-L1 and pS6 in CTCs [...] Read more.

The PD-1/PD-L1 axis provides CTCs an escape route from the immune system. Phosphorylation of the ribosomal protein S6 is implicated in the same pathway, following mTOR activation. The aim of the study was to investigate the expression of PD-L1 and pS6 in CTCs from NSCLC patients under Osimertinib treatment at a single cell level. CTCs were isolated using ISET from NSCLC patients’ blood [37 at baseline, 25 after the 1st cycle, and 23 at the end of treatment (EOT)]. Staining was performed using immunofluorescence. Cytokeratin-positive (CK⁺) CTCs were detected in 62% of patients. CK⁺PD-L1⁺CD45⁻ and CK⁺pS6⁺ phenotypes were detected in 38% and 41% of the patients at baseline, in 28% and 32% after 1st cycle, and in 30% and 35% at EOT, respectively. Spearman’s analysis revealed statistically significant correlations between PD-L1 and pS6 phenotypes at all time points. Survival analysis revealed that CK⁺pS6⁺ (p = 0.003) and CK^lowpS6⁺ (p = 0.021) phenotypes after 1st cycle were related to significantly decreased one-year progression-free survival (PFS_12m) and PFS, respectively. CK⁺PD-L1⁺CD45⁻phenotype at baseline and after 1st cycle showed a trend for decreased PFS_12m. Increased expression of PD-L1/pS6 in CTCs of Osimertinib-treated NSCLC patients implies the activation of the corresponding pathway, which is potentially associated with poor clinical outcomes. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

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17 pages, 2257 KiB

Open AccessArticle

Synergistic Analysis of Circulating Tumor Cells Reveals Prognostic Signatures in Pilot Study of Treatment-Naïve Metastatic Pancreatic Cancer Patients

by Sarah Owen, Emily Prantzalos, Valerie Gunchick, Vaibhav Sahai and Sunitha Nagrath

Biomedicines 2022, 10(1), 146; https://doi.org/10.3390/biomedicines10010146 - 11 Jan 2022

Cited by 3 | Viewed by 2184

Abstract

Pancreatic ductal adenocarcinoma is typically diagnosed at late stages and has one of the lowest five-year survival rates of all malignancies. In this pilot study, we identify signatures related to survival and treatment response found in circulating tumor cells (CTCs). Patients with poor [...] Read more.

Pancreatic ductal adenocarcinoma is typically diagnosed at late stages and has one of the lowest five-year survival rates of all malignancies. In this pilot study, we identify signatures related to survival and treatment response found in circulating tumor cells (CTCs). Patients with poor survival had increased mutant KRAS expression and deregulation of connected pathways such as PI3K-AKT and MAPK signaling. Further, in a subset of these patients, expression patterns of gemcitabine resistance mechanisms were observed, even prior to initiating treatment. This work highlights the need for identifying patients with these resistance profiles and designing treatment regimens to circumvent these mechanisms. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

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Review

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17 pages, 6959 KiB

Open AccessReview

Clinical Applications of Cancer-Associated Cells Present in the Blood of Cancer Patients

by Cha-Mei Tang and Daniel L. Adams

Biomedicines 2022, 10(3), 587; https://doi.org/10.3390/biomedicines10030587 - 02 Mar 2022

Cited by 10 | Viewed by 2395

Abstract

The ability to obtain tumor material from cells in the blood of cancer patients provides a significant benefit over the use of tumor tissue as a diagnostic to make treatment decisions. However, the traditionally defined circulating tumor cell (CTC) has been shown to [...] Read more.

The ability to obtain tumor material from cells in the blood of cancer patients provides a significant benefit over the use of tumor tissue as a diagnostic to make treatment decisions. However, the traditionally defined circulating tumor cell (CTC) has been shown to be useful only in some cases. A recently identified type of circulating stromal cell, which appears to be more frequent than CTCs, was found engulfing tumor material at the tumor site and then entering the blood stream. These cells were defined as cancer-associated macrophage-like cells (CAMLs). Together, CTCs and CAMLs may be able to provide information for cancer detection and diagnosis, without the use of tissue. CTCs and CAMLs have many clinical applications, three of which are summarized in this review: for prognosis, as companion diagnostics, and for residual disease monitoring. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

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12 pages, 293 KiB

Open AccessReview

Circulating Tumor Cells as a Tool to Untangle the Breast Cancer Heterogeneity Issue

by Tania Rossi, Giulia Gallerani, Giovanni Martinelli, Roberta Maltoni and Francesco Fabbri

Biomedicines 2021, 9(9), 1242; https://doi.org/10.3390/biomedicines9091242 - 16 Sep 2021

Cited by 7 | Viewed by 1623

Abstract

Breast cancer (BC) is a disease characterized by high degrees of heterogeneity at morphologic, genomic, and genetic levels, even within the same tumor mass or among patients. As a consequence, different subpopulations coexist and less represented clones may have a selective advantage, significantly [...] Read more.

Breast cancer (BC) is a disease characterized by high degrees of heterogeneity at morphologic, genomic, and genetic levels, even within the same tumor mass or among patients. As a consequence, different subpopulations coexist and less represented clones may have a selective advantage, significantly influencing the outcome of BC patients. Circulating tumor cells (CTCs) represent a rare population of cells with a crucial role in metastatic cascade, and in recent years have represented a fascinating alternative to overcome the heterogeneity issue as a “liquid biopsy”. However, besides the raw enumeration of these cells in advanced epithelial tumors, there are no CTC-based assays applied in the clinical practice to improve personalized medicine. In this review, we report the latest findings in the field of CTCs for intra-tumoral heterogeneity unmasking in BC, supporting the need to deepen their analysis to investigate their role in metastatic process and include the molecular characterization in the clinical practice. In the future, CTCs will be helpful in monitoring patients during treatment, as well as to better address therapeutic strategies. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

15 pages, 326 KiB

Open AccessReview

Clinical Relevance of Circulating Tumor Cells in Prostate Cancer Management

by Wojciech A. Cieślikowski, Andrzej Antczak, Michał Nowicki, Maciej Zabel and Joanna Budna-Tukan

Biomedicines 2021, 9(9), 1179; https://doi.org/10.3390/biomedicines9091179 - 08 Sep 2021

Cited by 17 | Viewed by 1845

Abstract

Given the low specificity of the routinely used biomarker prostate-specific antigen, circulating tumor cell (CTC) enumeration seems to be particularly useful in the monitoring of prostate cancer. In this review, we focused on a few aspects of CTC enumeration in prostate malignancies: prognostic [...] Read more.

Given the low specificity of the routinely used biomarker prostate-specific antigen, circulating tumor cell (CTC) enumeration seems to be particularly useful in the monitoring of prostate cancer. In this review, we focused on a few aspects of CTC enumeration in prostate malignancies: prognostic value in metastatic and non-metastatic tumors, role in the monitoring of treatment outcomes, use as a surrogate marker for survival, and other applications, mostly for research purposes. CTC enumeration, without a doubt, offers an attractive perspective in the management of prostate cancer. However, the vast majority of available data about the role of CTC in this malignancy originate from randomized studies of anticancer agents and do not necessarily translate into real-world clinical practice. Further, most studies on the application of CTC in prostate cancer patients were limited to advanced stages of this malignancy. Meanwhile, the role of CTC in the early stages of prostate cancer, in which some patients may present with occult disseminated disease, is still relatively poorly understood, and should thus be studied extensively. Other obstacles in the widespread application of CTC enumeration in routine clinical practice include considerable discrepancies in the number of cells determined with various commercially available systems. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

19 pages, 857 KiB

Open AccessReview

Circulating Tumor Cells: Technologies and Their Clinical Potential in Cancer Metastasis

by Jerry Xiao, Paula R. Pohlmann, Claudine Isaacs, Benjamin A. Weinberg, Aiwu R. He, Richard Schlegel and Seema Agarwal

Biomedicines 2021, 9(9), 1111; https://doi.org/10.3390/biomedicines9091111 - 30 Aug 2021

Cited by 26 | Viewed by 4197

Abstract

Circulating tumor cells (CTCs) are single cells or clusters of cells within the circulatory system of a cancer patient. While most CTCs will perish, a small proportion will proceed to colonize the metastatic niche. The clinical importance of CTCs was reaffirmed by the [...] Read more.

Circulating tumor cells (CTCs) are single cells or clusters of cells within the circulatory system of a cancer patient. While most CTCs will perish, a small proportion will proceed to colonize the metastatic niche. The clinical importance of CTCs was reaffirmed by the 2008 FDA approval of CellSearch^®, a platform that could extract EpCAM-positive, CD45-negative cells from whole blood samples. Many further studies have demonstrated the presence of CTCs to stratify patients based on overall and progression-free survival, among other clinical indices. Given their unique role in metastasis, CTCs could also offer a glimpse into the genetic drivers of metastasis. Investigation of CTCs has already led to groundbreaking discoveries such as receptor switching between primary tumors and metastatic nodules in breast cancer, which could greatly affect disease management, as well as CTC-immune cell interactions that enhance colonization. In this review, we will highlight the growing variety of isolation techniques for investigating CTCs. Next, we will provide clinically relevant context for CTCs, discussing key clinical trials involving CTCs. Finally, we will provide insight into the future of CTC studies and some questions that CTCs are primed to answer. Full article

(This article belongs to the Special Issue The Biology of Circulating Tumor Cells)

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