Morphological analysis of circulating tumour cells in patients undergoing surgery for non‐small cell lung carcinoma using the isolation by size of epithelial tumour cell (ISET) method

Hofman, Véronique; Long‐Mira, Elodie; Bonnetaud, Christelle; Vignaud, Jean-Michel; Fléjou, Jean‐François; Lantuéjoul, Sylvie; Piaton, Éric; Mourad, Nathalie; Butori, Catherine; Selva, Éric; Marquette, Charles-Hugo; Poudenx, M.; Sibon, Stéphanie; Kelhef, Sabrina; Vénissac, Nicolas; Jaı̈s, Jean-Philippe; Mouroux, Jérôme; Molina, Thierry Jo; Vielh, Philippe; Hofman, Paul

doi:10.1111/j.1365-2303.2010.00835.x

Cited by 115 publications

(126 citation statements)

References 39 publications

(93 reference statements)

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“…27,28 and clinical validation. 29 In addition, this technique has been show to be extremely sensible and specific as cells with malignant phenotype were found only in patients with cancer and never in healthy volunteers when tested on ISET (reviewed by Chinen et al, 2014). 30 Thus, we suggest that the method described here using CTCs isolated by ISET can be used for the most common mutations, as it is faster, cheaper and practical.…”

Section: Discussionmentioning

confidence: 89%

Detection of KRAS mutations in circulating tumor cells from patients with metastatic colorectal cancer

Buim

Fanelli

Souza

et al. 2015

Cancer Biology & Therapy

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Background: Quantification of Circulating Tumor Cells (CTCs) as a prognostic marker in metastatic colorectal cancer (mCRC) has already been validated and approved for routine use. However, more than quantification, qualification or characterization of CTCs is gaining importance, since the genetic characterization of CTCs may reflect, in a real time fashion, genetic profile of the disease. Objective: To characterize KRAS mutations (codon 12 and 13) in CTCs from patients with mCRC and to compare with matched primary tumor. Additionally, correlate these mutations with clinical and pathological features of patients. Methods: Blood samples were collected from 26 patients with mCRC from the AC Camargo Cancer Center (São Paulo-Brazil). CTCs were isolated by ISET technology (Isolation by Size of Epithelial Tumors; Rarecells Diagnostics, France) and mutations analyzes were performed by pyrosequencing (QIAGEN). Results: KRAS mutation was detected in 7 of the 21 cases (33%) of samples from CTCs. In matched primary tumors, 9 of the 24 cases (37.5%) were found KRAS mutated. We observed that 5 of the 9 samples with KRAS mutation in their primary tumor had also KRAS mutation in CTCs, meaning a concordance of 71% of matched cases (P D 0.017). KRAS mutation neither on primary tumor nor in CTCs was associated with clinical-pathological parameters analyzed. Conclusion: Faced with a polyclonal disease like colorectal cancer, which is often treated with alternating and successive lines of chemotherapy, real time genetic characterization of CTCs, in a fast and feasible fashion, can provide important information to clinical management of metastatic patients. Although our cohort was limited, it was possible to show a high grade of concordance between primary tumor and CTCs, which suggests that CTCs can be used as surrogate of primary tumors in clinical practice, when the knowledge of mutation profile is necessary and the primary tumor is not available.

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Section: Discussionmentioning

confidence: 89%

Detection of KRAS mutations in circulating tumor cells from patients with metastatic colorectal cancer

Buim

Fanelli

Souza

et al. 2015

Cancer Biology & Therapy

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“…[16][17][18] Five milliliters of whole blood was diluted up to 8 ml with filtration buffer containing 0.2% paraformaldehyde. Then, each sample was harvested by a membrane with an 8-mm diameter and cylindrical pores.…”

Section: Ctcs/ctm Detection and Analysismentioning

confidence: 99%

“…Imaging of CTCs was captured with a confocal microscope (Olympus, DP130, Japan) with a 40£ or 100£ magnification objective. Cells isolated in this study were assigned as CTCs only if they met the following morphological criteria 4,17,18 ]: 1) the nuclear-cytoplasmic ratio was greater than 0.8; 2) the long diameter of nucleus was larger than 18 mm; 3) irregularity (thickened, sunken and wrinkled) of the nuclear membrane; 4) a large nucleoli or abnormal mitotic figures; or 5) the presence of tumor cell aggregations. CTC clusters formed by at least 3 tumor cells were called circulating tumor microemboli (CTM).…”

Section: Ctcs/ctm Detection and Analysismentioning

confidence: 99%

“…In the present study, we evaluate the association between EBV activation and CTC detection in NPC patients. We applied the method of Isolation by Size of Epithelial Tumor (ISET), which has been proven to be useful in detection of various cancer cells 16,17,18 ], to identify CTCs in NPC patients. Moreover, immunohistochemical staining using the CK5/6 (cytokeratin5/6) and P63 plus in situ hybridization using the EBV-specific biomarker of EBERs (EBV-encoded RNAs) was used to help differentiate NPC cells from normal blood cells.…”

Section: Introductionmentioning

confidence: 99%

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The association between circulating tumor cells and Epstein-Barr virus activation in patients with nasopharyngeal carcinoma

Huang

et al. 2017

Cancer Biology & Therapy

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Background: Circulating tumor cells (CTCs) and microemboli (CTM) are attracting increasing attention in medical biology and clinical practice. However, the clinical relevance of CTCs in nasopharyngeal carcinoma (NPC) has not yet been ascertained, and no study has focused on the influence of Epstein-Barr virus (EBV) status on CTCs in NPC patients. These issues were therefore examined. Methods: Peripheral blood samples were prospectively obtained from 33 NPC patients before treatment. CTCs and CTM were captured using the Isolation by Size of Epithelial Tumor (ISET) method. Immunohistochemistry on CK5/6 (cytokeratin5/6) and P63, as well as in situ hybridization of EBERs (EBV-encoded RNAs) were used to validate the harvested tumor cells. Results: Out of 33 NPC patients, CTCs were detected in 22 cases (66.7%), and CTM were observed in 2 cases (6.1%). CTCs were presented in all stages of NPC patients but had no association with the TNM stages (all P > 0.05). The presence of CTCs appeared to correlate with EBV activation status. Among the total NPC patients, the EBV VCA-IgA levels in CTC-positive cases were higher than that in CTC-negative cases (negative vs. positive: 3.88 vs. 4.86, P D 0.016). A similar result was observed in the patients without distant metastasis (negative vs. positive: 3.76 vs. 4.95, P D 0.009). When the number of CTCs was considered, CTC counts were found to correlate with EBV VCA-IgA (R D 0.382, P D 0.041) and EBV DNA load (R D 0.396, P D 0.033) for all NPC patients as well as NPC patients without distant metastases. Conclusions: These findings strongly suggested detectable CTCs/CTM in all stages of NPC patients and support a positive correlation between CTCs and EBV activation in NPC patients.

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“…Some of these technologies are based on the segregation of malignant and benign tumor cells cytomorphologically from nonhematological circulating cells (65)(66)(67). Methods based on size-specific discrimination of epithelial tumor cells are also used (65)(66)(67). A new approach is to work with xenograft CTCs by producing CTC-derived transplants.…”

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confidence: 99%

Techniques for maximizing the performance of molecular pathology testing: responsibilities of all pathologists

Uzun

Sarıoğlu²

2017

TJPATH

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Molecular pathological analysis has an expanding role in patient diagnosis and management. The performance of these techniques relies on excellent laboratory procedures. However, the crucial step is obtaining the best samples for molecular analysis. Archiving and selection of these are the responsibilities of all pathologists even if they are not working at a center with molecular pathological facilities.This review focuses on the features of different types of materials for molecular pathological analysis. Many steps that might affect the results, including communication between the pathologist and the oncology team, features of different types of materials (cytological, tissue blocks, biopsy, circulating tumor cells (CTCs) and cell-free circulating nucleic acids), effects of tissue processing, methods for selecting the best material, and tissue saving and tumor enrichment methods are discussed. The procedures for referral to a center for molecular pathological analysis are also mentioned.Awareness of the importance of the cytopathological and histopathological material of the patients for future molecular pathological analysis by pathologists is of the utmost importance.Key Words: Molecular Pathology, Tissue saving, Tumor enrichment, Tumor percentage INTRODUCTION "Molecular Pathology" is one of the pathological methods with expanding role in patient diagnosis, prediction of prognosis and treatment. Tissue and cellular material from patients have always been valued in pathology laboratories; however they gain additional importance in the molecular pathology era with the increase in new disease classifications according to molecular changes and new targeted therapy options being determined with predictive molecular testing.While the value of diseased and normal tissue is expanding with the advances in immunohistochemistry and molecular pathology, minimally invasive techniques have taken the place of radical processes in tissue sampling along with developments in the technology. These techniques are better for the comfort of the patient but they also have disadvantages, as they provide smaller amounts of tissue for diagnosis. The management of tissues obtained with these techniques can be problematic for pathologists. The application of individualized treatment regimens developed in recent years has depended on molecular studies. The adequacy of the obtained material and its reliability for molecular investigation are of utmost importance in many cases.The diagnosis of lung carcinomas is usually made with small biopsies or cytological materials and these materials are generally the only options for further studies. Transthoracic, transbronchial fine needle aspirations, forceps biopsies, bronchial brushes, and washes are some of these materials (1). A molecular examination requires the best process and good sample quality. The most important factor for the success of molecular tests is the number of tumor cells and the percentage of mutant cells (2). The requirements for tumor cell number and mutation perc...

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Morphological analysis of circulating tumour cells in patients undergoing surgery for non‐small cell lung carcinoma using the isolation by size of epithelial tumour cell (ISET) method

Abstract: Identification of CTCs in resectable NSCLC patients, using ISET technology and according to cytopathological criteria of malignancy, appears to be a new and promising field of cytopathology with potential relevance to lung oncology.

Cited by 115 publications

References 39 publications

Detection of KRAS mutations in circulating tumor cells from patients with metastatic colorectal cancer

Detection of KRAS mutations in circulating tumor cells from patients with metastatic colorectal cancer

The association between circulating tumor cells and Epstein-Barr virus activation in patients with nasopharyngeal carcinoma

Techniques for maximizing the performance of molecular pathology testing: responsibilities of all pathologists

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