Size-Based Isolation of Circulating Tumor Cells in Lung Cancer Patients Using a Microcavity Array System

Background Epithelial cell adhesion molecule (EpCAM)-based enumeration of circulating tumor cells (CTC) has prognostic value in patients with solid tumors, such as advanced breast, colon, and prostate cancer. However, poor sensitivity has been reported for non-small cell lung cancer (NSCLC). To address this problem, we developed a microcavity array (MCA) system integrated with a miniaturized device for CTC isolation without relying on EpCAM expression. Here, we report the results of a clinical study on CTCs of advanced lung cancer patients in which we compared the MCA system with the CellSearch system, which employs the conventional EpCAM-based method. Methods Paired peripheral blood samples were collected from 43 metastatic lung cancer patients to enumerate CTCs using the CellSearch system according to the manufacturer’s protocol and the MCA system by immunolabeling and cytomorphological analysis. The presence of CTCs was assessed blindly and independently by both systems. Results CTCs were detected in 17 of 22 NSCLC patients using the MCA system versus 7 of 22 patients using the CellSearch system. On the other hand, CTCs were detected in 20 of 21 small cell lung cancer (SCLC) patients using the MCA system versus 12 of 21 patients using the CellSearch system. Significantly more CTCs in NSCLC patients were detected by the MCA system (median 13, range 0–291 cells/7.5 mL) than by the CellSearch system (median 0, range 0–37 cells/7.5 ml) demonstrating statistical superiority (p = 0.0015). Statistical significance was not reached in SCLC though the trend favoring the MCA system over the CellSearch system was observed (p = 0.2888). The MCA system also isolated CTC clusters from patients who had been identified as CTC negative using the CellSearch system. Conclusions The MCA system has a potential to isolate significantly more CTCs and CTC clusters in advanced lung cancer patients compared to the CellSearch system.

[1]  D. Altman,et al.  STATISTICAL METHODS FOR ASSESSING AGREEMENT BETWEEN TWO METHODS OF CLINICAL MEASUREMENT , 1986, The Lancet.

[2]  K. Pienta,et al.  Circulating Tumor Cells Predict Survival Benefit from Treatment in Metastatic Castration-Resistant Prostate Cancer , 2008, Clinical Cancer Research.

[3]  Tomoko Yoshino,et al.  Size-selective microcavity array for rapid and efficient detection of circulating tumor cells. , 2010, Analytical chemistry.

[4]  Chwee Teck Lim,et al.  Versatile label free biochip for the detection of circulating tumor cells from peripheral blood in cancer patients. , 2010, Biosensors & bioelectronics.

[5]  Michael Morse,et al.  Relationship of circulating tumor cells to tumor response, progression-free survival, and overall survival in patients with metastatic colorectal cancer. , 2008, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[6]  Bernard Lacour,et al.  Impact of cytomorphological detection of circulating tumor cells in patients with liver cancer , 2004, Hepatology.

[7]  Han Wei Hou,et al.  Pinched flow coupled shear-modulated inertial microfluidics for high-throughput rare blood cell separation. , 2011, Lab on a chip.

[8]  Yukiko Nakamura,et al.  Prognostic Impact of Circulating Tumor Cells in Patients with Small Cell Lung Cancer , 2012, Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer.

[9]  Anil K Sood,et al.  A novel platform for detection of CK+ and CK- CTCs. , 2011, Cancer discovery.

[10]  Haruko Takeyama,et al.  High-density microcavity array for cell detection: single-cell analysis of hematopoietic stem cells in peripheral blood mononuclear cells. , 2009, Analytical chemistry.

[11]  K. Isselbacher,et al.  Isolation of circulating tumor cells using a microvortex-generating herringbone-chip , 2010, Proceedings of the National Academy of Sciences.

[12]  H. Hansen,et al.  Lung cancer. , 1990, Cancer chemotherapy and biological response modifiers.

[13]  Jonathan W. Uhr,et al.  Tumor Cells Circulate in the Peripheral Blood of All Major Carcinomas but not in Healthy Subjects or Patients With Nonmalignant Diseases , 2004, Clinical Cancer Research.

[14]  A. Gemma,et al.  F1000 highlights , 2010 .

[15]  Caroline Dive,et al.  Circulating tumor cells as a window on metastasis biology in lung cancer. , 2011, The American journal of pathology.

[16]  Alison Stopeck,et al.  Circulating tumor cells, disease progression, and survival in metastatic breast cancer. , 2004, The New England journal of medicine.

[17]  K. Schütze,et al.  Isolation by size of epithelial tumor cells : a new method for the immunomorphological and molecular characterization of circulatingtumor cells. , 2000, The American journal of pathology.

[18]  Li Zhang,et al.  Preliminary Investigation of the Clinical Significance of Detecting Circulating Tumor Cells Enriched from Lung Cancer Patients , 2009, Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer.

[19]  Mitch Dowsett,et al.  Cell filtration‐laser scanning cytometry for the characterisation of circulating breast cancer cells , 2003, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[20]  T. Molina,et al.  Detection of circulating tumor cells as a prognostic factor in patients undergoing radical surgery for non‐small‐cell lung carcinoma: comparison of the efficacy of the CellSearch Assay™ and the isolation by size of epithelial tumor cell method , 2011, International journal of cancer.

[21]  Tony J. Pircher,et al.  Detection of EpCAM-Negative and Cytokeratin-Negative Circulating Tumor Cells in Peripheral Blood , 2011, Journal of oncology.

[22]  Mieke Schutte,et al.  Anti-Epithelial Cell Adhesion Molecule Antibodies and the Detection of Circulating Normal-Like Breast Tumor Cells , 2009, Journal of the National Cancer Institute.

[23]  P. Paterlini-Bréchot,et al.  Circulating tumor cells (CTC) detection: clinical impact and future directions. , 2007, Cancer letters.

[24]  J Lawry,et al.  Monitoring tumour cells in the peripheral blood of small cell lung cancer patients. , 2002, Cytometry.

[25]  Caroline Dive,et al.  Tumorigenesis and Neoplastic Progression Evaluation of Circulating Tumor Cells and Serological Cell Death Biomarkers in Small Cell Lung Cancer Patients Undergoing Chemotherapy , 2010 .

[26]  Giuseppe Naso,et al.  Epithelial-mesenchymal transition and stemness features in circulating tumor cells from breast cancer patients , 2011, Breast Cancer Research and Treatment.

[27]  Jeffrey W. Clark,et al.  Anaplastic lymphoma kinase inhibition in non-small-cell lung cancer. , 2010, The New England journal of medicine.

[28]  G. Assmann,et al.  Isolation of prostate-derived single cells and cell clusters from human peripheral blood. , 1996, Cancer research.

[29]  Nobuyuki Yamamoto,et al.  Microcavity array system for size-based enrichment of circulating tumor cells from the blood of patients with small-cell lung cancer. , 2013, Analytical chemistry.

[30]  N. Tsubota,et al.  Circulating Tumor Cell as a Diagnostic Marker in Primary Lung Cancer , 2009, Clinical Cancer Research.

[31]  D. Planchard,et al.  A direct comparison of CellSearch and ISET for circulating tumour-cell detection in patients with metastatic carcinomas , 2011, British Journal of Cancer.

[32]  Bo Lu,et al.  3D microfilter device for viable circulating tumor cell (CTC) enrichment from blood , 2011, Biomedical microdevices.

[33]  Caroline Dive,et al.  Clinical significance and molecular characteristics of circulating tumor cells and circulating tumor microemboli in patients with small-cell lung cancer. , 2012, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[34]  Caroline Dive,et al.  Analysis of Circulating Tumor Cells in Patients with Non-small Cell Lung Cancer Using Epithelial Marker-Dependent and -Independent Approaches , 2012, Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer.

[35]  Brigitte Rack,et al.  Detection of Circulating Tumor Cells in Peripheral Blood of Patients with Metastatic Breast Cancer: A Validation Study of the CellSearch System , 2007, Clinical Cancer Research.

[36]  A. Beckett,et al.  AKUFO AND IBARAPA. , 1965, Lancet.

[37]  Hongshen Ma,et al.  Cell separation based on size and deformability using microfluidic funnel ratchets. , 2012, Lab on a chip.

[38]  Alessandro Lugli,et al.  Frequent EpCam protein expression in human carcinomas. , 2004, Human pathology.

[39]  D. Planchard,et al.  Detection of circulating tumour cells with a hybrid (epithelial/mesenchymal) phenotype in patients with metastatic non-small cell lung cancer , 2011, British Journal of Cancer.

[40]  P. Jänne,et al.  A new device for rapid isolation by size and characterization of rare circulating tumor cells. , 2011, Anticancer research.

[41]  J. A. van der Laak,et al.  Circulating tumour tissue fragments in patients with pulmonary metastasis of clear cell renal cell carcinoma , 2009, The Journal of pathology.

[42]  Satoshi Nakasono,et al.  Leukocyte counting from a small amount of whole blood using a size‐controlled microcavity array , 2012, Biotechnology and bioengineering.

[43]  Caroline Dive,et al.  Evaluation and prognostic significance of circulating tumor cells in patients with non-small-cell lung cancer. , 2011, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.