In vitro validation of an ultra-sensitive scanning fluorescence microscope for analysis of Circulating Tumor Cells

Analysis of circulating tumor cells (CTC) holds promise of providing liquid biopsies from patients with cancer. However, current methods include enrichment procedures. We present a method (CytoTrack®), where CTC from 7.5 mL of blood is stained, analyzed and counted by a scanning fluorescence microscope. The method was validated by breast cancer cells (MCF‐7) spiked in blood from healthy donors. The number of cells spiked in each blood sample was exactly determined by cell sorter and performed in three series of three samples spiked with 10, 33 or 100 cells in addition with three control samples for each series. The recovery rate of 10, 33 and 100 tumor cells in a blood sample was 55%, 70% and 78%, percent coefficient of variation (CV%) for samples was 59%, 32% and 18%, respectively. None of the control samples contained CTC. In conclusion, the method has been validated to highly sensitively detect breast cancer cells in spiking experiments and should be tested on blood samples from breast cancer patients. The method could benefit from automation that could reduce the CV%, and further optimization of the procedure to increase the recovery.

[1]  E. Lianidou,et al.  Circulating tumor cells in breast cancer: detection systems, molecular characterization, and future challenges. , 2011, Clinical chemistry.

[2]  D. Dearnaley,et al.  Characterization of ERG, AR and PTEN gene status in circulating tumor cells from patients with castration-resistant prostate cancer. , 2009, Cancer research.

[3]  Nada Jabado,et al.  New technologies for the detection of circulating tumour cells. , 2010, British medical bulletin.

[4]  E. Galleher,et al.  CIRCULATING "CANCER CELLS". , 1964, JAMA.

[5]  E. Lianidou,et al.  Circulating tumor cells as emerging tumor biomarkers in breast cancer , 2011, Clinical chemistry and laboratory medicine.

[6]  Leon W.M.M. Terstappen,et al.  Circulating Tumor Cells versus Imaging—Predicting Overall Survival in Metastatic Breast Cancer , 2006, Clinical Cancer Research.

[7]  Michael Koutsilieris,et al.  Detection of the circulating tumor cells in cancer patients. , 2010, Future oncology.

[8]  B. Naume,et al.  Detection and clinical relevance of early disseminated breast cancer cells depend on their cytokeratin expression pattern , 2011, Breast Cancer Research and Treatment.

[9]  Joshua M. Kunken,et al.  Fluid biopsy in patients with metastatic prostate, pancreatic and breast cancers , 2012, Physical biology.

[10]  A. Jemal,et al.  Global Cancer Statistics , 2011 .

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

[12]  G. Doyle,et al.  Significance of Circulating Tumor Cells Detected by the CellSearch System in Patients with Metastatic Breast Colorectal and Prostate Cancer , 2009, Journal of oncology.

[13]  J. Bono,et al.  All circulating EpCAM+CK+CD45- objects predict overall survival in castration-resistant prostate cancer. , 2010, Annals of oncology : official journal of the European Society for Medical Oncology.

[14]  Jia Fan,et al.  Circulating tumor cells: advances in detection methods, biological issues, and clinical relevance , 2011, Journal of Cancer Research and Clinical Oncology.

[15]  E. S. Pearson,et al.  Tests for departure from normality. Empirical results for the distributions of b2 and √b1 , 1973 .

[16]  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.

[17]  A. Órfão,et al.  Loss of CD34(+) hematopoietic progenitor cells due to washing can be reduced by the use of fixative-free erythrocyte lysing reagents. , 2000, Journal of immunological methods.

[18]  A. Jemal,et al.  Global cancer statistics , 2011, CA: a cancer journal for clinicians.

[19]  Wenjun Guo,et al.  The Epithelial-Mesenchymal Transition Generates Cells with Properties of Stem Cells , 2008, Cell.

[20]  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.

[21]  G. Hortobagyi,et al.  Circulating tumor cells and [18F]fluorodeoxyglucose positron emission tomography/computed tomography for outcome prediction in metastatic breast cancer. , 2009, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[22]  S. Eccles,et al.  Metastasis: recent discoveries and novel treatment strategies , 2007, The Lancet.

[23]  Jaw-Yuan Wang,et al.  Detection of KRAS Oncogene in Peripheral Blood as a Predictor of the Response to Cetuximab Plus Chemotherapy in Patients with Metastatic Colorectal Cancer , 2009, Clinical Cancer Research.

[24]  Alison Stopeck,et al.  Circulating tumor cells: a novel prognostic factor for newly diagnosed metastatic breast cancer. , 2005, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.