Circulating tumor cells (CTCs) in blood of breast cancer patients:
Cytological detection and technical characterization
Enrica Bresaola, Mara Jo Miller, Marco Picozzi, Maria Teresa Sandri, Chiara Casadio
______________________Cytology Unit, Department of Pathology & Laboratory Medicine
European Institute of Oncology, Milan, Italy
Introduction
Circulating tumor cells (CTCs) were described for the first time more than one century ago and their occurrence in the bloodstream fit with the hematogenous theory of metastatization
Currently, their detection may play a pivotal role in the prognosis and prediction of therapy efficacy, providing us with insights into the clinical outcome, cell dissemination, drug resistance and treatment-induced cell death
Purpose
To assess the feasibility of detecting CTCs in blood samples of breast cancer patients using the Thin Prep® cytological preparation after a concurrent analysis with the CellSearch System (Veridex LLC, Warren, NJ)
To further characterize these cells according to estrogen receptor immunoreactivity and Her-2/neu gene status evaluation by fluorescence in situ hybridization (FISH) analysis
Materials and Methods
7.5 ml of whole blood were drawn from breast cancer patients into the CellSave Preservative Tube containing a cellular preservative and processed within 72 hours
The CellSearch Profile kit was then utilized to separate the CTCs by treatment with iron particles coated with antibodies against the Epithelial Cell Adhesion Molecule (EpCAM) for capturing CTCs
CTCs were then magnetically separated out and concentrated into a remaining aliquot of 1ml
Materials and Methods
This aliquot was centrifuged at 1700 rpm for 7 minutes and the supernatant discarded
The pellet was then added directly into the Preservcyt® vial for subsequent processing of ThinPrep slides
The slides were colored with H&E and evaluated microscopically
Estrogen receptor immunostaining and FISH analysis were carried out according to previously refined laboratory methods
Results
A total of 106 blood samples, where the CellSearch System obtained CTCs, were further evaluated cytologically
Of these, 60 were negative and 46 were positive for
malignant cells (range: 1-615) (Figure 1)
Figure 1
Results Immunocytochemistry for estrogen receptor was
performed in 7 samples and only one case showed ER-positive tumor cells
10 cases were analyzed by FISH: 7 cases had no amplification while in 3 cases no more cells were detected
+ve cell -ve cell
Conclusions
Cytological detection of CTCs in blood specimens from breast cancer patients can be useful in providing samples for testing predictive indicators of prognosis and clinical response during therapy
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