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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
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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

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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

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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

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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 HE and evaluated
  microscopically
• Estrogen receptor immunostaining and FISH
  analysis were carried out according to previously
  refined laboratory methods

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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)

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Figure 1
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Results

• Immunocytochemistry for estrogen receptor was
  performed in 7 samples and only one case showed
  ER-positive tumor cells

ve cell
-ve cell

• 10 cases were analyzed by FISH 7 cases had no
  amplification while in 3 cases no more cells were
  detected

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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