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. 2009 Jun 5:2:24.
doi: 10.1186/1756-8722-2-24.

Identification of circulating tumour cells in early stage breast cancer patients using multi marker immunobead RT-PCR

Michael P Raynor  1 , Sally-Anne Stephenson, Kenneth B Pittman, David C A Walsh, Michael A Henderson, Alexander Dobrovic
Affiliations

Identification of circulating tumour cells in early stage breast cancer patients using multi marker immunobead RT-PCR

Michael P Raynor et al. J Hematol Oncol. .

Abstract

Introduction: The ability to screen blood of early stage operable breast cancer patients for circulating tumour cells is of potential importance for identifying patients at risk of developing distant relapse. We present the results of a study of the efficacy of the immunobead RT-PCR method in identifying patients with circulating tumour cells.

Results: Immunomagnetic enrichment of circulating tumour cells followed by RT-PCR (immunobead RT-PCR) with a panel of five epithelial specific markers (ELF3, EPHB4, EGFR, MGB1 and TACSTD1) was used to screen for circulating tumour cells in the peripheral blood of 56 breast cancer patients. Twenty patients were positive for two or more RT-PCR markers, including seven patients who were node negative by conventional techniques. Significant increases in the frequency of marker positivity was seen in lymph node positive patients, in patients with high grade tumours and in patients with lymphovascular invasion. A strong trend towards improved disease free survival was seen for marker negative patients although it did not reach significance (p = 0.08).

Conclusion: Multi-marker immunobead RT-PCR analysis of peripheral blood is a robust assay that is capable of detecting circulating tumour cells in early stage breast cancer patients.

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Figures

Figure 1
Figure 1
IB RT-PCR sensitivity test on dilutions of MDA-MB-453 in normal blood. Cells were added to 10 mls of normal blood. Lane 1, M = pUC19/HpaII marker. Lanes 2–4, no added cells. Lanes 5–7, 10 added cells. Lanes 8–10, 100 added cells, Lanes 11–13, 1000 added cells. Lane 14, C = cDNA positive control. Lane 15, R = RT negative control. Lanes 16–18, N = PCR negative controls. Product size is shown in base pairs (bp). A genomic DNA band is seen above the RT-PCR band for ELF3. The legend shows cells/10 ml of blood.
Figure 2
Figure 2
IB RT-PCR analysis for a representative group of 10 breast cancer patients. Patient number corresponds to numbers provided in Table 2. Product sizes are shown in base pairs. M = pUC19/HpaII marker. A genomic DNA band is seen above RT-PCR band for ELF3.
Figure 3
Figure 3
Disease free survival for 56 patients with early stage breast cancer. Comparing patients positive for RT-PCR markers (2 or more) with patients negative for RT-PCR markers (0 or 1).
See this image and copyright information in PMC

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