Yonsei Med J.  2017 Jan;58(1):19-26. 10.3349/ymj.2017.58.1.19.

Detection of Circulating Tumor Cells in Breast Cancer Patients Using Cytokeratin-19 Real-Time RT-PCR

• Park HS
• Han HJ
• Lee S
• Kim GM
• Park S
• Choi YA
• Lee JD
• Kim GM
• Sohn J
• Kim SI

• KMID: 2374184
• DOI: http://doi.org/10.3349/ymj.2017.58.1.19

Abstract

PURPOSE
The roles of circulating tumor cells (CTCs) as predictive and prognostic factors, as well as key mediators in the metastatic cascade, have been investigated. This study aimed to validate a method to quantify CTCs in peripheral blood using a real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay for cytokeratin (CK)-19 and to evaluate the utility of this assay in detecting CTCs in breast cancer patients.
MATERIALS AND METHODS
Real-time monitoring PCR of fluorescently labeled specific hybridization probes for CK-19 mRNA was established. Peripheral blood samples from 30 healthy donors, 69 patients with early breast cancer, 47 patients with locally advanced breast cancer, and 126 patients with metastatic breast cancer were prospectively obtained and analyzed for CTC detection.
RESULTS
CK-19 mRNA was not detectable in healthy subjects using the real-time RT-PCR method. The detection rates of CK-19 mRNA in breast cancer patients were 47.8% for early breast cancer (33/69), 46.8% for locally advanced breast cancer (22/47), and 61.1% for metastatic breast cancer (77/129). The detection rate of CK-19-positive CTCs in metastatic disease was slightly higher than early or locally advanced breast cancer; however, the detection rate according to disease burden was not statistically different (p=0.097). The detection rate was higher in patients with pleural metastasis (p=0.045). CTC detection was associated with poor survival (p=0.014).
CONCLUSION
A highly specific and sensitive CK-19 mRNA-based method to detect CTCs in peripheral blood in breast cancer patients can be used in further prospective studies to evaluate the predictive and prognostic importance of CTCs.

Keyword

Cytokeratin-19 mRNA; circulating tumor cells; RT-PCR; peripheral blood samples; metastatic breast cancer

MeSH Terms

Biomarkers, Tumor/*blood
Breast Neoplasms/blood/*pathology
Female
Humans
Keratin-19/*blood/genetics
*Neoplastic Cells, Circulating
Prognosis
Prospective Studies
RNA, Messenger/*blood
Real-Time Polymerase Chain Reaction
*Reverse Transcriptase Polymerase Chain Reaction/methods
Biomarkers, Tumor
Keratin-19
RNA, Messenger

Figure

• 

  Fig. 1 Ct values according to treatment setting in patients with CTCs. High Ct values correlated with low levels of gene expression, whereas low Ct values correlate with high levels of gene expression. The box plots represent the median, maximal, and minimal number of Ct values. There was no significant difference in terms of Ct values among three groups (p=0.16). CTC, circulating tumor cell; EBC, early breast cancer; LBC, locally advanced breast cancer; MBC, metastatic breast cancer.

• 

  Fig. 2 Overall survival according to the detection of CTCs defined by CK-19 mRNA-positive cells. CTC, circulating tumor cell; CK, cytokeratin.

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