Efficient leukocyte depletion by a novel microfluidic platform enables the molecular detection and characterization of circulating tumor cells
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Eva Obermayr1, Elisabeth Maritschnegg1, Christiane Agreiter1, Nina Pecha1,2, Paul Speiser1,2, Samir Helmy-Bader2, Sabine Danzinger3, Michael Krainer4, Christian Singer3 and Robert Zeillinger1
1Molecular Oncology Group, Department of Obstetrics and Gynecology, Comprehensive Cancer Center, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria
2Department of Obstetrics and Gynecology, Clinical Division for General Gynecology and Gynecological Oncology, Comprehensive Cancer Center, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria
3Department of Obstetrics and Gynecology, Division of Senology, Comprehensive Cancer Center, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria
4Department of Medicine I, Clinical Division of Oncology, Comprehensive Cancer Center, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna, Austria
Eva Obermayr, email: [email protected]
Keywords: circulating tumor cells, microfluidics, gene expression analysis, ovarian cancer
Received: December 15, 2016 Accepted: September 30, 2017 Published: November 28, 2017
RT-qPCR is a highly sensitive approach to detect rare transcripts, as derived from circulating tumor cells (CTCs) in the blood of cancer patients. However, the presence of unwanted leukocytes often leads to false positive results. Here, we evaluated whether the micro-fluidic ParsortixTM technology is appropriate to remove these leukocytes and thereby finally to improve the overall approach.
In this study, we established a workflow including the micro-fluidic ParsortixTM technology for the molecular detection of CTC related transcripts. Background levels of EpCAM, PPIC, TUSC3, and MAL2 were efficiently removed due to an up to 106-fold depletion of leukocytes. The presence of these gene markers was observed in ParsortixTM-enriched blood samples from patients with primary and recurrent gynecological cancer (32% and 14%), as well as in 86% of the metastatic breast cancer samples, at a very high specificity. In the ovarian cancer samples, PPIC was the most prominent gene marker, contributing to all positive cases and at least to 70% of the positive cases after pre-amplification of the respective target genes. Expanding the analytical panel up to 29 gene markers further increased the positivity rate (primary gynecological cancer: 95%, recurrent gynecological cancer: 100%, metastatic breast cancer: 92%).
The established workflow strongly improved the overall molecular analysis of the target cells by the efficient removal of contaminating cells, and, thereby offers great promise for the molecular characterization of CTCs.
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