Abstract 4960: Molecular characterization of in-vivo isolated EpCAM-positive circulating tumor cells in high-risk prostate cancer patients
Authors: Markou Athina N. , Lazaridou Marifili , Paraskevopoulos Panagiotis , Chen Shukun , Kroneis Thomas , Swierczewska Monika , Budna Joanna , Kuske Andra , Gorges Tobias M. , Zabel Maciej , Sedlmayr Peter , Alix-Panabieres Catherine , Pantel Klaus , Lianidou Evi S. 
Published: July 2016

Abstract
Introduction: CTC have been verified as prognostic markers for disease progression in various cancer types. The main aim of the EU project “CTC-SCAN” is to validate the number of CTC isolated from patient's blood as a prognostic marker for relapse in high-risk prostate cancer patients treated with primary radical prostatectomy or radiotherapy. In this study, we present our results on gene expression profiling of CTCs that were isolated, using the CellCollector, a novel clinical device designed for the in vivo isolation of EpCAM-positive CTCs.
Patients and methods: We first developed and validated 3 multiplex and 3 single-plex highly sensitive RT-qPCR assays amplifying:a)Epithelial markers:CK-19,EpCAM,E-CAD & PBGD, b)Stem cell markers:PSCA,ALDH1,CD133& HPRT, c) EMT markers: TWIST, VIM, N-CAD and B2M and d)PSA, e)TMPRSS2-ERG fusion, f)Plastin-3. 62 patients and 36 healthy volunteers participated in the study. After in vivo isolation, total RNA was extracted from captured cells,followed by cDNA synthesis. RT-qPCR was performed for the molecular characterization of captured cells. In all cases, peripheral blood was also collected for CTC analysis by CellSearch and the EPISPOT.
Results: The findings of our study are summarized in Table 1. Briefly, in 13/15(87%) samples, in which at least one cell was detected by CellSearch, we detected the expression of at least one gene. In 28/47 samples, negative by CellSearch, we detected the expression of several genes by the developed RT-qPCR assays. In 9/14 samples that were exclusively found to be positive by EPISPOT for PSA immunospots, at least one of the analyzed genes was also expressed.
Conclusions: In vivo isolation of CTC in combination with a downstream molecular analysis is minimally invasive, and in combination with high specific and sensitive RT-qPCR assays for CTC detection and molecular characterization seems very promising. Comparison studies with the CellSearch and the EPISPOT have shown a higher sensitivity, but a poor agreement. in vivo(n = 18)in vitro(n = 18)GENEPOSITIVE(%)POSITIVE(%)POSITIVE(%)CK-1920(32%)1(0.5%)0(0%)E-CAD0(0%)0(0%)0(0%)EpCAM16(26%)1(0.5%)0(0%)CD1330(0%)0(0%)0(0%)ALDH111(18%)0(0%)0(0%)PSCA0(0%)0(0%)0(0%)VIMENTIN10(16%)0(0%)0(0%)TWIST15(24%)0(0%)0(0%)N-CAD11(18%)0(0%)0(0%)PLASTIN-36(10%)0(0%)0(0%)PSA6(10%)0(0%)0(0%)TMPRSS:ERG0(0%)0(0%)0(0%)CELLSEARCH(cut-off&gt;1cells/ml)15(24%)0(0%)EPISPOT15(24%)0(0%)
Citation Format: Athina N. Markou, Marifili Lazaridou, Panagiotis Paraskevopoulos, Shukun Chen, Thomas Kroneis, Monika Swierczewska, Joanna Budna, Andra Kuske, Tobias M. Gorges, Maciej Zabel, Peter Sedlmayr, Catherine Alix-Panabieres, Klaus Pantel, Evi S. Lianidou. Molecular characterization of in-vivo isolated EpCAM-positive circulating tumor cells in high-risk prostate cancer patients. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4960.