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Open Access Highly Accessed Research article

Pancreatic cancer circulating tumour cells express a cell motility gene signature that predicts survival after surgery

Gregory Sergeant1, Rudy van Eijsden2, Tania Roskams3, Victor Van Duppen4 and Baki Topal1*

Author Affiliations

1 Department of Abdominal Surgery, University Hospitals Leuven, Herestraat 49, Leuven, 3000, Belgium

2 VIB Nucleomics Core, KU Leuven, Herestraat 49, Leuven, 3000, Belgium

3 Department of Pathology, University Hospitals Leuven, Herestraat 49, Leuven, 3000, Belgium

4 Laboratory of Experimental Hematology, KU Leuven, Herestraat 49, Leuven, 3000, Belgium

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BMC Cancer 2012, 12:527  doi:10.1186/1471-2407-12-527

Published: 16 November 2012

Abstract

Background

Most cancer deaths are caused by metastases, resulting from circulating tumor cells (CTC) that detach from the primary cancer and survive in distant organs. The aim of the present study was to develop a CTC gene signature and to assess its prognostic relevance after surgery for pancreatic ductal adenocarcinoma (PDAC).

Methods

Negative depletion fluorescence activated cell sorting (FACS) was developed and validated with spiking experiments using cancer cell lines in whole human blood samples. This FACS-based method was used to enrich for CTC from the blood of 10 patients who underwent surgery for PDAC. Total RNA was isolated from 4 subgroup samples, i.e. CTC, haematological cells (G), original tumour (T), and non-tumoural pancreatic control tissue (P). After RNA quality control, samples of 6 patients were eligible for further analysis. Whole genome microarray analysis was performed after double linear amplification of RNA. ‘Ingenuity Pathway Analysis’ software and AmiGO were used for functional data analyses. A CTC gene signature was developed and validated with the nCounter system on expression data of 78 primary PDAC using Cox regression analysis for disease-free (DFS) and overall survival (OS).

Results

Using stringent statistical analysis, we retained 8,152 genes to compare expression profiles of CTC vs. other subgroups, and found 1,059 genes to be differentially expressed. The pathway with the highest expression ratio in CTC was p38 mitogen-activated protein kinase (p38 MAPK) signaling, known to be involved in cancer cell migration. In the p38 MAPK pathway, TGF-β1, cPLA2, and MAX were significantly upregulated. In addition, 9 other genes associated with both p38 MAPK signaling and cell motility were overexpressed in CTC. High co-expression of TGF-β1 and our cell motility panel (≥ 4 out of 9 genes for DFS and ≥ 6 out of 9 genes for OS) in primary PDAC was identified as an independent predictor of DFS (p=0.041, HR (95% CI) = 1.885 (1.025 – 3.559)) and OS (p=0.047, HR (95% CI) = 1.366 (1.004 – 1.861)).

Conclusions

Pancreatic CTC isolated from blood samples using FACS-based negative depletion, express a cell motility gene signature. Expression of this newly defined cell motility gene signature in the primary tumour can predict survival of patients undergoing surgical resection for pancreatic cancer.

Trial Registration

Clinical trials.gov NCT00495924

Keywords:
Circulating tumour cells; Pancreatic ductal adenocarcinoma; Gene expression profiling; p38 – MAPK signaling; Transforming growth factor - β1; Cancer cell migration; Cell motility