A pilot study to explore circulating tumour cells in pancreatic cancer as a novel biomarker

Abstract

Background: Obtaining tissue for pancreatic carcinoma diagnosis and biomarker assessment to aid drug development is challenging. Circulating tumour cells (CTCs) may represent a potential biomarker to address these unmet needs. We compared prospectively the utility of two platforms for CTC enumeration and characterisation in pancreatic cancer patients in a pilot exploratory study.

Patients and methods: Blood samples were obtained prospectively from 54 consenting patients and analysed by CellSearch and isolation by size of epithelial tumour cells (ISET). CellSearch exploits immunomagnetic capture of CTCs-expressing epithelial markers, whereas ISET is a marker independent, blood filtration device. Circulating tumour cell expression of epithelial and mesenchymal markers was assessed to explore any discrepancy in CTC number between the two platforms.

Results: ISET detected CTCs in more patients than CellSearch (93% vs 40%) and in higher numbers (median CTCs/7.5 ml, 9 (range 0-240) vs 0 (range 0-144)). Heterogeneity observed for epithelial cell adhesion molecule, pan-cytokeratin (CK), E-Cadherin, Vimentin and CK 7 expression in CTCs may account for discrepancy in CTC number between platforms.

Conclusion: ISET detects more CTCs than CellSearch and offers flexible CTC characterisation with potential to investigate CTC biology and develop biomarkers for pancreatic cancer patient management.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1

CTC detection by CellSearch and ISET. Comparison of CTC numbers in pancreatic cancer patients using the CellSearch and ISET methods. The number of cells per 7.5 ml blood is shown for all recruited patients, 53 patients had CellSearch analysis and 27 had ISET analysis.

Figure 2

PFS and OS for patients without and with CTCs. (A) Progression-free survival of patients without CTCs vs those with CTCs using the CellSearch platform. Median survival (days) 140 vs 94 hazard ratio 0.63. P=0.13 (log rank). (B) Overall survival of patients without CTCs vs those with CTCs using the CellSearch platform. Median survival (days) 164 vs 127 hazard ratio 0.7. P=0.26 (log rank).

Figure 3

Examples of EpCAM and Pan CK C-11 tumour staining. Examples of staining intensity of (A) an EpCAM 3+ tumour, (B) an EpCAM 2+ tumour, (C) an EpCAM 1+ tumour. (D) A CK-positive tumour, (E) a CK weakly positive tumour, (F) a CK-negative tumour.

Figure 4

Examples of differential expression of EpCAM and Pan CK C-11 staining of CTCs. (A) Epithelial cell adhesion molecule-negative staining in patient 60, (B) EpCAM weakly positive staining in patient 54, (C) EpCAM-positive staining in patient 51. (D) C-11-negative staining in patient 51, (E) C-11 weakly positive staining in patient 42, (F) C-11-positive staining in patient 50.

Figure 5

CTCs and CTMs captured on ISET filters. (A–C) Three different patient samples, filtered through ISET membranes (pores are dark circles) and stained with CD45 antibody so that leukocytes can be excluded; these are shown in brown. Circulating tumour cells were counterstained with haematoxylin. (C) A large CTM. The membrane pores are all 8 μm in diameter, which gives scale to these images. (D) A CTM from patient 39 (pancreatic head primary and liver, lung and abodminal wall metastases) demonstrates similar morphology to (E). (E) A haematoxylin and eosin (H&E)-stained tumour block from the same patient photographed at the same magnification.

Figure 6

Differential expression of epithelial and mesenchymal markers. CTMs from patient 50 were stained with (A) Pan CK (C-11), (B) E-Cadherin, (C) and (D) Vimentin, and (E) CK 7. The membrane pores are all 8 μm in diameter, which gives scale to these images.