Download Plastin-3 Transformed Circulatin Tumor Cells in Hepatic Portal Vein

Circulatin Rare Cells
Application Note
P l a s t i n - 3 Tr a n s f o r m e d C i r c u l a t i n Tu m o r C e l l s i n H e p a t i c P o r t a l
Ve i n o f P a n c r e a t i c C a n c e r P a t i e n t f o r P r e d i c t i n g L i v e r M e t a s t a s i s
Introduction
Materials & Methods
Pancreatic cancer is an asymptomatic disease which
is usually diagnosed during advanced stage with high
incidence of liver metastasis [1].
Evidence of liver metastasis is a principal determinant of
disease management and clinical outcome in pancreatic
cancer patients [2].
Whipple operation is a beneficial surgical procedure for
localized pancreatic cancer, whereas palliative
procedures are recommended for metastatic disease.
Plastin-3, a metastatic-specific gene located on
chromosome Xq23, is an epithelial mesenchymal
transition (EMT) marker associated with colorectal and
breast cancers [3].
Presence of Plastin-3 transformed circulating tumor
cells (CTCs) in hepatic portal vein could serve as a
clinical predictor of liver metastasis.
We applied CytoQuest™ CR positive enrichment system,
EpCAM, PanCK and Plastin-3 monoclonal antibodies to
capture and identify epithelial CTCs in hepatic portal
vein of a pancreatic cancer patient.
Hepatic portal vein blood of pancreatic cancer patient
was collected in Heparin Tube (02-689-6, BD).
7.5 ml blood was prepared for collecting the
peripheral mononuclear cell (PBMC) by density gradient
centrifugation using Leucosep® (163290P, Greiner Bio One) and Histopaque®-1077 (10771, Sigma-Aldrich).
PBMC fraction was harvested and resuspended in Wash
Medium.
Resuspended PBMC was loaded into the CytoQuest™
CR System and CTCs were captured by EpCAM
(KA4586, Abnova) immobilized CytoChipNano (U0095,
Abnova).
Immunofluorescence staining for detecting CTCs was
performed using PanCK, CD45, Plastin-3 (KA4586,
Abnova) and DAPI as the instruction of protocol.
Imaging was performed using Nikon Eclipse Ti-E
fluorescent inverted microscope.
Results
CTC Counts: In 7.5 ml blood of pancreatic cancer patient, 2 cells count as CTC (PanCK+, Plastin3+, CD45-, DAPI+).
Merged
PanCK
Plastin3
CD45
Nucleus
Figure 1: Representative images of CTC (white arrow) and WBCs (yellow arrow) from pancreatic cancer patient, CTC was detected by using immunofluorescence staining for PanCK (FITC, green), Plastin-3
(Alexa647, red) CD45 (PE, orange) and Nucleus (DAPI, blue).
Circulatin Rare Cells
Application Note
Discussions
Effective clinical management of pancreatic cancer
patients depends on a complete medical workup to
determine exact nature of metastasis.
A clinical predictor of liver metastasis would be highly
desirable before Whipple procedure for “localized”
disease to assess the probability of metastasis before
surgery.
Access to the hepatic portal vein for sampling and
detection of EMT-transformed circulating tumor cells
would be a valuable adjunct to the clinical management.
A combination of EpCAM, PanCK and Plastin-3
monoclonal antibodies successfully identify circulating
pancreatic cancer cells using an antibody-based,
positive enrichment.
This case study is the first report of Plastin-3 EMT
expression in pancreatic CTCs identified with EpCAM
and PanCK epithelial cell markers.
A larger cohort study will provide clinically evidence
for establishing Plastin-3 transformed epithelial CTCs
as a predictor of liver metastasis in “localized”
pancreatic cancer patients.
References
1. Berger HG, Rau B, Gansauge F, Poch B, Link KH. Treatment of
pancreatic cancer: challenge of the facts. World journal of surgery,
2003, 27(10):1075-85.
2. Li D, Xie K, Wolff R. Abbruzzese JL Pancreatic cancer, Lancet,
2004, 363(9414):1049-57.
3. Yokobori T, et al. Plastin3 is a novel marker for circulating tumor
cells undergoing the epithelial-mesenchymal transition and is
associated with colorectal cancer prognosis. Cancer research, 2013,
73(7):2059-69.
4. Lyberopoulou A, et al. Mutational analysis of circulating tumor
cells from colorectal cancer patients and correlation with primary
tumor tissue. PloS one, 2015, 10(4):e0123902.
Date of issue January 2016
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