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TITLE OFcirculating
POSTERtumor
IN CAPITALS
EpCAM negative
cells in metastatic
lung cancer
enriched
by filtration
(ARIAL
NARROW
96PT
BOLD)
1, Arjan G.J. Tibbe2, Cees J.M. van Rijn3, T. Jeroen N. Hiltermann4, Harry J.M. Groen4, Leon W.M.M. Terstappen1
Sanne de Wit1*, Guus van Dalum1#, Aufried
Lenferink
FirstT.M.name
last
name authors, underline presenting author
1 Department
of Medical Cell BioPhysics, MIRA institute, University of Twente, Enschede, The Netherlands; 2 VyCAP, Deventer, The Netherlands;
3 University of Wageningen, Wageningen the Netherlands; 4 Department of Pulmonary diseases, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands
Name Chair, MIRA Univesity of Twente (Arial narrow 48pt regular)
Summary
Presence of circulating tumor cells (CTC) in patients with lung cancer is associated with poor survival. The gold standard for the enumeration of
CTC is CellSearch, which uses an enrichment step of cancer cells based on the EpCAM protein present on those cells. The number of CTC found
in lung cancer patients is very low, raising the question whether CTC can be found with a different phenotype that are missed by the CellSearch
system. To investigate this, a device was designed that collects the sample material of individual samples that are discarded by CellSearch after
EpCAM enrichment. EpCAM-positive CTC were isolated using the CellSearch system and EpCAM-negative CTC were isolated from blood using
filtration. Extra cytokeratin (CK) markers were added to the CellSearch system to broaden the coverage of all CK-positive CTC .
Methods
1
Patient data
ASCD
CellSearch
Patient sample
2
In patients, we found EpCAM-positive CTC in CellSearch (A) and in the
CellSearch Waste we found EpCAM-negative CTC (B). The additional CK markers
show that the expression of CK is heterogeneous in the CTC population (CKFITC).
When examining all types
of CTC, the total number of
CTC found in patients
increases (C). However,
there is no correlation
between all types of CTC in
each sample with a
Spearman’s Rho of 0.022
(D). Also, EpCAM-negative
CTC show no association
with survival (E). The CTC in
CellSearch are associated
with poor survival in these
patients, also when the
extra
CK-markers
are
included (F).
A
B
EpCAM+ CTC in CellSearch
EpCAM– CTC in CellSearch Waste
top view
3
D
E
The Automatic Sample Collection Device (ASCD) detects
the presence of blood in the CellSearch Waste and
distributes each sample in a colonical tube (1). After
collection, the blood is passed with constant pressure of
100 mbar through a microsieve containing 5 μm pores
(2). After filtration of the sample, the slide is removed
from the consumable for staining with antibodies to
distinguish between white blood cells (WBC, red) and
cancer cells (green) (3).
C
F
Conclusion
The number of CTC in lung cancer patients is doubled by
expanding the CellSearch assay by filtration of the
discarded blood and the extra cytokeratin markers. The
CellSearch CTC are associated with poor survival, whereas
EpCAM-negative CTC show no association with survival.
Detailed molecular characterization is necessary to
confirm that the EpCAM-negative CTC found after
filtration are indeed tumor cells, and to determine their
difference with the EpCAM-positive CTC.
* Research funded by the EU FP7 CTCTrap Project
# Research funded by Janssen Diagnostics
CTC in patient samples
≥1
≥5
CTC in CellSearch
41%
14%
Patients extra due to
discarded CTC
35%
28%
CTC & discarded CTC
(n=29)
76%
41%
CTC in CellSearch
43%
14%
Patients extra due to
extra CK
11%
4%
CTC & extra CK CTC
(n=28)
54%
18%
All CTC
83%
41%