Isolation and characterization of circulating tumor cells in patients with metastatic colorectal cancer.
OBJECTIVE
Development of targeted therapeutic agents in colorectal cancer (CRC) is impeded by the lack of a noninvasive surrogate of drug effect. This pilot study evaluated the ability of immunomagnetic separation to isolate circulating tumor cells (CTCs) and of the fluorescent microscope system and flow cytometry to enumerate and characterize CTCs from patients with metastatic CRC.


METHODS
Fifty patients with metastatic CRC contributed 50 mL of blood at treatment initiation and disease evaluation timepoints. Fresh tumor specimens were obtained from 17 patients for comparison of circulating and in situ tumor cell characteristics. Epithelial cells were magnetically isolated from whole blood targeting the antiepithelial cell adhesion molecule (EpCAM). Circulating tumor cells were defined as EpCAM isolated, cytokeratin positive, nuclear stain positive, and CD45 negative. Total RNA was isolated from EpCAM-enriched CTCs and multigene reverse-transcriptase polymerase chain reaction analyses were performed.


RESULTS
The median number of CTCs detected by flow cytometry was 2/7.5 mL blood. Mean change in cell count was significantly different for patients with tumor progression versus nonprogression (+6.7 vs. +0.2/7.5 mL; P = 0.001). A correlation was noted between mean fluorescence intensity (flow cytometry) of cytokeratin in CTC and matched tumor specimens (r = 0.79, P = 0.06). Nearly 80% (15 of 19) of samples with >or= 2 CTCs expressed >or= 1 epithelial marker gene (CK19, CK20, carcinoembryonic antigen, or epidermal growth factor receptor).


CONCLUSIONS
Isolating and characterizing CTCs from patients with metastatic CRC is feasible. Change in the CTC number might reflect clinical status, and flow cytometric and gene expression data suggest similarity of circulating and in situ tumor cells. Further evaluation of CTCs for pharmacodynamic and clinical monitoring in patients with CRC is warranted.
Cohen
Steven J
SJ
Department of Medical Oncology, Fox Chase Cancer Center, Philadelphia, PA 19111-2497, USA. s_cohen@fccc.edu
Alpaugh
R Katherine
RK
Gross
Steve
S
O'Hara
Shawn M
SM
Smirnov
Denis A
DA
Terstappen
Leon W M M
LW
Allard
W Jeffrey
WJ
Bilbee
Maryann
M
Cheng
Jonathan D
JD
Hoffman
John P
JP
Lewis
Nancy L
NL
Pellegrino
Ann
A
Rogatko
André
A
Sigurdson
Elin
E
Wang
Hao
H
Watson
James C
JC
Weiner
Louis M
LM
Meropol
Neal J
NJ
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002469:Cell Separation; D015179:Colorectal Neoplasms / Q000097:blood / Q000473:pathology*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D009362:Neoplasm Metastasis; D009360:Neoplastic Cells, Circulating / Q000473:pathology*; D010865:Pilot Projects; D020133:Reverse Transcriptase Polymerase Chain Reaction