Circulating tumor cells are cells that have detached from a primary tumor and circulate in the bloodstream. CTCs may constitute seeds for subsequent growth of additional tumors (metastasis

Metastasis

Metastasis, or metastatic disease , is the spread of a disease from one organ or part to another non-adjacent organ or part. It was previously thought that only malignant tumor cells and infections have the capacity to metastasize; however, this is being reconsidered due to new research...

) in different tissues.

In 1869 Thomas Ashworth obeserved circulating tumor cells in the blood of a man with metastatic cancer. He postulated that “cells identical with those of the cancer itself being seen in the blood may tend to throw some light upon the mode of origin of multiple tumours existing in the same person”. A thorough comparison of the morphology

Morphology (biology)

In biology, morphology is a branch of bioscience dealing with the study of the form and structure of organisms and their specific structural features....

 of the circulating cells to tumor cells from different lesions led Ashworth to conclude that “One thing is certain, that if they [CTC] came from an existing cancer structure, they must have passed through the greater part of the circulatory system to have arrived at the internal saphena vein of the sound leg”. Cancer research has demonstrated the critical role circulating tumor cells play in the metastatic spread of carcinoma

Carcinoma

Carcinoma is the medical term for the most common type of cancer occurring in humans. Put simply, a carcinoma is a cancer that begins in a tissue that lines the inner or outer surfaces of the body, and that generally arises from cells originating in the endodermal or ectodermal germ layer during...

s. Technology with the requisite sensitivity and reproducibility to detect CTC in patients with metastatic disease was developed only recently.

Frequency of CTC

The detection of CTCs may have important prognostic and therapeutic implications but because their numbers can be very small, these cells are not easily detected. Circulating tumor cells are found in frequencies in the order of 1-10 CTC per mL of whole blood in patients with metastatic disease. For comparison, a mL of blood contains a few million white blood cells and a billion red blood cells, see figure 1. This low frequency means that a key component of methods to detect CTC is the enrichment method.
First evidence indicates that CTC markers applied in human medicine are conserved in other species. Five of the more common markers including CK19 are also useful to detect CTC in the blood of dogs with malignant mammary tumors.

Clinical utility

To date, a variety of research methods have been developed to isolate and enumerate CTC. The only U.S. Food and Drug Administration

Food and Drug Administration

The Food and Drug Administration is an agency of the United States Department of Health and Human Services, one of the United States federal executive departments...

 (FDA) cleared methodology for enumeration of CTC in whole blood is the CellSearch system. Extensive clinical testing done using this method shows that presence of CTC is a strong prognostic factor for overall survival

Survival rate

In biostatistics, survival rate is a part of survival analysis, indicating the percentage of people in a study or treatment group who are alive for a given period of time after diagnosis...

 in patients with metastatic breast, colorectal or prostate cancer, see figure 2

CellSearch Method

Blood is sampled in an EDTA tube with an added preservative. Upon arrival in the lab, 7.5mL of blood is centrifuged and placed in a preparation system. This system first enriches the tumor cells immunomagnetically by means of ferrofluidic nanoparticles conjugated to epithelial cell adhesion molecule (EpCAM). Subsequently it stains the sample with a nuclear stain, and fluorescent antibody conjugates against CD45 and cytokeratin 8, 18 and 19 (CK). The sample is scanned on an analyzer, which takes images of the nuclear stain, the cytokeratin stain and the CD45 stain. A trained operator looks at the images of cells to identify those cells that are positive for the nuclear stain and the cytokeratin stain and negative for the CD45 stain. If the cell meets these criteria, the operator judges whether the morphology is suitable for a tumor cell. If the total number of tumor cells found is 5 or more, a sample is positive. In studies done on prostate, breast and colon cancer patients, median survival of metastatic patients with positive samples is about half the median survival of metastatic patients with negative samples.

Morphological definition

Morphological appearance is judged by human operators and is therefore subject to large inter operator variation. Several CTC enumeration methods exist which use morphological appearance to identify CTC, which may also apply different morphological criteria. A recent study in prostate cancer showed that many different morphological definitions of circulating tumor cells have similar prognostic value, even though the absolute number of cells found in patients and normal donors varied by more than a decade between different morphological definitions.

Further characterisation of CTC

Some drugs are particularly effective against cancers which fit certain requirements. For example Herceptin

Trastuzumab

Trastuzumab is a monoclonal antibody that interferes with the HER2/neu receptor.The HER receptors are proteins that are embedded in the cell membrane and communicate molecular signals from outside the cell to inside the cell, and turn genes on and off...

 is very effective in patients who are Her2

HER2/neu

HER-2 also known as proto-oncogene Neu, receptor tyrosine-protein kinase erbB-2, CD340 or p185 is an enzyme that in humans is encoded by the ERBB2 gene. Over expression of this gene is correlated with higher aggressiveness in breast cancers...

 positive, but much less effective in patients who are Her2 negative. Once the primary tumor is removed, biopsy of the current state of the cancer through traditional tissue typing is not possible anymore. Often tissue sections of the primary tumor, removed years prior, are used to do the typing. Further characterisation of CTC may help determining the current tumor phenotype. FISH assays has been performed on CTC to as well as determination of IGF-1R, Her2, Bcl-2

Bcl-2

Bcl-2 is the founding member of the Bcl-2 family of apoptosis regulator proteins encoded by the BCL2 gene. Bcl-2 derives its name from B-cell lymphoma 2, as it is the second member of a range of proteins initially described in chromosomal translocations involving chromosomes 14 and 18 in...

, [ERG (gene)|ERG], PTEN

PTEN (gene)

Phosphatase and tensin homolog is a protein that, in humans, is encoded by the PTEN gene. Mutations of this gene are a step in the development of many cancers....

, AR

Androgen receptor

The androgen receptor , also known as NR3C4 , is a type of nuclear receptor that is activated by binding of either of the androgenic hormones testosterone or dihydrotestosterone in the cytoplasm and then translocating into the nucleus...

 status using immunofluorescence

Immunofluorescence

Immunofluorescence is a technique used for light microscopy with a fluorescence microscope and is used primarily on biological samples. This technique uses the specificity of antibodies to their antigen to target fluorescent dyes to specific biomolecule targets within a cell, and therefore allows...

.