Daniel F. Hayes, MD
Despite being discovered more than 150 years ago, tumor cells present in the blood of patients with cancer are only now inspiring significant research efforts. Technological advancements have allowed the isolation and enrichment of these rare cells and, as potential metastatic “emissaries,” they have significant potential for improving the detection and treatment of advanced and possibly even early-stage disease.
While a plethora of studies have demonstrated prognostic value in the number of circulating tumor cells (CTCs) present in a patient’s bloodstream, the ability to use CTCs to inform clinical decision-making remains elusive. Newer analytical techniques and single-cell molecular profiling are contributing to an improved understanding of CTCs that is already being applied in clinical trials. Leading experts propose that while we aren’t quite there yet, promising data are set to mature in the coming years.
Evolution of CTC Technology
In the mid-1800s, an Australian physician noted “cells identical with those of the cancer” in the blood of a patient with metastatic disease. Scientists at the time postulated that these CTCs, as they came to be known, had broken off of the original tumor and might supply the “seeds” for the metastatic spread of cancer by traveling through the bloodstream and lymphatic system and lodging in capillaries around distant organs.
Further study of CTCs was hindered by the fact that they are so rare—there are typically 1 to 10 CTCs per 10 mL of blood—so it was difficult to isolate them from the mass of other cells in a blood sample. It wasn’t until relatively recently that effective methods were developed to routinely isolate and enrich CTCs from blood samples of patients with cancer and CTCs began to receive wider attention.
Thanks in large part to the development of a wide range of companion analytical technologies for CTCs since the 1980s, there has been an explosion in the number of CTC-related scientific publications. According to Daniel F. Hayes, MD, who has been involved in CTC work for 15 years, at last count there were more than 40 CTC platforms at various stages of development. “Of those, there are only three or four that I think have actually been shown to have analytical validity, that if you run the assay five times you get the same results,” said Hayes, who is the Stuart B. Padnos Professor of Breast Cancer Research at the University of Michigan Comprehensive Cancer Center in Ann Arbor.
Hayes has been involved in the development of CellSearch, a Janssen Diagnostics system that is the only CTC tool that the FDA has approved. He has received research funding from CellSearch manufacturers and has current and pending patents with Janssen for novel CTC technologies.
In order to isolate and enrich CTCs, the majority of analytical technologies use a method known as positive selection, which exploits biological (eg, presence of tumor-associated antigens) or physical (eg, variations in size or density) properties that are specific to CTCs and absent in normal blood cells.
The CellSearch system, for example, takes advantage of the fact that CTCs typically express the epithelial cell–specific antigen—the epithelial cell adhesion molecule (EpCAM). It uses magnetic particles coated with antibodies that bind to Ep- CAM-expressing cells. The cells are then stained with cytokeratin and CD45 antibodies, markers of epithelial and white blood cells, respectively, and a DNA dye. The stained cells are visualized in a chamber and a computer imaging algorithm is used to identify cells that are intact, have a nucleus, and express cytokeratins, but do not express CD45. These cells are classed as CTCs. In recent years, interest has shifted toward the development of CTC technologies that employ socalled negative depletion.
Mehmet Toner, PhD, said in an interview that positive selection has several limitations that drove this shift. “You need to know something on the surface of CTCs that is absent on blood cells so you can positively capture these rare cells, but what we’ve learned over the past 10 years is that these cells are very dynamic in their phenotype,” said Toner, a professor of Surgery (Biomedical Engineering) at the Massachusetts General Hospital (MGH), Harvard Medical School, and founding director of the National Institutes of Health BioMicroElectroMechanical Systems (BioMEMS) Resource Center. “You also need to know something about the cancer behind the CTCs, which limits the utility of the assay in a broader sense.” Negative depletion assays, on the other hand, filter away red blood cells, white blood cells, and platelets, and leave only CTCs behind. “Instead of going after the needle in the haystack, as with positive selection, you get rid of the haystack and leave the needle behind,” Toner explained.