|New Targeted Fluorescent-Imaging Compound
Allows Researchers to Detect Viable Cancer Cells in Mice
Researchers have developed a new type of imaging compound that
allows them to visualize viable breast cancer cells that have spread
to the lungs in mice. The compound binds to a protein called HER2,
which is found on the surface of some breast cancer cells, and
it glows, or fluoresces, only when taken inside living cells. This
method of targeting and activation allowed researchers to detect
specific types of live cancer cells in a mouse model of breast
cancer. The study, by researchers at the National Cancer Institute
(NCI), part of the National Institutes of Health, and in Japan,
appeared online Dec. 7, 2008, in Nature Medicine.
Previously developed fluorescent compounds that are activated
inside the body's cells have the limitation that, once they are
turned on, they continue to fluoresce even after they diffuse to
new locations, making it difficult to distinguish viable tumor
cells from normal tissue or dead or damaged tumor cells. The research
team, led by Hisataka Kobayashi, M.D., Ph.D., at the Molecular
Imaging Program of NCI's Center for Cancer Research (CCR), in collaboration
with Yasuteru Urano, Ph.D., at the University of Tokyo, created
an imaging compound that is turned on only when it is inside a
living cell and stops fluorescing when it leaves the cell, as would
happen when the cell dies or becomes damaged. The compound also
can be engineered to target specific types of cancer cells.
"Imaging compounds designed by our concept may have applications
in the clinic," said Kobayashi. "These compounds may allow clinicians
to monitor a patient's response to cancer therapy by allowing them
to visualize whether a drug hits its target and whether hitting
the target leads to shrinkage of the tumor. With additional research
and extensive testing in humans, these compounds may also be adapted
for use in endoscopy procedures and for use as a surgical aid to
improve removal of tumors."
The team created the activatable, cancer-targeting compound by
joining a drug called trastuzumab (Herceptin), which is an antibody
that binds to HER2 and is used to treat HER2-positive breast cancer,
to a modified version of a small fluorescent complex known as BODIPY.
This complex fluoresces only under acidic conditions, such as those
found inside cellular structures called lysosomes, which are sac-like
compartments inside cells that contain enzymes that break down
large molecules the cell does not need. When the activatable BODIPY-antibody
compound encounters a HER2-positive breast cancer cell, the trastuzumab
portion binds to HER2 proteins on the cell's surface, and then
the cell takes the HER2-activatable complex inside. When this complex
is processed inside the cell and enters the acidic environment
of a lysosome, BODIPY becomes activated and fluoresces.
"Our design concept is very versatile and can be used to detect
many types of cancer," said Kobayashi. "Unlike other activatable
fluorescent compounds, our compound consists of a targeting agent
and a fluorescing agent that act independently. We can target the
fluorescing agent to different types of cancer cells by using any
antibody or molecule that is internalized by the targeted cells
after it binds to the cell's surface proteins."
Using a mouse model, the Kobayashi team examined the potential
of the activatable compound for detecting tumors within the body.
They injected either the activatable compound, or a control that
always fluoresces, into the tail of mice that had HER2-positive
breast cancer tumors that had spread to their lungs. One day later,
the investigators found fluorescence from the activatable compound
only in lung tumors, whereas the "always on" control produced fluorescence
in lung tumors, normal lung tissue, and the heart.
To confirm that the activatable compound was primarily processed
by, or specific for, HER2-positive tumor cells, the researchers
induced lung metastases in mice by intravenously injecting both
HER2-positive tumor cells and tumor cells that carried the gene
for red fluorescent protein (RFP) instead of the HER2 gene. After
administering the fluorescence compounds, they found that the activatable
compound produced fluorescence only in the HER2-positive tumors,
whereas the "always on" control produced fluorescence in HER2-positive
tumors and the surrounding tissues as well as the RFP-positive
Of the 472 HER2-positive tumors examined in mice with the activatable
compound, only three showed fluorescence from both the activatable
compound and the RFP (false positive), indicating that the activatable
compound had a 99 percent tumor detection accuracy, or specificity,
for HER2-positive tumors. The "always on" control had a specificity
of less than 85 percent.
The team also confirmed that the activatable compound detects
only living cells. Thirty minutes after they exposed tumor tissues
to alcohol to kill the cells, they found that the fluorescence
of the activatable compound significantly decreased in tumor tissue,
whereas the fluorescence of the "always on" control showed minimal
In another series of experiments, the researchers demonstrated
the versatility of their design concept by linking a BODIPY complex
to a molecule that targets the surface of mouse ovarian cancer
cells. This compound allowed the researchers to detect clusters
of live ovarian cancer cells that had spread to the peritoneum,
or the tissue lining the walls of the abdomen, of mice.
For more information on Dr. Kobayashi's research in NCI's Molecular
Imaging Program, please go to http://ccr.cancer.gov/labs/lab.asp?labid=175.
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and their families, through research into prevention and cancer
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Urano Y., Asanuma D., Hama Y, Koyama Y, Barrett T, Kamiya M, Nagano T, Watanabe T, Hasegawa A, Choyke P.L., and Kobayashi H. Selective molecular imaging of viable cancer cells with pH-activatable fluorescence probes. Nature Medicine. Online December 7, 2008.