Johns Hopkins scientists have by chance discovered
that a widely used means of
illuminating cancer cells could undermine studies of the
potential value of experimental
anticancer drugs because the natural "pump" that cells use
to clear out the chemical light
source alters their chemistry.
"Researchers who use markers involving luciferase may
want to double-check their
findings," said Martin Pomper, associate professor of
pharmacology and molecular sciences at Johns
Scientists have increased their use of "glowing"
markers to track cellular activity
in rats and mice, in part, because the tactic is
noninvasive and more humane for the
To make the cells glow, scientists use a gene encoding
luciferase, an enzyme that
causes a chemical reaction responsible for the tiny glow in
transfer the luciferase gene into the genetic machinery of
cancer cells, which then are
injected into an animal, and the enzyme's glow signals the
response of a cell to an
But in a chance discovery, Pomper, collaborating with
John Laterra, professor of
oncology at Johns Hopkins and the Kennedy Krieger
Institute, found that a cellular "pump" that
automatically rids cancer cells of its glowing
contents over time can distort test results. They stumbled
on the saboteur pump during
experiments with a bioluminescent marker in mice that is
designed to test the effect of
an experimental drug on a cancer-causing gene pathway
Pomper's work focused on an anti-hedgehog compound
called HhAntag-691 that his
team hoped would turn off the pathway with a single dose.
The way the study was
designed, his team first bioilluminated the cancer cells
with the luciferase gene, then
introduced HhAntag-691 into the animals. If it worked,
researchers would see no glow in
the cancer cells because the hedgehog pathway would be
switched off, failing to activate
other components in the hedgehog pathway that turn on
"But when we added the anti-hedgehog compound, the
cells glowed brighter rather
than getting dimmer," said Pomper, whose team from the
Johns Hopkins Kimmel Cancer
Center and Russell H. Morgan Department of Radiology
observed a threefold increase in
glow output. The glow is measured by means of a photograph
taken through the animal's
skin with a camera that detects bioluminescent wavelengths.
"We thought this was
bizarre and repeated the experiment many times."
In an effort to figure out their problem, the
researchers repeated the experiment
in a cell extract, a replica of the cell environment
without the intact cell itself. In this
test, the glow dipped threefold. "This suggested that
natural pumps on the surfaces of
intact cancer cells pump out luciferin, which reacts with
intracellular luciferase to cause
the cells to glow," said Pomper, whose report on the
subject appeared in the Oct. 1 issue
of Cancer Research.
Pomper's team identified the chemical pump, called
ABCG2/BCRP, which is one of
nearly 50 of its type. And they warn drug developers to be
wary of it.
"If you want to use bioluminescence to test a drug's
action, make sure the cells
don't have this pump," Pomper said.
The trouble is that many cancers express this pump,
and shutting it off would
complicate the test, according to Pomper. Still, the
discovery has tipped off the
researchers to a new target and a test to find inhibitors
"We're now looking for inhibitors of the pump, which,
when coupled with standard
chemotherapy, could lead to less drug resistance than
currently seen for most tumors,"
Funding for the research was provided by the National
Institutes of Health and
the Dana Foundation.
Other participants in the research are Yimao Zhang,
Joseph P. Bressler, Jeff Neal,
Bachchu Lal and Hyo-Eun C. Bhang, all of Johns Hopkins.