The tuberculosis bacterium requires a specific enzyme
to cause persistent infection, a consortium of researchers at Rockefeller University and three other institutions have found. The discovery
suggests that targeting the enzyme could improve therapies for TB,
which claims more lives each year than any other bacterial infection.
The finding, published in the Aug. 17 issue of the
journal Nature, resulted from a multidisciplinary effort
by investigators at Rockefeller, Albert Einstein College of Medicine,
Washington University in St. Louis and Texas A & M University,
each of whom focused on different aspects of the problem.
The enzyme produced by the bacterium, isocitrate lyase
(ICL), allows the TB microbe Mycobacterium tuberculosis to
use fatty acids as a source of energy. Fatty acids are the most
abundant source of stored energy in the body's cells. The new
findings suggest that these reserves may be exploited by "intracellular"
pathogens like M. tuberculosis, which make their living by
parasitizing the cells of the host. The researchers have shown in
a mouse model of TB that disabling the gene for ICL in M.
tuberculosis crippled the bacterium in the disease's
later, persistent phase.
"If we can block an enzyme that allows M.
tuberculosis to persist, that might provide an avenue for attacking
the bug in its latent phase," says lead author John McKinney,
Ph.D., assistant professor and head of Rockefeller's Laboratory
of Infection Biology. "A drug that targeted persistence would
be quite different from conventional TB drugs, which target processes
required for bacterial growth. ICL is not present in humans, so
blocking its activity should not cause harmful side effects in the
When M. tuberculosis infects the body,
the immune system rallies to fight the invader but is unable to
eradicate the microbe completely. A stalemate is achieved in which
the bacteria continue to live in infected tissue, although their
multiplication is inhibited. The infection can persist in a latent
or chronic state for the lifetime of the host and can flare up if
the immune system is later weakened.
The bug's persistence in the body has been the
main barrier to clearing the TB pathogen with drugs. Current treatments
require three to five drugs for at least six months, a regimen many
patients are unable or unwilling to follow. The drugs can have unpleasant
side effects and in some cases cannot be taken because they interfere
with other medications. Consequently, most patients fail to complete
therapy unless their compliance is closely monitored.
"A drug that inhibited ICL activity or synthesis
might allow us to shorten the duration of chemotherapy from six
months or more to just a few weeks," McKinney says. "That
would have an enormous impact on patient compliance and cure rates."
The multidisciplinary project grew out of research
that McKinney initiated as a postdoctoral fellow in the laboratory
of William R. Jacobs, Ph.D., a bacterial geneticist at the Albert
Einstein College of Medicine and investigator with the Howard Hughes
Medical Institute. Jacobs and McKinney, a 1994 Rockefeller University
graduate, attacked the problem using bacterial genetics and animal
infection models. Co-author David Russell, Ph.D., formerly of Washington
University and now at Cornell University, focused on cell biology
and enzymology. James Sacchettini, Ph.D., of Texas A & M University,
solved the physical structure of ICL using the imaging technique
known as X-ray crystallography.
In the Nature paper, the researchers demonstrated
ICL's link to persistence of the TB bug by infecting mice with
two strains of M. tuberculosis-a "wild-type"
strain containing the gene for ICL and a mutant strain that lacked
the gene. During the early acute phase of infection, growth of the
two strains was identical. Once infection entered the chronic phase,
however, the mutant bacteria were largely eliminated by the immune
system while the wild-type bacteria maintained peak loads in the
animals. Not surprisingly, the mutant bacteria were less virulent.
"We're keeping mice that were infected with the mutant
more than a year ago-they're still alive and vigorous,"
The research suggested that M. tuberculosis
reprograms its carbon metabolism in response to the activation of
certain immune cells-called macrophages-in the host. This
idea was confirmed in subsequent experiments. Russell and colleagues
showed that the mutant bacteria were robust in "resting"
macrophages but were unable to survive in immune-activated macrophages.
The requirement for ICL was correlated with the production of ICL
by the bacteria, which was low in normal macrophages, high in activated
macrophages. These results pointed to a link between the immune
response of the host and the requirement for ICL. To confirm this
link, McKinney and colleagues showed that when the mutant bacteria
were injected into immune-deficient mice, their virulence was restored.
"This is a surprising finding-that the immune
response of the host dictates the basic carbon metabolism of the
pathogen," McKinney says. "I don't know of any precedent
for this in the scientific literature."
A separate paper, published by the same consortium
of investigators in the August issue of Nature Structural Biology,
reveals the three-dimensional structure of ICL. The work was led
by structural biologist James Sacchettini, whose group also determined
the structure of ICL in complex with specific inhibitory molecules
that blocked the ability of the bacteria to use fatty acids as an
energy source. Knowledge of the enzyme's shape is guiding current
efforts by collaborators at Glaxo Wellcome Medicines Research Centre
in Stevenage, UK to identify drugs that will inhibit ICL's
function. These efforts, led by Ken Duncan, Ph.D., have already
paid off handsomely, with several novel ICL inhibitors already identified
and under study.
McKinney says the collaboration among the scientists
at the various institutions was the only way the TB research could
progress so quickly.
"None of us working alone could have gotten
us where we are. Tuberculosis R&D is slow, difficult, and expensive,
and it's not a high priority in the U.S. because it mostly
affects people in developing countries," he says. "Drug
companies are understandably reluctant to invest a lot of money
up front in basic research on TB. But by attacking the problem simultaneously
from many angles, we've made ICL an attractive target, and
in just a few years we've moved from an idea to having leads
that could form the basis of a full drug-discovery program. Considering
that there have been no new drugs developed specifically for TB
since the 1960s, that's not too bad."
TB looms as a major public health issue that is intensifying.
About 16 million people worldwide have active TB, and an estimated
eight million new cases occur each year. Nearly two billion individuals-a
third of the global population-are believed to harbor the infection
in its latent form. In the United States and elsewhere, the spread
of the human immunodeficiency virus (HIV) and the compromised immune
systems that result have greatly increased the number of people
susceptible to the TB germ. A drug that targeted persistent bacteria
could be used as an adjunct to conventional treatments to clear
TB infections much more quickly. That could translate into millions
of lives saved every year.