MemoryBlog: Infectious Disease Research in and Around New Orleans

Cached story from here:

The
Road Worth Traveling

By
Fran Simon

fsimon@tulane.edu

Photography
By Jackson Hill

Snow blanketed the wooded 500 acres north of Lake Pontchartrain back in
1963, a rare occurrence in Southeast Louisiana, during construction of
a new research facility for Tulane University. The pristine, white powder was an auspicious sign to set the stage for a new era in Tulane research and a clean slate for the old woods, once the site of a rice plantation that also manufactured bricks.

Originally opened in November 1964 as the Delta Regional Primate Research Center, now the Tulane
National Primate Research Center is the largest of the eight centers in
the National Primate Research Center program funded by the National
Institutes of Health. The Tulane
center houses approximately 5,000 primates of several different
species, most of which live in outdoor breeding colonies.

"Nonhuman primates are limited and precious resources, so scientists at Tulane
and collaborating institutions only use them when other research models
cannot be used," says Andrew Lackner, who became director of the center
in 2001. "Without research in primates there would be no vaccines for
polio or Hepatitis B and advances in our understanding of AIDS would be
greatly delayed. A variety of animal models are used in research but
the primates are particularly useful because of their close genetic and
physiologic relatedness to humans. This is particularly true for
studies of infectious disease where nonhuman primates may be the only
species other than humans susceptible to a particular infectious
diseases."

Located 35
miles north of New Orleans in Covington, the primate center currently
employs 240 people, including 30 doctoral-level scientists. Many of the
staff would not consider working anywhere else--33 employees have been
at the center for at least 20 years.

In 2003, the
center supported research being conducted by 335 scientists from 32
states and 13 countries. This research resulted in 162 publications
with an additional 30 in press. The primate center's grant portfolio of
funding for research projects has tripled since Lackner joined Tulane,
and total funding (including construction) has increased fourfold. In
the current fiscal year, total funding is approaching $40 million.

The primary areas of focus today at the Tulane
National Primate Research Center are infectious diseases, including
biodefenserelated work, gene therapy, reproductive biology and
neuroscience.

THE MISSING LINK IN BIODEFENSE

The Tulane primate center is playing a key role in the federal strategic plan for biodefense
research. The National Institutes of Health has established a
nationwide group of multidisciplinary centers as Regional Centers of
Excellence for Biodefense and Emerging Infectious Diseases with about $350 million over five years. The Tulane primate center is collaborating with scientists at two of the eight biodefense centers, located at the University of Texas Medical Branch at Galveston and at Duke University.

In addition, Tulane
received a $13.6 million competitive grant from the National Institute
of Allergy and Infectious Diseases to build a new regional
biocontainment laboratory. The focus of research in the facility will
be on the development of diagnostics, therapeutics and vaccines for
protection from potential bioterrorist attacks and new, naturally
occurring "emerging" infectious disease. The biocontainment lab will be
a biosafety level-three facility adhering to the most stringent
standards. The Tulane
primate center has safely operated a biosafety level-three laboratory
for more than a decade for work on a variety of infectious diseases.
The center's expertise with infectious disease research using primates
was a significant factor in winning the grant. The Tulane
primate center was the only primate center in the nation to receive
funding for construction of a regional biocontainment laboratory.

Tulane University will invest about $5 million in the project.

"Tulane
has a long history of studying infectious diseases at the primate
center," Lackner says. "Our scientists are among those at the forefront
of this area of research. The infectious diseases program focuses on
HIV/AIDS, malaria, Lyme disease, West Nile virus, tuberculosis,
microsporidiosis and other diseases. These are multidisciplinary
studies involving investigators in multiple divisions at the primate
center and collaborators from elsewhere at Tulane and other institutions."

A BREED APART

"Veterinary medicine is the foundation of everything we do at the primate center, and Tulane
follows the strictest procedures of animal care," says Rudolf Bohm,
chair of veterinary medicine. The research program at the Tulane
primate center has been awarded continuous full accreditation since
1983 by the Association for Assessment and Accreditation of Laboratory
Animal Care International, a private, nonprofit organization that
promotes the responsible treatment of animals in science through a
voluntary accreditation program. To earn and maintain accreditation, a
research program undergoes periodic, extensive assessment and rigorous
evaluations by independent panels of experts from around the world.

"An accredited
program must demonstrate that it is going beyond the minimal
requirements to exhibit excellence in animal care and use," says Bohm,
who supervises nearly 100 veterinarians and support staff. "We develop
improved techniques of animal care and strive to continually improve
the veterinary care for animals here and at other primate centers. We
have derived about 1,000 animals that are free of viruses that are
dangerous to monkeys and which might infect humans, with an $8 million
National Institutes of Health grant.

"In addition
to developing better characterized populations of primates, the
veterinary facility is involved in refining the use of animal models by
developing the use of state-of-the-art diagnostics and surgical
procedures," Bohm says. "In the past five years, we have utilized
minimally invasive surgery widely for a number of infectious disease
and reproductive biology studies. And we commonly utilize more advanced
imaging modalities, including MRI, for both clinical and research use."

A NEW APPROACH

While researchers around the country and at Tulane continue their quest for an effective AIDS vaccine, scientists at the Tulane
primate center are making strides in the fight against HIV infection,
hoping to take a new approach to halting the HIV epidemic. Since the
epidemic began, 60 million people have been infected with HIV worldwide
and an estimated 45 million more people will be infected by 2010,
according to the Joint United Nations Programme on HIV/AIDS. Once
considered a disease that spreads primarily through homosexual sex and
intravenous drug use, AIDS increasingly is striking heterosexuals.

Until
recently, most HIV vaccine research concentrated on the body's immune
system as a whole and HIV transmission through blood. The Tulane
team has shown that major sites for HIV transmission and early viral
replication are the mucosal linings of the body--particularly the
gastrointestinal tract and vagina. Their research focusing on the
mucosal immune system is supported by nine grants, primarily from the
National Institutes of Health, totaling more than $3.5 million for the
current year.

"Women are
much more vulnerable to HIV infection than men during sex, partly
because mucosal surfaces contain many of the cells that the virus
targets, and women have larger mucosal surface areas that can tear to
further facilitate viral infection," says comparative pathologist
Ronald Veazey.

In addition to Veazey, the key members of the Tulane
team that have received these HIVrelated grants are Preston Marx,
Andrew Lackner, Louis Martin and Karol Sestak. They work together on a
variety of projects involving mechanisms of HIV transmission,
pathogenesis and vaccine development.

Veazey and
Marx, who has made major contributions to our understanding about the
origins of HIV, are conducting research into a topical microbicide that
may reduce the risk of transmitting HIV infection from men to women.
"We're working on a molecular barrier, a coating on the cells that
would prevent the virus from 'seeing' the cells," Veazey says. Veazey
and Marx have shown that such a barrier known as a topical
microbicide--a germ-fighting barrier that would be placed in the vagina
before sexual intercourse--prevented SIV infection, the monkey
correlate of HIV, in most of the monkeys in their study. They
co-authored a paper about this "proof of principle" discovery in the
prestigious journal Nature Medicine (2003). This has led to a
tremendous increase in interest in commercial development of a topical
microbicide to prevent HIV transmission.

The National
Institute of Allergy and Infectious Diseases has awarded Veazey and his
team a five-year, $2 million grant for the project "Mechanisms of CD4
Depletion and Proliferation in SIV" (the monkey form of HIV). Veazey
hopes this project will provide greater understanding of how the virus
overcomes the body's immune system. The team also received an
additional four-year grant of more than $1.6 million to study the
pathogenesis of neonatal SIV infection.

In their work
to perfect an effective HIV vaccine, Marx and other scientists at the
primate center are focusing on new strategies to incorporate their
knowledge of the mucosal immune system. Sestak also received a twoyear,
$500,000 grant from the National Institutes of Health to collaborate
with a vaccine company in Sweden and John Clements, professor and chair
of microbiology and immunology at Tulane, who holds a patent on a mucosal adjuvant that has been shown to boost the immune system.

"So far, the
vaccines that are being tested in humans do not target the mucosal
immune system," Lackner says. "These traditional vaccines have not
proven effective. By targeting the mucosal tissues, our hope is to
block HIV infection at an earlier stage."

AN EMERGING DISEASE

Of great
concern to the National Institutes of Health, the Centers for Disease
Control and Prevention, and scientists alike are emerging infectious
diseases--new diseases that will require development of new or improved
treatments and diagnostic procedures. Lyme disease is one to watch.
While the number of humans infected with Lyme disease each year is
relatively smallýabout 20,000 in the United States--the number of new
cases of Lyme disease has been increasing steadily during the last 10
years. Lyme disease is a bacterial infection spread by tick bite that
can lead to arthritis, heart rhythm irregularities and nervous system
abnormalities, including numbness, pain, paralysis of the facial
muscles, meningitis and neurocognitive symptoms. Rare deaths from Lyme
disease have been reported.

In addition to
human infection, each year hundreds of thousands of dogs are bitten by
ticks harboring the spirochete that causes Lyme disease.

Bacteriologist
Mario Philipp and colleagues were the first to develop a monkey model
for Lyme disease that led to the development of a new diagnostic test,
now patented by Tulane.
The U.S. Food and Drug Administration and Department of Agriculture
approved the test, which is more sensitive and specific than previous
tests. It is now the most widely used test for Lyme disease in
veterinary practice.

Philipp and
his team recently received a four-year Centers for Disease Control
grant of more than $1 million and a five-year, $2.25 million grant from
the National Institute of Neurological Disorders and Stroke to continue
studying the pathogenesis of the Lyme disease spirochete in the rhesus
monkey. "Our goal is to advance understanding of the effects of Lyme
disease on the central nervous system by determining the mechanisms of
neural injury using primate brain cells in culture as well as
experiments in vivo," Philipp says.

He believes
the spirochetes that cause Lyme disease produce inflammation that leads
to degeneration and death of neurons in the brain. This research may
have broader applications for diseases such as AIDS dementia,
Parkinson's disease and Alzheimer's disease by explaining the role of
inflammation in killing neurons in the brain.

Lyme disease
is important as an emerging infectious disease as well as a model for
other neurologic disorders. Scientists have demonstrated that the ticks
found in Louisiana are fully competent to transmit Lyme disease
infection, although they appear not to be infected with the Lyme
disease spirochete.

"In doing this
work, we owe a lot to our graduate students and postdoctoral fellows,"
says Philipp, who has trained 17 young scientists in the last 10 years.
"The infrastructure of the primate center, along with highly skilled
veterinary support, enables this multidisciplinary research."

A UNIQUE OPPORTUNITY

While many
degenerative illnesses of the brain are caused by infectious diseases
and are related to aging, others are genetic disorders that afflict
babies who are born with a defective chromosome. Several years ago,
veterinarians at the Tulane
primate center began to notice that animals born in a particular group
of rhesus monkeys in the breeding colony had a devastating neurological
disorder. Some of the afflicted rhesus newborns died within a few days
of birth, while others died before adolescence. Tulane
identified the first and only monkey model in the world of a human
genetic disorder, Krabbe's disease. Though a rare genetic disease, the
monkeys with Krabbe's may one day provide the key to curing many other
inborn errors of metabolism, such as Tay-Sachs disease, in which
children lack vital enzymes to break down their food and eliminate
waste. About one child in every 7,000 born has a lysosomal storage
disorder like Tay-Sachs disease and Krabbe's disease. There are about
40 such disorders.

Bruce Bunnell
heads up a team that is using various approaches of gene therapy and
regenerative medicine to explore potential methods that might cure
Krabbe's disease in the monkeys. If they find a solution, they may help
countless children born every year with these devastating genetic
diseases.

In one
approach, Bunnell and his team are collaborating with Chromos, a
Canadian company exploring a proprietary synthetic chromosome
technology it has developed. Their sights are on the possibility of
replacing the damaged chromosome in the monkeys with Krabbe's disease.
But many questions must be asked--and answered--before attempting gene
therapy in the monkeys.

"The Krabbe's
disease animals are a rare and valuable resource," Bunnell says. The
team is testing different strategies that might be used to deliver
chromosomes into stem cells, tracking expression of the chromosomes to
determine if the systems work. The chromosomes in question are marked
with a chemical that glows green with fluorescence under a microscope.
The team has been able to show that the chromosomes have been
introduced successfully into the target cells. The next hurdles in the
race for a genetic cure are laid out clearly in Bunnell's mind.

"This summer,
we need to continue our bench studies to see if we can make any enzyme,
if it can be expressed at high levels over time, and if the stem cells
can maintain their ability to differentiate into other types of cells
such as brain cells." All this before the team begins to work with
normal animals, to determine if stem cells can be injected into the
monkeys' brains where, the scientists hope, the cells will become brain
cells that glow green with the synthetic chromosome.

"We're in a
unique situation here, with the models, the necessary skills and tools
to explore methods of cellular transplantation, bone-marrow
transplantation and other techniques compared with traditional,
drugbased therapies," Bunnell says. "Ultimately, the solutions probably
will involve combination therapy. It will be many years before human
trials with gene therapy, but right now there is no available therapy
whatsoever for any of the lysosomal storage diseases."

People used to
fret that there was no way to prevent polio, but now polio has been
eradicated. And HIV infection once was considered a death sentence,
while now it is considered a chronic disease that can be managed with a
variety of medications. Much as the Lyme disease work may one day
benefit patients with Parkinson's disease, Bunnell hopes that the
gene-therapy technology his team is building will help Parkinson's
patients re-grow normal brain cells. Hopefully, within his own
lifetime.

A SELF-SUSTAINING POPULATION

The eight
centers in the national primate research program house about 25,000
primates, including about 15,000 rhesus monkeys. About half of the
primate center monkeys in the national program are kept for breeding.
Yet, a government study in 2003 found that it will take up to 10 years
to produce a self-sustaining rhesus population capable of meeting the
demand for research monkeys.

Rhesus
monkeys, the species most commonly used in biomedical research, mate
only during a certain period each year. Each female that becomes
pregnant produces only one offspring, which the rhesus mother nurses
for up to a year. Nature has many ways to foil scientists who are
trying to increase the rhesus population.

Enter
embryologist Hans-Michael Kubisch and the team of scientists who are
using assisted reproductive technology. After coming to Tulane
four years ago, Kubisch has established an in vitro fertilization
system for the rhesus monkeys at the primate center. Kubisch's team
gives carefully chosen female monkeys hormones, to "superovulate" them,
the precious eggs are collected, the scientists place the eggs in a
dish with sperm retrieved from male monkeys, from 70 to 80 percent of
the eggs become fertilized, and the resulting embryos are frozen in
liquid nitrogen when they are two days old. Then, at the opportune
moment, the reproductive technology team implants two embryos in a
rhesus that plays surrogate mother. So far, the team has seen eight
healthy, normal monkeys born from six pregnancies using these
techniques-- including two rare sets of twins.

Now, Kubisch
is shifting focus to more complicated challenges. For the first time
anywhere in the world, a pigtail monkey surrogate mother at the Tulane primate center gave birth to a rhesus.

"This
represents the first cross-species surrogate newborn produced in such
evolutionarily divergent monkeys," Kubisch says.

"The rhesus monkey and the pigtail monkey are thought to be several million years apart on the evolutionary tree."

The advantage?
While rhesus monkeys are ready to carry offspring only at one time each
year, pigtail monkeys breed all year. If the scientists can use pigtail
monkeys as surrogate mothers for rhesus monkeys all year long, the
window of opportunity for producing more rhesus offspring opens wider.

In the future,
Kubisch's team plans to begin work with the monkeys in the Krabbe's
disease colony, with the goal of producing more monkeys with the rare
genetic mutation that might serve as models of Krabbe's disease in
humans.

"We can select
each female only four or five times for super-ovulation. So if we are
able to obtain 20-30 eggs with each attempt, we might obtain 100 eggs
total from each female with the Krabbe's trait," Kubisch says.

While it might
seem that female monkeys are more valuable than the males that are
born, Kubisch says, "I still get excited about every birth that results
from embryo transfer, and I can't help thinking, 'I knew you when you
were just a single cell in a petri dish!'"

Just as each
precious monkey in the national primate research program begins with a
single cell, every research project at the Tulane
National Primate Research Center--the success of which depends on these
very monkeys--takes baby steps on the paths to new discoveries.

Fran Simon is director of public relations for the Tulane University Health Sciences Center.

Tulanian

Summer 2004