frogs and HIV

[NicoleRussell]

Frogs may help in fight against HIV
by Leigh MacMillan, Vanderbilt University Medical Center
October 1, 2005


A new weapon in the battle against HIV may come from an unusual source -- a
small tropical frog.


Investigators at Vanderbilt University Medical Center reported this month in
the Journal of Virology that compounds secreted by frog skin are potent
blockers of HIV infection.

The findings could lead to topical treatments for preventing HIV
transmission, and they reinforce the value of preserving the Earth's
biodiversity.

"We need to protect these species long enough for us to understand their
medicinal cabinet," said Louise A. Rollins-Smith, Ph.D., associate professor
of Microbiology & Immunology, who has been studying the antimicrobial
defenses of frogs for about six years. Frogs, she explained, have
specialized granular glands in the skin that produce and store packets of
peptides, small protein-like molecules. In response to skin injury or alarm,
the frog secretes large amounts of these antimicrobial peptides onto the
surface of the skin to combat pathogens like bacteria, fungi and viruses.


Derya Unutmaz, M.D., left, Louise Rollins-Smith, Ph.D., and Scott
VanCompernolle, Ph.D., discovered that compounds made by frogs block HIV
infection. photo by Dana Johnson
Rollins-Smith happens to have the laboratory next door to Derya Unutmaz,
M.D., associate professor of Microbiology & Immunology. During a hallway
chat one day, the two decided it would be interesting to investigate whether
any frog peptides have activity against human viruses, specifically HIV, the
focus of Unutmaz's group.

Postdoctoral fellow Scott E. VanCompernolle, Ph.D., screened 15
antimicrobial peptides from a variety of frog species for their ability to
block HIV infection of T cells, immune system cells targeted by HIV. He
found several that inhibited HIV infection without harming the T cells.

NSF funds Vanderbilt University Medical Center study of amphibian decline
Researchers study frog peptides as anti-microbial agents, including HIV
blockers

Frogs around the world are in trouble. And as species are lost, so are their
biological treasures. The National Science Foundation has awarded a team of
Vanderbilt University Medical Center investigators a four-year grant to
study amphibian declines in Central America and California.

"Amphibian skin has long been favored in folklore for its medicinal
properties," said Louise A. Rollins-Smith, Ph.D., associate professor of
Microbiology & Immunology and principal investigator of the new grant.
"Frogs are a rich source of potentially useful molecules that might work
against human pathogens."

Rollins-Smith collaborated with Derya Unutmaz, M.D., assistant professor of
Microbiology & Immunology, and other Vanderbilt scientists to show this
month that compounds from frog skin block HIV infection.

Frogs produce and secrete compounds called antimicrobial peptides to fight
off bacteria, fungi and viruses that land on their skin, Rollins-Smith
explained.

"Frogs have evolved over millennia to combat such pathogens, so we want to
learn from the frog as much as we can about these molecules," she said.

With the new grant, Rollins-Smith and her team will be investigating the
antimicrobial defenses of declining frog populations that are facing a
particular skin fungus. Postdoctoral fellow Douglas C. Woodhams, Ph.D., will
be traveling to sites in Panama and in California to collect samples of the
skin peptides from affected frogs.

"Our goal is to study frog populations that are ahead of an epidemic of this
fungus, and those that are behind an epidemic to see if the ones that have
survived have beneficial protective peptides," Rollins-Smith said.

The Mass Spectrometry Research Center at Vanderbilt is particularly valuable
to the team's studies. Using mass spectrometry, it is possible to
characterize the array of peptides in the samples and rapidly focus on and
sequence those that might be antimicrobial. "We hope to figure out which
species are most vulnerable to this fungal pathogen so that they can be the
focus of greater conservation efforts," Rollins-Smith said.

The studies may also reveal new antimicrobial peptides which could be useful
blockers of human pathogens, she added.
The peptides appear to selectively kill the virus, perhaps by inserting
themselves into the HIV outer membrane envelope and creating "holes" that
cause the virus particle to fall apart, Unutmaz said.

"We like to call these peptides WMDs -- weapons of membrane destruction,"
Unutmaz said. It is curious that the antimicrobial peptides do not harm the
T cells at concentrations that are effective against the virus, he noted,
since HIV's outer membrane is derived from, and therefore essentially
identical to, the cellular membrane. The investigators have proposed that
the peptides act selectively on the virus in part because of its small size
relative to cells.

The ability of the peptides to destroy HIV was enticing, but to be really
effective as antimicrobial agents, they need to prevent transmission of HIV
from dendritic cells to T cells, Unutmaz said.

Dendritic cells, he explained, are the sentinels of the immune system. They
hang out in the mucosal surface tissues, scanning for invading pathogens.

"Their purpose in life is to capture the enemy, bring it to the lymph
node -- the command center -- and present it to the general, the T cell, to
activate a battle plan," Unutmaz said. "It's a very efficient system that
has allowed us to survive many insults, pathogens and viruses."

But HIV is a wily foe. When it is picked up at the mucosal surface by a
sentinel dendritic cell, it somehow evades destruction. Instead, it hides
inside the cell, waiting to invade the T cell with a Trojan Horse-like
mechanism. The ability of HIV to remain hidden in the dendritic cell,
avoiding destruction by circulating antibodies and immune system cells, "may
explain why after 20 years we don't have a vaccine for this virus," Unutmaz
said.

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To test the effectiveness of the frog peptides in preventing HIV
transmission, VanCompernolle first allowed cultured dendritic cells to
capture active HIV. He then incubated the HIV-harboring dendritic cells with
antimicrobial peptides, washed the peptides away, and added T cells.

"Normally the dendritic cell passes the virus to the T cell, and we get very
efficient infection of the T cell," Unutmaz said. "But when we treated the
dendritic cells with peptides, the virus was gone, completely gone.

"This was a great surprise."

The finding was puzzling, he added, since the prevailing notion is that HIV
captured by dendritic cells is hidden and protected. The investigators
currently are using imaging technologies to test the hypothesis that HIV is
actually cycling to the dendritic cell surface.

"We think maybe it's popping its head out, looking around for a T cell, and
then going back inside to hide until it cycles out again," Unutmaz said. If
peptide is present outside the cell, "it targets the virus that pops up and
kills it." Preliminary experiments suggest that the hypothesis is correct.

"This is very exciting, as it suggests that these peptides could be very
effective since the virus now has nowhere to hide," Unutmaz said. "And if
this cycling is really happening, we may be able to generate a vaccine that
will target virus captured by dendritic cells."

The frog peptides are an exceptional tool for probing "what the virus knows
about the dendritic cell that we don't know," Unutmaz added. "How does HIV
manage to survive and cycle back and forth to the cell membrane? If we can
understand that, we'll find the gaps, and that will open a whole new
universe of targets for intervention."

The investigators learned this week that the American Foundation for AIDS
Research will fund their continuing quest to understand how the frog
peptides kill HIV in dendritic cells. Their plans include imaging how the
peptides work, screening additional frog peptides for activity, and testing
peptides on a mucosal cell system to study the feasibility of developing
them as prophylactics against HIV infection.

"If we are able to learn the mechanisms these peptides are using to kill
HIV, it might be possible to make small chemical molecules that achieve the
same results," Unutmaz said. Such chemicals would be more practical as
therapeutic microbicides, he said. "This study is a great example of how
collaboration across disciplines leads to big discoveries," Unutmaz said.

Other members of the Department of Microbiology and Immunology assisted the
investigators by providing viruses for testing. The team found that
membrane-coated viruses were susceptible to destruction by the frog
peptides, but non-coated viruses, such as reovirus and adenovirus, were not
affected.

R. Jeffery Taylor, Ph.D., Kyra Oswald-Richter, Ph.D., Jiyang Jiang, Ph.D.,
Bryan E Youree, M.D., Christopher R. Aiken, Ph.D., and Terence S. Dermody,
M.D., at Vanderbilt are co-authors of the study. The research was supported
by the National Institutes of Health, the Elizabeth B. Lamb Center for
Pediatric Research, and the National Science Foundation.


This is an adapted press release from the Vanderbilt University Medical
Center. The original version appears at Frogs may aid in HIV fight: study.