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FUNGI secrete compounds that kill bacteria, and our exploitation of
such ‘antibiotics’ has transformed the face of modern medicine. But we still
have only a few drugs that are effective against diseases caused by fungi
and protozoa. A newly discovered chemical armoury, evolved by creatures
as diverse as moths and frogs, may provide the solution. They may even be
used ultimately to treat some human cancers.

Some animals produce peptides – small proteins – that can puncture the
membranes of bacterial cells, killing the bacteria in the process. Hans
Boman and his colleagues at the University of Stockholm were pioneers in
this field. They described the defence system of the giant silk moth, Hyalophora
cecropia. Once infected, the moth makes specialised proteins that disrupt
the membranes of bacteria, causing the cells to ‘lyse’, or burst and die.
Researchers have long thought that these ‘lytic’ peptides lyse only bacteria.

Several years ago, my colleagues and I at Louisiana State University
decided to repeat some of this published work. We chemically synthesised
one of a family of lytic peptides, cecropin B, and several of its derivatives
that had minor changes in their sequence of amino acids. As expected, these
peptides killed every type of bacteria we tested. The peptides disrupt the
bacterial membrane, leading to lysis as the cells lose their ability to
control their fluid balance. Electron micrographs show large pores in the
membranes.

We wondered whether the peptides might also kill simple eukaryotic cells,
such as fungi and protozoa. These cells differ from bacteria in having a
membrane-bound nucleus and an outer cell membrane. After all, in nature,
the insects that produce cecropins also risk infection by pathogens other
than bacteria. We did indeed find that the peptides killed yeast cells,
which are eukaryotes. Next, we tested our theory on another eukaryotic cell,
the protozoan that causes malaria, Plasmodium falciparum. Amazingly, we
found that high concentrations of lytic peptides eliminated this parasite
from red blood cells infected in the laboratory. We also eliminated Trypanosoma
cruzi, the protozoan that causes Chagas’ disease, from cells infected in
a laboratory culture.

These experiments suggested that our peptides could kill pathogens that
grow and develop inside human tissue. This type of infection is particularly
difficult to treat because the pathogens are hidden within our own cells.
The peptides, it seemed, could somehow seek out and destroy the pathogens
lurking within. The question was: how? To complicate matters, we had also
shown that the peptides had no effect on normal mammalian cells, even at
high concentrations.

To try to solve this puzzle, my colleagues and I designed a new peptide
in which we altered slightly more than half of the amino acids in cecropin
B’s peptide chain. We deliberately exchanged like for like, replacing the
originals with others that were alike in chemical type. To our surprise,
this new peptide was even more lytic than the natural cecropin B. Because
of its enhanced activity, we nicknamed this artificial peptide Shiva-1,
after the Hindu god of destruction.

At first, we found it difficult to understand how this new peptide could
be more potent than cecropin B. As a rule, a protein’s biological activity
is intimately related to its structure, which depends directly on the sequence
of amino acids that make it up. By altering the original molecule, we should
have rendered it inactive. The fact that this did not happen gave a clue
as to how the peptide lyses cells. The amino acids we exchanged were alike
in chemical type, so the altered peptide retains certain physical properties
of the original molecule. This seems to explain why other natural proteins
are lytic, even though their sequence of amino acids bears no resemblance
to that of the cecropins. Examples include sarcotoxins from the flesh fly
Sarcophaga peregrina and magainins, from the skin of frogs. Evolution has
evidently found more than one way to make a lytic peptide.

So we might be able to design new peptides that lyse only certain kinds
of cells by manipulating their sequence of amino acids. But the question
remains: why are normal mammalian cells resistant to lysis? We suspected
that the answer lies in their highly organised and well-developed cytoskeleton.
The cytoskeleton attaches to the inner surface of the cell membrane at many
places and so maintains the cell’s shape and integrity. By exerting pressure
at its many points of attachment, the cytoskeleton might prevent normal
cells treated with lytic peptides from swelling beyond the bursting point.
We devised a simple experiment to test our hypothesis: we exposed normal
mammalian cells to two chemicals, cytochalasin D and colchicine, that destroy
the cytoskeleton. The chemicals did make the cells sensitive to lysis, so
it looks as if the cytoskeleton is probably the key. Although the membranes
of normal cells suffer some disruption, their cytoskeleton keeps the cells
from bursting.

In the same way, cells with a defective cytoskeleton should become more
sensitive to lytic proteins. Many cancerous cells have an inferior cytoskeleton.
We tested several types of cancerous cell and found that most are killed
by levels of peptide that leave normal cells unscathed. So lytic peptides
may ultimately be useful in treating some human cancers.

Cells infected by viruses may also be more sensitive to lysis. Such
cells fuse together to form giant ‘syncytial’ cells. The fusion causes aberrations
in the cytoskeleton of individual cells. We found that the peptides can
lyse cells infected with several different viruses including herpes simplex
II, the cause of genital herpes, and HIV, which causes AIDS.

Low doses of lytic peptides will also make cells divide more rapidly.
Both skin cells and cells of the immune system seem to respond to peptides
in this manner. It might be possible to design drugs that will enhance the
immune system and help wounds to heal. But we do not yet know how the peptides
might exert such an effect. They might make the cells more permeable to
ions and nutrients, so hastening their growth. Or they might be shaped rather
like proteins designed to promote cell growth; if so, they might interact
directly with membrane receptors to induce cells to divide.

So do we normally produce our own lytic peptides? I decided to look
for them among peptides that function as disrupters of membranes. Some proteins
carry a ‘signal peptide’ that enables them to pass across membranes, moving
through the cell or outside it. Many proteins, for instance, are secreted
into the bloodstream, and each protein of this type carries a signal peptide
that enables it to pass through the membrane barrier. I searched the computerised
database of signal sequences accumulated over the past several years, looking
for sequences that resembled the lytic peptides. Only one sequence matched
the pattern that is essential for the range of activities exerted by our
lytic peptides. This unique signal sequence comes from the human protein
beta-fibrinogen, which plays a key role in the formation of blood clots.

We synthesised the signal sequence from beta-fibrinogen and found that
it is as lytic as the other peptides we made. It also causes cells to proliferate.
Remarkably, it kills cancer cells even better than our other synthetic peptides.
So it may offer us a unique tool with which to fight cancer.

Why does this signal peptide look so much like a lytic peptide? Could
it be functioning as an ancient form of protection in people? It is tempting
to speculate, because beta-fibrinogen is one of the suite of natural substances
known as ‘acute phase’ proteins. These are manufactured especially under
conditions of stress, such as illness or injury. We have not yet found the
beta-fibrinogen signal peptide in human blood; this is rather like looking
for a needle in a haystack. But whatever its function in humans, it may
still point the way to powerful therapeutic compounds. The challenge that
we now face is to find practical applications in fighting diseases in humans.
Only then can we say that lytic peptides are harbingers of a new age in
the treatment of disease.

Dr Jesse Jaynes is in the department of biochemistry and the agricultural
center at the Louisiana State University in Baton Rouge.