快猫短视频

ILYA Raskin finishes the last of his paperwork, gets up and heads out of his
office at Rutgers University in New Jersey. The sun is setting, and he is
heading off to tend his herd. It鈥檚 milking time.

The air is hot and humid in the milking shed. But all is quiet鈥攖here
are no cows and no milking machines, just a long, shallow tank holding row upon
row of small green plants. Here, Raskin has spent the past four years
unravelling one of botany鈥檚 best-kept secrets: 鈥渢ickle鈥� plants with bacteria or
fungi, and you can milk their roots for the chemicals they produce. Tapping the
molecular riches of the radish, lupin or tomato plant, he believes, could answer
the pharmaceuticals industry鈥檚 prayers. A new source of cancer drugs,
bug-busting antibiotics and鈥攊f all goes to plan鈥攁 whole lot more
besides.

Turning plants into medicine chests is already a big money-spinner: around
one quarter of all drugs on the pharmacy shelves started out in fields and
forests. Aspirin, for example, was originally extracted from the bark of the
willow tree, a heart drug called atropine from deadly nightshade, and the
anti-cancer drug taxol from the Pacific yew tree.

Yet, says Raskin, the traditional 鈥渉unter-gatherer鈥� approach to tracking down
new medicines in plants hasn鈥檛 lived up to the expectations of the
pharmaceuticals industry. Part of the problem, he says, is that their technique
is all wrong. 鈥淭he industry has been using ways of extracting the active
ingredients from plants that haven鈥檛 changed for decades.鈥� Bioprospectors uproot
plants from their natural habitat and lug them back to their faraway labs.
There, they crush the leaves and stems, use solvents to extract active
ingredients from a complex mixture of compounds, then analyse them. While this
sounds fairly straightforward, it is costly, time-consuming and frequently
frustrating.

The biggest problem, Raskin believes, is that chemists simply don鈥檛
appreciate how sensitive and complex plant chemistry can be. They use harsh
solvents such as methanol to extract the active components. In many cases, these
solvents鈥攐r the enzymes released from ruptured plant cells鈥攁lter or
smash up the metabolites that the chemists are after, making it difficult to
separate them from other, less useful components. They end up retrieving
unpredictable amounts of the valuable compounds, or losing them altogether.

Exasperated, many have turned to more sophisticated techniques such as
combinatorial chemistry to create the drugs of tomorrow. 鈥淭hey just didn鈥檛
believe that nature did it best,鈥� says Raskin.

But he still believes that plants offer the best route to novel biochemicals.
Under attack, humans can choose to fight or run, but plants are literally rooted
to the spot. When they sense danger, they resort to chemical warfare, defending
themselves with a powerful arsenal of molecular missiles concocted specially for
the occasion.

Under attack

These compounds vary according to the type of threat they face, whether
infection, injury or an extreme change in temperature. Under intense ultraviolet
light, for example, most plants protect their leaves by manufacturing potent
antioxidants called flavonoids. And when attacked by fungi or bacteria, other
plants manufacture antimicrobial chemicals called phytoalexins. Plants, says
Raskin, are chemical chameleons.

So how could he access this amazing chemical factory without grinding up the
plants and washing the pulp with aggressive solvents? The answer, he quickly
realised, lay just out of sight.

Next time you lie on a lawn or walk along the edge of a field of corn, spare
a thought for the incredible subterranean root network that lies beneath. Just
one hectare of rye, for example, can grow a colossal 140 million kilometres of
roots, enough to stretch from the Earth to the Sun. This immense system plays a
number of vital roles. It anchors the plants firmly in the ground, and acts as a
surface through which water and nutrients can be absorbed. So roots must live an
intimate and dangerous existence among hoards of hungry parasites,
root-crunching insects, worms and a host of soil bacteria and fungi. Without the
protection of the thorns, bark or waxy coatings that the aerial parts of a plant
rely on, roots have evolved to live by their chemical wits. They respond to
their enemies by formulating and releasing a variety of specific chemical
鈥渨eapons鈥� to keep the attackers at bay.

In 1996, Raskin and his colleagues were working on phytoremediation鈥攁
technique whereby plants clean up contaminated soil by absorbing chemicals through their roots
(快猫短视频, 20 December 1997, p 26). But
Raskin鈥檚 team began to wonder if the process could be reversed. Could they turn
the roots into tiny chemical factories and somehow control the production
line?

The idea was to trick a plant into thinking that it was under attack from
bacteria or fungi. Then, they hoped, when the plant鈥檚 鈥渋mmune system鈥� responded
to the threat, the root system would release an appropriate cocktail of
chemicals.

Sure enough, testing showed that with the right triggers, plant
roots could be persuaded to produce an amazing array of chemicals. Seeing the
commercial potential of this technique, in 1998 Raskin set up a company called
Photosynthetic Harvest鈥攏ow called Phytomedics.

Raskin quickly realised that the best way to milk plant roots was to grow
them hydroponically鈥攊n a bath of water and nutrients
(see Diagram). It鈥檚
far easier to extract chemicals from water than to get at those dissolved in
toxic organic solvents. To mimic what happens during attack by fungi, for
example, he added chitosan to the water bathing the roots. Chitosan is a
component of the walls of many pathogenic fungi and Raskin discovered that it
makes the ideal 鈥渆licitor鈥� for antifungal compounds. Then, after 24 hours or so,
when the plant had had time to react, the researchers would collect the water
from around their roots and concentrate the chemicals in it by freeze-drying and
chromatography. Finally, screening for active components could begin in
earnest.

Their chitosan elicitation technique really did seem to work. Last year,
Raskin published a paper in the Journal of Experimental Botany (vol 50,
p 1553) showing that chitosan persuaded lupin roots to produce 20 times as much
of an anti-cancer substance, genistein, as they would without the
trigger.

A more general elicitor is salicylic acid. In the late 1980s, Raskin鈥檚 team
found that this compound was a vital part of the plant鈥檚 immune system. When
plants are attacked by viruses, bacteria or fungi, salicylic acid is released at
the site of injury, and circulates throughout the plant to trigger the
production of protective agents (快猫短视频, 19 February, p 24).

Taking fingerprints

Raskin and his colleagues at Phytomedics have now studied over 1600 plant
species, each stimulated with up to four different bacterial and fungal
elicitors, such as derivatives of salicylic acid. For each species they have
produced a unique biochemical map or 鈥渇ingerprint鈥� of the chemicals it secretes,
confirming the technique鈥檚 potential for creating and identifying an enormous
variety of novel chemicals.

Not surprisingly, the drugs industry is more than interested. Phytomedics is
working with the US-based healthcare company Johnson & Johnson,
pharmaceuticals giant American Home Products and the US National Cancer
Institute in a search for novel drugs.

Researchers at the National Cancer Institute have already tested some of
Raskin鈥檚 plant extracts on cancer cells in the lab. Raskin says that he has 24
promising compounds, and Phytomedics is looking for corporate partners to take
the anti-cancer drugs to the next step: clinical trials.

The antimicrobial programme is also looking good. 鈥淲e have at least 20 leads,
but of course it is too early to say if they will make drugs,鈥� says Raskin. Two
of the leads have, however, structures completely unrelated to any existing
antibiotics鈥攁 boon in an age of antibiotic-resistant bacteria. And
researchers at Phytomedics have shown that these two agents are powerful killers
of staphylococcal bacteria, a cause of pneumonia and deadly blood infections.
Raskin also hopes that one of his substances could zap fungal infections.
鈥淔ungal infections are a major killer of people who are immunocompromised,鈥� says
Raskin.

Adrian Parr, of the Food Research Institute in Norwich, has spent years
investigating plant metabolites and believes that Raskin鈥檚 technique looks
promising. 鈥淚t may be of benefit when looking for novel specialist molecules,鈥�
he says. He adds that although plants contain thousands of different chemicals,
only a few hundred may be elicited from their roots. This turns out to be an
advantage in the hunt for novel drugs, since it gives chemists a more manageable
selection to sort through, and even increases their chances of a 鈥渉it鈥�.

But Raskin has even bigger dreams. He believes roots also offer the ideal
collection point for recombinant proteins manufactured inside genetically
modified plants. Although scientists have determined the structure of complex
human proteins鈥攕uch as antibodies and hormones鈥攁nd the genes that
code for them, they have not yet found a simple way to make them in a test tube.
So, most medicinal proteins are made by inserting the relevant gene into living
protein 鈥渇actories鈥� such as bacterial or animal cells, and letting them do the
leg work.

But there鈥檚 a catch. When they鈥檙e produced naturally, proteins undergo
chemical 鈥渢weaking鈥� in cells鈥攖he attachment of sugars or lipids, for
example鈥攖o turn them into their active form. However, different types of
cell modify proteins in different ways. A mammalian protein made inside
bacteria, for example, may not work as well as the real thing, if it works at
all. Worse still, our immune systems might see it as foreign, provoking a
dangerous immune reaction.

By contrast, Raskin claims, plants are perfect protein architects: in many
cases the modifications carried out in plant cells creates a more
鈥渉uman-looking鈥� protein. Add to that the lower cost of purifying proteins from
water rather than alcohol, and secreting recombinant proteins through roots
becomes a highly attractive option. As Raskin likes to put it: 鈥淲hy grind up the
cow to extract the milk?鈥�

As proof of the technique, last year Raskin鈥檚 team published the results of a
study in which they attached a special DNA sequence, called an endoplasmic
reticulum signal peptide, to the genes coding for three 鈥渢est鈥� proteins鈥攁
human placental enzyme, a bacterial enzyme and a fluorescent protein from a
jellyfish. The signal peptide acts as a flag to the plant, telling it to package
up and excrete the protein through the roots.

The genes were engineered into tobacco plants, and Raskin showed that around
80 per cent of the three proteins manufactured by the plant were secreted into
the solution surrounding the roots鈥攁 process that Raskin dubs
鈥渞丑颈锄辞蝉别肠谤别迟颈辞苍鈥�.

On paper, rhizosecretion certainly seems to be cost-effective. Raskin
estimates that making proteins using mammalian cells costs approximately
$5000 per gram, and using bugs costs $500 per gram. But add
purification, removing tannins and pigments, for example, and the final bill can
double or triple, he says. 鈥淲e think we can produce around 100 kilograms of
unpurified protein for each acre of greenhouse space each year, which works out
at around $5 per gram,鈥� says Raskin. Even with purification, the final
cost may be no more than $100 per gram. About a year ago, Raskin began
testing another idea which he hopes could one day complement rhizosecretion:
phyllosecretion. First thing in the morning, the leaves of many plants,
especially those from the tropics, become covered with drops of dew. This is
extracellular fluid that is squeezed out of the plant by water pressure鈥攁
process called guttation鈥攁nd Raskin has found that this dew contains a
variety of plant proteins.

Now he hopes to harness guttation too. Genetically modify the plant and every
morning, Raskin hopes, you could wash your recombinant proteins off the leaves,
much like collecting maple syrup. The results of his initial study have already
been submitted for publication and suggest that it may be possible to collect
high yields of drugs from 鈥渓eaf sweat鈥�. 鈥淚t鈥檚 a very exciting new finding,鈥� he
says.

Bud Ryan, a botanist at Washington State University, agrees that the
technology of rhizo- and phyllosecretion could be cost-effective for those
making recombinant proteins. 鈥淚t is a highly efficient system and could be used
to make proteins in bulk,鈥� he says. Only time will tell if plant-derived
proteins provoke immune reactions. But there should be one important advantage:
proteins derived from other mammals can be infected with viruses that may jump
species. 鈥淧lant-derived proteins could be safer, as there is little risk that
they will be infected with viruses [that can infect humans],鈥� Ryan says.

Whether or not Phytomedics鈥檚 technologies generate the blockbuster drugs of
the future, they return the private life of the root to centre stage. 鈥淭he role
of the chemicals released from roots has been an enigma for ages,鈥� says Parr.
鈥淎t first they were just thought to be waste materials, but we are beginning to
realise that they play a key role in the interface between the root and its
别苍惫颈谤辞苍尘别苍迟.鈥�

For the moment, Raskin is concentrating on antimicrobial drugs鈥攖argeted
at antibiotic-resistant bacteria including Staphylococcus and
Cryptosporidium, and fungi such as Candida, Aspergillus
and Cryptococcus鈥攁s well as anti-cancer drugs. But in the future
he鈥檚 also hoping to hunt for drugs to treat diabetes and cardiovascular
diseases. Raskin鈥檚 work may open doors for environmentally friendly drug
developers, but it has also encouraged botanists to get back to their roots.
鈥淏otany is changing,鈥� says David Tepfer, a plant biologist at INRA at Versailles
in France. 鈥淩oots have always been considered dirty things hidden from
view鈥攂ut they are coming back into fashion.鈥�