Plant-derived scavengers prowl the body for nerve toxins

Arizona State University ^ | November 23, 2010 | Unknown

[decimon]

The brain is forever chattering to itself, via electrical impulses sent along its hard-wired neuronal "Ethernet." These e-messages are translated into chemical transmissions, allowing communication across the narrow cleft separating one neuron from another or between neurons and their target cells. Of the many kinds of molecules involved in this lively chemical symposium, acetylcholine is among the most critical, performing a host of functions in the central and peripheral nervous system. This delicate cholinergic design however is highly vulnerable. It can fall victim to inadvertent or deliberate poisoning by a class of compounds known as organophosphates—chemicals found in a range of pesticides as well as weaponized nerve agents.

Now Tsafrir Mor, a biochemist in the Center for Infectious Diseases and Vaccinology at the Biodesign Institute at Arizona State University has shown that human butyrylcholinesterase (BChE), a so-called bioscavenging molecule, can be produced synthetically—from plants. Further, Mor and his colleagues have demonstrated the effectiveness of plant-derived BChE in protecting against both pesticide and nerve agent organophosphate poisoning.

The group's research, recently reported in the Proceedings of the National Academy of Science (PNAS), shows promise not only for protecting the nervous system from the effects of organophosphates, but also for gaining a firmer understanding of acetylcholine-linked diseases such as Alzheimer's Dementia and possibly for use against drug overdose and addiction, particularly cocaine. PNAS has selected the important paper as an Editor's Choice.

In the developing world, accidental pesticide poisonings are common. Organophosphate compounds are also the method of choice for many suicides in poor, agricultural regions. The development of far more lethal organophosphates engineered to kill humans has continued apace since Nazi Germany invented them and Cold War adversaries, the United States and the Soviet Union refined and stockpiled these agents.

Following the collapse of the USSR, weaponized organophosphate poisons have proliferated, occasionally falling into the hands of rogue states or terrorist organizations, as these lethal nerve toxins are relatively easy and inexpensive to manufacture and store. The threat of a nerve agent assault on civilians, like the sarin attack in the Tokyo subway system in 1995, perpetrated by the religiously-motivated group Aum Shinrikyo, remains a chilling possibility. The need for effective protection and treatment for organophosphate poisoning is hence a vital concern for public health.

Currently, clinical treatment for exposure to organophosphates involves the use of chemicals like atropine, which can save lives and alleviate acute symptoms, but which fail to address long term neurological effects of such poisoning, which may include muscle weakness, seizures and convulsions, permanent brain defects and social or behavioral symptoms.

Bioscavengers, Mor explains, act as sentries in the body, seeking out and binding with unwanted substances and neutralizing or destroying them. The most heavily studied bioscavengers are the two human cholinesterases—acetylcholinesterase (AChE), which is produced by neurons in the brain and BChE, which is produced mainly by the liver and circulates in blood serum. In addition to their role in defending the body from damaging chemicals, cholinesterases perform a vital housekeeping function, mopping up molecules of acetylcholine, once their signaling tasks are complete.

AChE is a key enzyme bioscavenger that terminates transmission of nerve impulses in the cholinergic synapses of the brain and is also active in the neuromuscular junction, where the axons of motoneurons terminate on muscle cells. As Mor explains, "every time that you move a muscle, the transmission is done through acetylcholine, which is released at the end of the nerve cell and taken up by the receptor on the muscle, causing an influx of ions and contraction of the muscle cell." For this to be accomplished in a coordinated way, the nerve impulse must be cut off almost instantly. This is what the cholinesterases do.

While other neurotransmitters like serotonin are eliminated through reuptake, cholinesterases remove molecules of acetylcholine by hydrolyzing them. Hydrolysis is a chemical reaction in which a given molecule is split into two parts through the addition of a water molecule. AChE is supremely efficient in its catalytic activity, degrading about 25,000 molecules of acetylcholine per second.

Without a means of rapidly getting rid of acetylcholine molecules once they have performed their signaling duty, they flood the nervous system and in sufficient quantity, produce neuromuscular paralysis, and unregulated muscle contraction, eventually causing death due to respiratory and cardiac collapse. This fact, Mor says, makes the system something of an Achilles heel. Many organisms make use of this cholinergic matrix for both offensive and defensive purposes. Plants produce potent anti-cholinesterases to try to thwart herbivory by insects, which in some cases have evolved mechanisms to circumvent such defenses.

Mammals and birds have developed their own mechanisms for dealing with cholinesterase blocking agents. In humans, a particular gene codes for BChE, a closely related analogue of AChE, but one that circulates in blood, laying in wait to scavenge anti-cholinesterase molecules like those of organophosphate poisons. The effectiveness of BChE in neutralizing potentially deadly organophosphates has made it a highly attractive candidate for protecting against the effects of pesticides or nerve agents, as well as mitigating their effects post-exposure. While AChE occurs in the brain and is therefore tricky to acquire, BChE can be readily extracted from blood and stockpiled for future use.

The problem however is finding enough BChE. To protect a few thousand troops on the battlefield from nerve agent poisoning, the entire nation's blood supply would be required. Further, Mor points to many other applications in medicine that would make the production of a sizeable stockpile of BChE highly desirable. In addition to possible treatment for cholinergic ailments, BChE could be used post surgery for patients who lack the naturally occurring enzyme and therefore have difficulty recovering from the effects of anesthesia. There is also evidence that BChE may be useful for treating acute cocaine overdose and possibly as a prophylactic that would eliminate cocaine's euphoric effects, making users less likely to seek out the drug. Again, the challenge is producing the enzyme in sufficient quantity.

The solution Mor and his group have come up with is to use transgenic tobacco plants, modified to synthesize human BChE in their leaves. In a series of experiments outlined in the new paper, Mor's group was able to demonstrate successful protection from both pesticide and nerve agent organophosphate poisoning in two animal models. The team was also able to extend the half-life of the plant-derived BChE, more closely replicating the persistence in the bloodstream of naturally occurring BChE, thereby improving its effectiveness. This was accomplished by decorating the outer portion of the enzyme with Polyethylene glycol (PEG).

Mor stresses that much work remains, before synthetic BChE can be applied as a nerve agent antidote or for other clinical purposes. Currently, the plant-derived BChE acts stoichiometrically, meaning that a molecule of the enzyme is needed for every anti-cholinesterase molecule to be degraded. Future work is aimed at developing forms of the enzyme that can act catalytically against organophosphates, which would permit a far lower effective dose of BChE to be used to protect from poisoning or for treatment post-exposure.

###

This work was funded in part by the National Institutes of Health CounterACT Program through the National Institute of Neurological Disorders and Stroke under a consortium grant awarded to US Army Medical Research Institute of Chemical Defense and contracted to Dr. Mor under a research cooperative agreement. It is a continuation of earlier work originally under support from the Defense Advanced Research Projects Agency (DARPA).

In addition to Dr. Mor's appointment with the Biodesign Institute at Arizona State University he is a professor in the School of Life Sciences.

*Geyer BC, *Kannan L, *Garnaud PE, Broomfield CA, Cadieux CL, *Cherni I, Hodgins SM, Kasten SA, *Kelley K, *Kilbourne J, Oliver ZP, Otto TC, *Puffenberger I, Reeves TE, *Robbins N, 2nd, *Woods RR, Soreq H, Lenz DE, Cerasoli DM, *Mor TS (2010) Plant-derived human butyrylcholinesterase, but not an organophosphorous-compound hydrolyzing variant thereof, protects rodents against nerve agents. Proc Natl Acad Sci U S A, in press (available online at http://www.pnas.org/content/early/2010/11/05/1009021107) .

*Present or former members of Mor Lab at ASU.

[decimon]

Maybe we can replace that atropine injector with a good cheroot.

[decimon]

Ping

[steveo]

Too bad the guy’s first name wasn’t Lester.

[decimon]

Too bad the guy’s first name wasn’t Lester.

It seems that Tsafrir means 'pleasant morning wind or morning sunlight' and Mor means 'myrth.'

[yefragetuwrabrumuy]

This is really important. As far as organophosphate nerve agents are concerned, there is not a whole lot of difference between an insects nervous system and that of a human.

Malathion, a common pesticide that Jerry Brown sprayed on a lot of southern California, is one of the least toxic of these chemicals, as far as *immediate* lethality goes. However, long term effects are unknown.

The acute LD50 (lethal dose for 50% of a group) of Malathion ranges between 1522mg and 1945mg/kg of body weight. To apply that to humans, a dose of 5 oz. would be fatal to a chemically sensitive human of 70kg (154lbs).

Parathion, the agricultural pesticide, invented by IG Farben in the 1940s, is so deadly that crop dusters must wear “space suits” with independent oxygen supplies.

The equivalent toxic dose of Parathion in humans is one tenth of an ounce. Military strength agents are much more potent.

[decimon]

This is not a direct reply, but...

Can’t tell from this news item if the nerve agents can have long-term effects because they are persistent in the body or because they damage they do may become manifest over time. It’s therefore not clear, to me, if this curative must be administered immediately to be of benefit. In any case, this looks like good news.

[yefragetuwrabrumuy]

While they don’t know the answer to that question, they do know that there are long term effects. I knew several men who had been “bitten” by military grade nerve agents, and always had time to listen to anything they had to say on the subject.

The first one had been working “tech escort” on a convoy transporting persistent nerve agent around the US, an infrequent event, to say the least. He unknowingly developed a hole in his suit. Quite abruptly, he showed textbook symptoms, of which there are about a dozen.

At that point, I was all ears, hoping he would wax eloquent.

“I felt really bad. Then they put 10 atropine injectors in my legs. Then I passed out and went into a coma for two weeks. Then I woke up and felt really bad. That’s all.” Just my luck, he was a poet.

The other guy was more interesting. He worked in a lab for a long time with persistent nerve agent, with a low vapor hazard. They had vent fans, wore gloves, goggles, and just surgical masks. If someone’s eyes started to water, they would go outside and smoke a cigarette.

Then they worked with non-persistent agent, with a high vapor hazard. They wore protective masks. Even so, after working with it for a while, his eyes started to water, so he went outside for a smoke.

He took one drag, and then he said “It felt like my lungs contracted to the size of two walnut meats.” He was out cold before he even started to fall down. He concluded by saying that he never smoked another cigarette again, nor wanted to in any way.

[bigheadfred]

Monitor 4.

Watched them dreamily as they sprayed it on the potato fields. Year after year. Asked the pilot one day how you could tell if you got a dose. He said “You just stop breathing”. They never used protective gear. Used it in the big greenhouse. Had a respirator. But absorption through the skin. You could always feel it. Started with a pain in the neck.

Heard it was the bastard child of a Nazi nerve gas.

Not sayin’ I’ve ever been right in the head. Overexposure to all the different crap over the years couldn’t have helped.

http://pesticideinfo.org/Detail_Chemical.jsp?Rec_Id=PC32881

[frithguild]

It seems that Tsafrir means 'pleasant morning wind

In my experience it also means "sure fire way to get kicked out of bed."

[decimon]

Don’t think I’ve encountered any nerve agents. But given the opportunity while in the Army, I messed with a spring-loaded atropine cartridge and injected my finger. No noticeable effect. I also spilled some CN powder on a finger. It burns.

[decimon]

I hear that causes cranial expansion.

[bigheadfred]

I hear that causes cranial expansion.

Extravagantly. But it's just an increase in bone density.

[glorgau]

As I seem to recall, organophosphates were implicated by some in causing “mad cow” prion formation.

Very interesting...

[SunkenCiv]

Thanks decimon.

[neverdem]

Plant-derived human butyrylcholinesterase, but not an organophosphorous-compound hydrolyzing variant thereof, protects rodents against nerve agents

The PDF is a FReebie accessed from the right sidebar of the abstract. It looks like PNAS posted it today. Making access to it today on PNAS is pretty quick for them.

[DoorGunner]

Some ld50s. (TEPP used to be used on hops-prost!)

 

Insecticide	Oral LD50 in Rats (mg/Kg)
Ogranophosphates
TEPP	1.1
Parathion	13
Malathion	1375
Dichlorvos	80
Diazinon	108
Trichlorfon	630
Ronnel	1250
Carbamates
Carbaryl	850
Bagon	83
Mobam	150
Aldicarb	0.8
Zectram	37
Organochlorines
DDT	113
Methoxychlor	6000
Aldrin	39
Dieldrin	46
Lindane	88
Botanicals
Nicotine	10-60
Rotenone	100-300
Pyrethroids	100-300

http://faculty.swosu.edu/scott.long/txcl/pesticid.htm

[yefragetuwrabrumuy]

Science is still terribly ignorant about prions. Apparently, by a normal process, they are created commonly from proteins in the body, and in turn, those proteins convert other proteins into identical prions. And then typically, other natural processes in the body destroy these prions as well. No harm takes place in this process.

But then a deadly prion emerges, formed from a protein that causes a destructive action, and converts other proteins to do the same, *and* is not destroyed by the body.

And for some reason these deadly prions are terribly hard to destroy. Ordinary cooking won’t destroy them. Typically they are neutralized by a combination of enzymes and detergents. Protein-prions are too simple to have genetic material in them.