The five victims of inhalation anthrax first experienced a mild fever, malaise, fatigue, muscle ache, non-productive cough and a sensation of precordial (chest) oppression. Their symptoms were initially interpreted as flu, pneumonia or similar illnesses. The second stage of their infection developed suddenly, with acute shortness of breath, cyanosis bluish-purple skin color denoting severe oxygen deprivation fever, speeded-up pulse and breathing marked by squeaky or whistling sounds in the lungs. That acute stage lasted less than 24 hours, when four of the five patients were dead.
No, this clinical account does not describe how five Americans in Florida, Washington, New York and Connecticut died late last year of inhalation anthrax. Rather, the report, in the December 1960 issue of The American Journal of Medicine, narrates the case histories of the five inhalation victims, plus four cutaneous anthrax patients, under the title: “An epidemic of inhalation anthrax, the first in the twentieth century. Clinical features.” This journal has just reprinted that four-decade-old article in its January 2002 issue.
Its editor-in-chief, Lee Goldman, remarked: “As best we can tell, this series of five cases of inhalation anthrax in 1957 at a goat-hair processing plant in Manchester, N.H., represents not only the first but also the last epidemic of inhalation anthrax in the United States during the 20th century. The editors of The American Journal of Medicine join with [the authors of the original article] and others in hoping that this information will help physicians and the public blunt the threat of bioterrorism.”
Anthrax: From Goat Hair To Grim Envelopes
Now, as the 21st century unfolds, today’s issue of Nature, dated Jan. 24, 2002, carries an article titled, “Structural basis for the activation of anthrax adenylyl cyclase exotoxin by calmodulin.” Its senior author is crystallographer Andrew Bohm, a scientist at the Boston Biomedical Research Institute in Watertown, Mass.
“In the light of the September 11 terror attack and the anthrax threats,” Bohm told BioWorld Today, “I think the significance of this paper is that it potentially opens the door to a new class of anti-anthrax drugs. Right now, anthrax poisoning is treated by antibiotics, which do an outstanding job of killing the bacteria. But paradoxically, once the bacteria have secreted their toxins, the toxins remain lethally active, even after the bacterium is dead. That’s one of the reasons why you still die from anthrax unless you’ve eradicated the bacteria before the disease has passed its critical point.
“Bacillus anthracis, the anthrax bacterium, makes three proteins,” Bohm explained. “Our Nature paper describes exactly how one of these, edema factor, is activated. Protective antigen [PA] is a transporter protein, which by itself is harmless. PA transports the two other two proteins that do the damage. One of these that can hurt you is lethal factor [LF], and the other hurtful one, edema factor [EF]. The first 260 amino acids of LF and EF are very similar. They bind PA, and that makes them targets for transport into cells. Once they’re in, LF is the protease that cuts MAP-kinase-kinase [an intracellular signaling molecule], and does a lot of damage that way.
“Our Nature paper exactly describes the activation of EF and the adenyl cyclase that makes it work,” Bohm went on. “So the idea now is: Okay, we’re going to target the bacterial product inside cells, and with 3-dimensional structures in hand the structures of EF and LF were published recently hopefully, we can go to structure-based drug design, specifically targeting the anthrax toxin.’
“When anthrax spores are inhaled into a human host,” Bohm pointed out, “lethal factor is far more important than edema factor in terms of killing cells and people. LF proteolytically inactivates MAP-kinase-kinase. This eventually leads to septic shock, which is significantly more damaging than edema factor. EF eventually leads to fluid accumulation in the lungs, which is a very serious part of anthrax pathogenesis. And it’s also been shown that cyclic AMP produced by EF helps suppress macrophages, which should be combating the anthrax bacteria.”
Besides releasing fluid into the lungs, edema factor makes lethal factor 10 to 200 times more virulent. EF is harmless until it comes in contact, inside an infected cell, with calmodulin, a ubiquitous protein in the body. When calmodulin binds to EF, it twists that toxin’s structure out of shape, creating a deep narrow pocket that appears to provide an ideal anti-anthrax drug target. That pocket looks easy to block with a small molecule.”
Deep Pockets Suggest Anti-Anthrax Drug
“The interesting thing about that pocket-shaped active site,” Bohm said, “is that it’s quite dissimilar from the pockets seen in other adenosine triphosphate-binding proteins. This pocket,” he continued, “contains a histidine that appears to be working as a catalytic base, something not seen in mammalian proteins. The shape of the pocket is completely different. We think there’s a very good chance that ATP-like molecules, which specifically bind to edema factor, will be found. Some such molecules have already been shown to inhibit the edema factor.” But he added, “What happens in the test tube and in the clinic are totally different things. That doesn’t mean that proteins are drugs.”
His lab is working in this drug-discovery direction.
“It’s fairly slow going,” Bohm allowed. “Basically, we’re well set up to do edema factor crystallography, and we’ve even done some rudimentary chemistry that synthesizes potential therapeutic compounds. But now we’re looking for some industrial cooperation, from companies that have more resources for medicinal chemistry than we do. We’ve spoken with a number of pharmaceutical and biotech firms, but there hasn’t been any strong interest yet. I think the fear is that there really isn’t that much profit in anthrax research. Even with the threat, the only buyers of anti-anthrax drugs are the Army and the government. I suspect the situation will change a little bit now that our Nature paper has come out.
“We are in the process of filing a patent application,” Bohm mentioned. ”It covers the use of the 3-D EF crystal structure to design drugs combating anthrax, and also two other bacterial pathogens which release edema-factor-like toxins. They are Pseudomonas aeruginosa, a highly infectious bacterium in hospitals, and Bordetella pertussis, which causes whooping cough. There’s a good chance,” Bohm concluded, “that these future drugs would be effective from one toxin to another.”