Of the Ten Plagues that befell wicked Pharaoh some 35 centuriesago, sixth on the list was the Plague of Boils. As the Bible relates: ". .. a boil breaking forth with blains upon man, and upon beast,throughout all the land of Egypt."
That cutaneous lesion was anthrax, one of the oldest recordedinfections known. Now in our own century, that biblical form of germwarfare figures in the arsenals of several modern military powers.
But anthrax as we know it today is a sudden fatal infection of lungsor intestines _ not merely a carbuncle on the skin with a coal-blackcenter. (The word "anthrax" means "coal" in Greek.)
Now molecular microbiologists at Harvard University School ofMedicine are beating the sword of Bacillus anthracis into ploughshares for improving the safety and efficacy of human vaccinesgenerally. Their paper in today's Proceedings of the NationalAcademy of Sciences (PNAS) bears the title: "Anthrax toxin-mediated delivery of a cytotoxic T-cell epitope in vivo."
Microbiologist/molecular geneticist Michael Starnbach is the article'ssenior author. "Its novelty," he told BioWorld Today, "is that this is anon-toxic-protein way of generating killer T cells, as opposed to anaked-DNA or adjuvant or viral vector approach. And we believethat's what the biotech industry would be interested in applying itfor," he added, "either for human vaccination purposes, or forresearch, to create a vaccine in vitro that can go into an animal modeland stimulate this subset of T lymphocytes."
That CTL subset is the hit-man of the immune system's cellular arm.(The humoral arm's enforcers are antibodies, which patrol the spacesbetween cells.) The Harvard team exploited the expertise of anthraxtoxin at smuggling molecules into cells.
"Those CTL epitopes [antigenic targets] are short peptides,"Starnbach explained. "They serve as markers of whether a cell isinfected with a pathogen, and as a trigger for the immune system torecognize and attack such infected cells."
Starnbach and his co-authors take as their starting material purifiedsequences from the anthrax bacterium's toxin. "B. anthracis' toxin,"he said, "consists of two proteins, PA, standing for protectiveantigen, and LF, its lethality factor."
When the team deleted the C-terminal end of LF's sequence, it lostits toxicity. In place of that lethal fragment, they fused replacementpeptides from another microorganism, Listeria monocytogenes _ aperpetrator in humans of food poisoning.
As reported in today's PNAS, Starnbach and his co-authors, afterextensive in vitro experimentation, injected a mixture of anthrax PAand their truncated LF/Listeria fragment fusion protein into a cohortof mice, and saline into control animals. "It demonstrated," he said,"that the mice made a CTL response against that nine-amino-acidListeria epitope, almost as if they'd been infected with whole liveListeria.
"We were also able to demonstrate that it's an extremely efficientprocess; you can use an enormously small amount of this vaccineproduct _ about 10 nanograms of the LF fusion package is sufficient_ to stimulate this response," he said.
As for the immunization effect, "animals injected with the two-protein vaccine and infected two days later with Listeria, the actualpathogen, had fewer live organisms than the many found in controls.So it appears to have a vaccinating effect." This partial protectioncontinued for at least six months.
Partial, because the vaccine did not confer sterilizing immunity,"completely clearing the animals of the pathogen, and protectingthem against death." That's the Harvard team's next goal; its nexttarget, viral pathogens.
On To Viral Models, Human Vaccines
"We are now pursuing this with pathogens," Starnbach said, "otherthan Listeria, which we really used as a model. We want to seewhether this anthrax fusion technology can be more protective _particularly in viral pathogens _ than what we reported with theListeria."
They now are working with the lymphocytic choriomeningitis RNAvirus, which infects mice, dogs, monkeys and guinea pigs, as well aspeople. "We're considering making some constructs against humanviruses as well," Starnbach allowed, "but are not really at a pointwhere we're ready to discuss those."
He did observe that "Potentially, viral epitopes from HIV,papillomavirus, cytomegalovirus, these kinds of pathogens, could useour system to generate human vaccines, by linking the epitopes ontoit, or the whole proteins from which they are derived."
Bacteria such as Shigella, he suggested, "would probably becandidates for a human vaccine and we're looking into that. But rightnow, most of our testing is in mice, for which Shigella is notpathogenic, so we're kind of limited by that.
"The other aspect of therapeutic vaccines," Starnbach continued, "arethe tumor antigens. Killer T cells not only recognize cells infectedwith various viruses, but also cells that are altered in their own self;that endogenously produce tumor antigens.
"Some of those epitopes," he observed, "have been identified, and wehave the potential to take those, link them onto this system, go backinto an animal, either to prevent establishment of a tumor in general,or as a therapeutic to try to reduce the tumor load, or prevent itsmetastasis."
Anthrax itself is not a candidate for immune protection, becauseeffective vaccines against it have long been available. "One of thefirst vaccines ever developed," Starnbach recalled, "was an anthraxvaccine, made in 1881 by Louis Pasteur. He cured the microorganismof its plasmid that makes the toxin, and injected the toxinlessbacterium into animals. They no longer got infected with virulentanthrax." n
-- David N. Leff Science Editor
(c) 1997 American Health Consultants. All rights reserved.