Anthrax is an infectious disease caused by the bacterium Bacillus anthracis. While the infection is often treatable in its early stages, once the disease has progressed beyond the “point of no return” after just a few days, it can be fatal.
Researchers at the University of Pittsburgh have now found that a cocktail of growth factors reversed would-be lethal cell damage in anthrax-infected mice by reactivating a pathway that is damaged by the bacterial toxin, suggesting that this approach could be adapted for use in human patients. “While only a few people die from anthrax in the United States each year, there is always the concern that the bacterium could be released on a large scale as a bioweapon,” said senior author Shihui Liu, MD, PhD, associate professor of medicine at the Pitt Schol of Medicine and member of the Aging Institute, a joint venture of Pitt and UPMC. “Because the early symptoms of anthrax are nonspecific and flu-like, the disease often isn’t diagnosed until it’s too late for current treatments to help. We need new approaches to treat this later stage of the disease.”
Describing their results in Nature Microbiology (“ERK pathway reactivation prevents anthrax toxin lethality in mice”), Liu and colleagues commented that the growth factor treatment strategy “… provides a means to potentially overcome the ‘point of no return’ for patients who have contracted the often-deadly inhalation anthrax.”
When B. anthracis enters the body through inhalation, ingestion, injection, or contact with skin, it produces proteins that combine to form lethal toxins. “The three components of the anthrax exotoxins: protective antigen (PA), lethal factor (LF), and edema factor (EF) are individually nontoxic, but they can pair to form two toxins: lethal toxin (LT, composed of LF + PA) and edema toxin (ET, composed of EF + PA),” the authors explained. “… LT, the most abundant toxin, is the major virulence factor, playing critical roles during multiple steps of the disease.”
During early infection, anthrax can be treated using antibiotics that eliminate the bacterium or antibodies that neutralize lethal toxin before it enters cells. But once inside cells, the toxin inactivates members of a group of enzymes known as MEKs by cleaving off one of their ends, disrupting important MAPK pathways, and rapidly causing widespread cellular, tissue, and organ damage—and death. “Lethal toxin (LT), the major virulence factor of B. anthracis, proteolytically inactivates MEKs and disables downstream ERK, p38, and JNK pathway signaling leading to tissue damage and mortality,” the investigators noted.
As they further pointed out, anthrax disease progresses rapidly, but the initially nonspecific, flu-like symptoms of inhalational anthrax in particular may mean that patients might not seek medical assistance until the disease has reached an advanced stage. “Mortality usually follows when the host fails to repair toxin-induced cellular damage, the so-called ‘point of no return’ for current therapies,” they noted. “There is currently no therapy available to deal with the cellular, tissue, and organ damage caused by the toxins reaching their molecular targets within susceptible cells.”
To learn more about the roles of MEK-controlled pathways in anthrax toxicity, Liu and colleagues generated mice with modified MEKs that were resistant to being cleaved by lethal toxin. These included MEK1 and MEK2, which control a pathway called ERK involved in cellular division and survival, and MEK3 and MEK6, which regulate the p38 pathway that’s involved in stress-induced defense. “… we sought to define the roles of these pathways in B. anthracis infection using various LF-resistant MEK transgenic mice and sought to explore the therapeutic benefit of MAPK pathway reactivation in reversing anthrax-induced mortality,” they commented.
Their experimental results showed that when exposed to a lethal toxin or to B. anthracis, mice with either modified MEK1/2 or MEK3/6 had much greater survival than normal animals, indicating that anthrax must inactivate both the ERK and p38 pathways to kill its host. “We found that LT inactivation of both the ERK and p38 pathways are essential for anthrax pathogenesis,” they wrote.
The scientists also found that in mice and human cells exposed to lethal toxin or to B. anthracis, a combination of three growth factors—epidermal growth factor (EGF), granulocyte-macrophage colony-stimulating factor (GM-CSF), and basic fibroblast growth factor (FGF2)—which are all individually approved as treatments for other conditions, reactivated the ERK pathway and brought them back from the point of no return. “Remarkably, this regimen reduced the mortality caused by LT challenge, enabling ~60% of the challenged mice to fully recover from the disease, whereas 100% of control mice succumbed to this challenge,” the investigators reported. “Further, this cocktail of growth factors also greatly reduced the mortality of B. anthracis-infected, mice, allowing 70% of the mice to survive the challenge, while only 10% of those without treatment survived.”
Liu added, “Because lethal toxin breaks MEK proteins by clipping off their ends, we thought that this cellular damage was irreversible. So we were really surprised to find that specific growth factors were able to reactivate the ERK pathway and rescue the cell.”
Because different types of cells in the body may require different growth factors to activate ERK, the researchers are now working to optimize a treatment for anthrax in humans. “These findings offer potential towards developing damage-limiting therapeutic strategies for anthrax,” they concluded in their report, suggesting that “These results provide a proof-of-concept that ERK pathway reactivation might be an effective, biologically based therapy to prevent anthrax-induced lethality.”