Development of new anthrax vaccines
Although minimizing side effects and reducing the number of doses needed of the current vaccine is beneficial, the ultimate goal is to develop a safe, more effective vaccine that would provide immunity to anthrax within a shorter period of time. To rationally develop a new anthrax vaccine, scientists need to understand how the anthrax vaccine works in humans to elicit an antibody response and provide protection.
For this purpose, Dr. Keitel and colleagues conducted studies to analyze in detail the interactions between PA, the primary immunogenic component of the current vaccine, as well as proposed next-generation anthrax vaccines, and the antibodies produced in humans. The results of this work can guide researchers in the design of more effective anthrax vaccines.
One candidate for a new anthrax vaccine that has been investigated is based on a purified recombinant PA protein (rPA102). The safety and immunogenicity of the rPA120 vaccine at different dosage levels were evaluated in a phase I clinical trial. The conclusions of this study were that the rPA120 vaccine did not cause serious side effects, and that the vaccine was effective in producing an anti-PA antibody response that increased with increasing vaccine dose levels and number of doses. Neutralization activity at the highest rPA120 dosage was similar to that seen in subjects receiving AVA injections.
Another candidate for an anthrax vaccine is composed of two segments of DNA that express PA and lethal factor (LF), two of the three components of anthrax toxins. In a VTEU-supported study, Drs. Keitel and El Sahly and colleagues, evaluated the safety of the DNA-based vaccine in healthy human volunteers and tested whether it could stimulate antibody responses that are associated with protection against infection. They found that the vaccine was generally well tolerated and that a greater percentage of subjects developed antibodies to PA or LF in the groups that received the higher doses of the vaccine.
The researchers further tested the DNA-based anthrax vaccine in nonhuman primates to evaluate immunogenicity and to determine if the vaccine could provide protection against challenge with a lethal dose of B. anthracis spores. Low levels of antibody were detected in the monkeys after vaccination. Importantly, 75% of the animals survived the lethal challenge. They concluded that the vaccine provided immunity at doses that are generally well tolerated by humans.
Dr. Joseph Petrosino and colleagues have been using an approach called functional genomics to identify and study B. anthracis proteins involved in conferring virulence to the bacterium. Two independent virulence plasmids (pXO1 and pXO2) distinguish B. anthracis from other Bacillus species. These plasmids encode 224 genes, including the three anthrax toxin subunits (PA, LF, and EF). The researchers used a novel technique that allowed them to analyze this large group of genes. The virulence plasmid genes were cloned, and their proteins were expressed and purified and used in screens to identify which were immunoreactive with serum from three different anthrax infection animal models, as well as from AVA-vaccinated humans. The anthrax proteins identified in such a screen would be possible candidates for the development of a new generation of anthrax vaccines.
Their study identified ten proteins that were detected by antibodies from all three anthrax animal models (rabbits, mice, and rhesus macaque monkeys); eight of these proteins had not previously been shown to be immunoreactive. When they tested serum obtained from humans who had received the AVA vaccine with the purified proteins, they detected an expected response to PA, but no response with any of the other proteins, suggesting that antibodies elicited by the current vaccine do not react with the proteins they identified using the animal models.
These results indicate that a multi-component subunit vaccine, containing PA and another protein, rather than PA alone, may provide greater protection, because antibodies would be raised to additional targets. Further work is needed to reveal which of the candidates identified in this study could be combined with PA to create an improved vaccine against anthrax.
An additional candidate that could potentially serve as an antigen for an anthrax vaccine is a recombinant heme transporter. Dr. Anthony Maresso has been investigating iron acquisition by B. anthracis. Iron is an essential nutrient used by almost all organisms. Bacterial pathogens must acquire iron in order to grow inside mammalian hosts. The host, however, limits the availability of free iron, thereby providing an effective defense strategy against infection. In response, bacteria have evolved clever ways to subvert host sequestration of iron. Dr. Maresso has uncovered five secreted proteins produced by B. anthracis which specifically bind and transport the body’s prominent iron-carrier molecule, heme. These proteins contain a conserved heme-binding domain referred to as the near-iron transporter (NEAT).
Dr. Maresso, along with Dr. Keitel and others, conducted a study to determine if recombinant NEAT proteins can protect against anthrax disease. They found that vaccination with the NEAT proteins protected mice from B. anthracis infection, and they characterized the immune response elicited by the vaccine in a mouse model of B. anthracis infection. They further obtained the crystal structure of one of the NEAT proteins and identified important functional parts of the protein. These results indicate that NEAT proteins should be considered in the development of an improved vaccine against anthrax and this work can be used to guide this development.
In a separate line of research, Dr. Maresso has identified and studied a group of small molecule inhibitors which specifically target and inactivate enzymes in the pathway of iron acquisition in pathogens like B. anthracis. An understanding of the mechanisms of iron uptake in B. anthracis will allow for the development of therapeutic agents to combat infections by related gram-positive bacteria.
In addition to the administration of the anthrax vaccine for someone who has been exposed to anthrax, treatment for anthrax infections includes use of the antibiotics ciprofloxacin, doxycycline, and penicillin G. The problem is that some strains of the anthrax bacterium are naturally resistant to penicillin, so that penicillin is ineffective in treating patients with these particular strains. Resistance to penicillin is often due to enzymes made by bacteria that are called β-lactamases (these enzymes basically break apart penicillin). One strain of B. anthracis makes two such enzymes (called Bla1 and Bla2). Dr. Timothy Palzkill and his group are studying these enzymes to learn in detail how they allow some strains of B. anthracis to resist antibiotics. Their goal is to develop compounds that can inhibit the action of the β-lactamase enzymes, so that they are no longer effective in blocking the action of antibiotics.