scholarly journals Neutralization of the anthrax toxin by antibody-mediated stapling of its membrane penetrating loop

2021 ◽  
Author(s):  
Fabian Hoelzgen ◽  
Ran Zalk ◽  
Ron Alcalay ◽  
Sagit Cohen Schwartz ◽  
Gianpiero Garau ◽  
...  
Keyword(s):  
Pore Formation ◽  
Protective Antigen ◽  
Neutralizing Antibody ◽  
Anthrax Toxin ◽  
Structural Basis ◽  
Severe Illness ◽  
Central Component ◽  
Important Target ◽  
Anthrax Infection ◽  
Anthrax Vaccines

Anthrax infection is associated with severe illness and high mortality. Protective antigen (PA) is the central component of the anthrax toxin, which is the main virulent factor of anthrax. Upon endocytosis, PA opens a pore in the membranes of endosomes, through which the toxin's cytotoxic enzymes are extruded. The PA pore is formed by a cooperative conformational change where PA's membrane-penetrating loops associate, forming a hydrophobic rim that pierces the membrane. Due to its crucial role in anthrax progression, PA is an important target of monoclonal antibodies-based therapy. cAb29 is a highly effective neutralizing antibody against PA. We determined the cryo-EM structure of PA in complex with the Fab portion of cAb29. We found that cAb29 neutralizes the toxin by clamping the membrane-penetrating loop of PA to a static region on PA's surface, thereby preventing pore formation. Therefore, our results provide the structural basis for the antibody-based neutralization of PA and bring to focus the membrane-penetrating loop of PA as a target for the development of better anti-anthrax vaccines.

scholarly journals Neutralization of the anthrax toxin by antibody-mediated stapling of its membrane-penetrating loop

2021 ◽  
Vol 77 (9) ◽  
Author(s):  
F. Hoelzgen ◽  
R. Zalk ◽  
R. Alcalay ◽  
S. Cohen-Schwartz ◽  
G. Garau ◽  
...  
Keyword(s):  
Pore Formation ◽  
Protective Antigen ◽  
Neutralizing Antibody ◽  
Anthrax Toxin ◽  
Structural Basis ◽  
Severe Illness ◽  
Central Component ◽  
Important Target ◽  
Anthrax Infection ◽  
Anthrax Vaccines

Anthrax infection is associated with severe illness and high mortality. Protective antigen (PA) is the central component of the anthrax toxin, which is one of two major virulence factors of Bacillus anthracis, the causative agent of anthrax disease. Upon endocytosis, PA opens a pore in the membranes of endosomes, through which the cytotoxic enzymes of the toxin are extruded. The PA pore is formed by a cooperative conformational change in which the membrane-penetrating loops of PA associate, forming a hydrophobic rim that pierces the membrane. Due to its crucial role in anthrax progression, PA is an important target for monoclonal antibody-based therapy. cAb29 is a highly effective neutralizing antibody against PA. Here, the cryo-EM structure of PA in complex with the Fab portion of cAb29 was determined. It was found that cAb29 neutralizes the toxin by clamping the membrane-penetrating loop of PA to the static surface-exposed loop of the D3 domain of the same subunit, thereby preventing pore formation. These results provide the structural basis for the antibody-based neutralization of PA and bring into focus the membrane-penetrating loop of PA as a target for the development of better anti-anthrax vaccines.

scholarly journals Combining Anthrax Vaccine and Therapy: a Dominant-Negative Inhibitor of Anthrax Toxin Is Also a Potent and Safe Immunogen for Vaccines

2005 ◽  
Vol 73 (6) ◽  
pp. 3408-3414 ◽  
Author(s):  
Benedikt A. Aulinger ◽  
Michael H. Roehrl ◽  
John J. Mekalanos ◽  
R. John Collier ◽  
Julia Y. Wang
Keyword(s):  
Protective Antigen ◽  
Present Report ◽  
Point Mutations ◽  
Dominant Negative ◽  
Anthrax Toxin ◽  
Central Component ◽  
Endosomal Trafficking ◽  
Dominant Negative Inhibitor ◽  
Antigen Presenting ◽  
Anthrax Vaccines

ABSTRACT Anthrax is caused by the unimpeded growth of Bacillus anthracis in the host and the secretion of toxins. The currently available vaccine is based on protective antigen (PA), a central component of anthrax toxin. Vaccination with PA raises no direct immune response against the bacilli and, being a natural toxin component, PA might be hazardous when used immediately following exposure to B. anthracis. Thus, we have sought to develop a vaccine or therapeutic agent that is safe and eliminates both secreted toxins and bacilli. To that end, we have previously developed a dually active vaccine by conjugating the capsular poly-γ-d-glutamate (PGA) with PA to elicit the production of antibodies specific for both bacilli and toxins. In the present report, we describe the improved potency of anthrax vaccines through the use of a dominant-negative inhibitory (DNI) mutant to replace PA in PA or PA-PGA vaccines. When tested in mice, DNI alone is more immunogenic than PA, and DNI-PGA conjugate elicits significantly higher levels of antibodies against PA and PGA than PA-PGA conjugate. To explain the enhanced immunogenicity of DNI, we propose that the two point mutations in DNI may have improved epitopes of PA allowing better antigen presentation to helper T cells. Alternatively, these mutations may enhance the immunological processing of PA by altering endosomal trafficking of the toxin in antigen-presenting cells. Because DNI has previously been demonstrated to inhibit anthrax toxin, postexposure use of DNI-based vaccines, including conjugate vaccines, may provide improved immunogenicity and therapeutic activity simultaneously.

scholarly journals Efficacy of a Vaccine Based on Protective Antigen and Killed Spores against Experimental Inhalational Anthrax

2008 ◽  
Vol 77 (3) ◽  
pp. 1197-1207 ◽  
Author(s):  
Yves P. Gauthier ◽  
Jean-Nicolas Tournier ◽  
Jean-Charles Paucod ◽  
Jean-Philippe Corre ◽  
Michèle Mock ◽  
...  
Keyword(s):  
Guinea Pigs ◽  
Protective Immunity ◽  
Protective Antigen ◽  
Virulent Strain ◽  
Neutralizing Antibody ◽  
Cutaneous Anthrax ◽  
Anthrax Vaccines ◽  
Mucosal Secretions ◽  
Highly Virulent ◽  
Lethal Doses

ABSTRACTProtective antigen (PA)-based anthrax vaccines acting on toxins are less effective than live attenuated vaccines, suggesting that additional antigens may contribute to protective immunity. Several reports indicate that capsule or spore-associated antigens may enhance the protection afforded by PA. Addition of formaldehyde-inactivated spores (FIS) to PA (PA-FIS) elicits total protection against cutaneous anthrax. Nevertheless, vaccines that are effective against cutaneous anthrax may not be so against inhalational anthrax. The aim of this work was to optimize immunization with PA-FIS and to assess vaccine efficacy against inhalational anthrax. We assessed the immune response to recombinant anthrax PA fromBacillus anthracis(rPA)-FIS administered by various immunization protocols and the protection provided to mice and guinea pigs infected through the respiratory route with spores of a virulent strain ofB. anthracis. Combined subcutaneous plus intranasal immunization of mice yielded a mucosal immunoglobulin G response to rPA that was more than 20 times higher than that in lung mucosal secretions after subcutaneous vaccination. The titers of toxin-neutralizing antibody and antispore antibody were also significantly higher: nine and eight times higher, respectively. The optimized immunization elicited total protection of mice intranasally infected with the virulentB. anthracisstrain 17JB. Guinea pigs were fully protected, both against an intranasal challenge with 100 50% lethal doses (LD50) and against an aerosol with 75 LD50of spores of the highly virulent strain 9602. Conversely, immunization with PA alone did not elicit protection. These results demonstrate that the association of PA and spores is very much more effective than PA alone against experimental inhalational anthrax.

scholarly journals Analysis of Defined Combinations of Monoclonal Antibodies in Anthrax Toxin Neutralization Assays and Their Synergistic Action

2012 ◽  
Vol 19 (5) ◽  
pp. 731-739 ◽  
Author(s):  
Miriam M. Ngundi ◽  
Bruce D. Meade ◽  
Stephen F. Little ◽  
Conrad P. Quinn ◽  
Cindi R. Corbett ◽  
...  
Keyword(s):  
Cell Culture ◽  
Monoclonal Antibodies ◽  
Protective Antigen ◽  
Neutralizing Antibody ◽  
Synergistic Action ◽  
Anthrax Toxin ◽  
Content Type ◽  
Antagonistic Effects ◽  
Antibody Interactions ◽  
New Vaccines

ABSTRACTAntibodies against the protective antigen (PA) component of anthrax toxin play an important role in protection against disease caused byBacillus anthracis. In this study, we examined defined combinations of PA-specific monoclonal antibodies for their ability to neutralize anthrax toxin in cell culture assays. We observed additive, synergistic, and antagonistic effects of the antibodies depending on the specific antibody combination examined and the specific assay used. Synergistic toxin-neutralizing antibody interactions were examined in more detail. We found that one mechanism that can lead to antibody synergy is the bridging of PA monomers by one antibody, with resultant bivalent binding of the second antibody. These results may aid in optimal design of new vaccines and antibody therapies against anthrax.

scholarly journals Structural basis for anthrax toxin receptor 1 recognition by Seneca Valley Virus

2018 ◽  
Vol 115 (46) ◽  
pp. E10934-E10940 ◽  
Author(s):  
Nadishka Jayawardena ◽  
Laura N. Burga ◽  
Richard A. Easingwood ◽  
Yoshimasa Takizawa ◽  
Matthias Wolf ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Protective Antigen ◽  
Therapeutic Option ◽  
Oncolytic Viruses ◽  
Anthrax Toxin ◽  
Bacterial Toxin ◽  
Structural Basis ◽  
Electron Microscopy Analysis ◽  
Toxin Receptor ◽  
Seneca Valley Virus

Recently, the use of oncolytic viruses in cancer therapy has become a realistic therapeutic option. Seneca Valley Virus (SVV) is a newly discovered picornavirus, which has earned a significant reputation as a potent oncolytic agent. Anthrax toxin receptor 1 (ANTXR1), one of the cellular receptors for the protective antigen secreted by Bacillus anthracis, has been identified as the high-affinity cellular receptor for SVV. Here, we report the structure of the SVV-ANTXR1 complex determined by single-particle cryo-electron microscopy analysis at near-atomic resolution. This is an example of a shared receptor structure between a mammalian virus and a bacterial toxin. Our structure shows that ANTXR1 decorates the outer surface of the SVV capsid and interacts with the surface-exposed BC loop and loop II of VP1, “the puff” of VP2 and “the knob” of VP3. Comparison of the receptor-bound capsid structure with the native capsid structure reveals that receptor binding induces minor conformational changes in SVV capsid structure, suggesting the role of ANTXR1 as an attachment receptor. Furthermore, our results demonstrate that the capsid footprint on the receptor is not conserved in anthrax toxin receptor 2 (ANTXR2), thereby providing a molecular mechanism for explaining the exquisite selectivity of SVV for ANTXR1.

scholarly journals Structure of heptameric protective antigen bound to an anthrax toxin receptor: A role for receptor in pH-dependent pore formation

2004 ◽  
Vol 101 (36) ◽  
pp. 13147-13151 ◽  
Author(s):  
D. B. Lacy ◽  
D. J. Wigelsworth ◽  
R. A. Melnyk ◽  
S. C. Harrison ◽  
R. J. Collier
Keyword(s):  
Pore Formation ◽  
Protective Antigen ◽  
Anthrax Toxin ◽  
Toxin Receptor ◽  
Ph Dependent

scholarly journals Comparison of Three Anthrax Toxin Neutralization Assays

2010 ◽  
Vol 17 (6) ◽  
pp. 895-903 ◽  
Author(s):  
Miriam M. Ngundi ◽  
Bruce D. Meade ◽  
Tsai-Lien Lin ◽  
Wei-Jen Tang ◽  
Drusilla L. Burns
Keyword(s):  
Protective Antigen ◽  
Reference Sample ◽  
Quantitative Agreement ◽  
Anthrax Toxin ◽  
Raw 264.7 ◽  
Serum Samples ◽  
Common Reference ◽  
Raw 264.7 Cell ◽  
Antibody Levels ◽  
Anthrax Vaccines

ABSTRACT Different types of anthrax toxin neutralization assays have been utilized to measure the antibody levels elicited by anthrax vaccines in both nonclinical and clinical studies. In the present study, we sought to determine whether three commonly used toxin neutralization assays—J774A.1 cell-, RAW 264.7 cell-, and CHO cell-based assays—yield comparable estimates of neutralization activities for sera obtained after vaccination with anthrax vaccines composed of recombinant protective antigen (rPA). In order to compare the assays, sera were assayed alongside a common reference serum sample and the neutralization titers were expressed relative to the titer for the reference sample in each assay. Analysis of sera from rabbits immunized with multiple doses of the rPA vaccine showed that for later bleeds, the quantitative agreement between the assays was good; however, for early bleeds, some heterogeneity in relative neutralization estimates was observed. Analysis of serum samples from rabbits, nonhuman primates, and humans immunized with the rPA vaccine showed that the relative neutralization estimates obtained in the different assays agreed to various extents, depending on the species of origin of the sera examined. We identified differences in the magnitudes of the Fc receptor-mediated neutralization associated with the J774A.1 cell- and RAW 264.7 cell-based assays, which may account for some of the species dependence of the assays. The differences in the relative neutralization estimates among the assays were relatively small and were always less than 2.5-fold. However, because toxin neutralization assays will likely be used to establish the efficacies of new anthrax vaccines, our findings should be considered when assay outputs are interpreted.

scholarly journals Prophylaxis and Therapy of Inhalational Anthrax by a Novel Monoclonal Antibody to Protective Antigen That Mimics Vaccine-Induced Immunity

2006 ◽  
Vol 74 (10) ◽  
pp. 5840-5847 ◽  
Author(s):  
Laura Vitale ◽  
Diann Blanset ◽  
Israel Lowy ◽  
Thomas O'Neill ◽  
Joel Goldstein ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Protective Antigen ◽  
Fc Receptor ◽  
Anthrax Toxin ◽  
Functional Assay ◽  
Human Mabs ◽  
Anthrax Spores ◽  
Anthrax Vaccines

ABSTRACT The neutralizing antibody response to the protective antigen (PA) component of anthrax toxin elicited by approved anthrax vaccines is an accepted correlate for vaccine-mediated protection against anthrax. We reasoned that a human anti-PA monoclonal antibody (MAb) selected on the basis of superior toxin neutralization activity might provide potent protection against anthrax. The fully human MAb (also referred to as MDX-1303 or Valortim) was chosen from a large panel of anti-PA human MAbs generated using transgenic mice immunized with recombinant PA solely on the basis of in vitro anthrax toxin neutralization. This MAb was effective in prophylactic and postsymptomatic treatment of rabbits exposed to aerosolized anthrax spores, and a single intramuscular injection of 1 mg/kg of body weight fully protected cynomolgus monkeys challenged with aerosolized anthrax spores. Importantly, MAb 1303 defines a novel neutralizing epitope that requires Fc receptor engagement for maximal activity. F(ab′)2 fragments of MAb 1303, which retain equivalent affinity for PA, are 10- to 100-fold less potent in neutralizing anthrax toxin in vitro. Addition of Fc receptor-blocking antibodies also greatly reduced the activity of MAb 1303. Moreover, we found that the neutralizing activity of mouse, rabbit, and human antisera elicited by PA vaccines was effectively abrogated by blocking Fc receptors. Selection of an anti-PA MAb by using a functional assay that is a surrogate for protection has resulted in the identification of a fully human MAb with potent activity in vivo and uncovered a previously unrecognized mechanism of antibody-mediated toxin neutralization that is important for currently used anthrax vaccines.

scholarly journals Anthrax Spores Make an Essential Contribution to Vaccine Efficacy

2002 ◽  
Vol 70 (2) ◽  
pp. 661-664 ◽  
Author(s):  
Fabien Brossier ◽  
Martine Levy ◽  
Michèle Mock
Keyword(s):  
Guinea Pigs ◽  
Protective Antigen ◽  
Humoral Response ◽  
Anthrax Toxin ◽  
Experimental Animals ◽  
Challenge Strain ◽  
Anthrax Spores ◽  
Acellular Vaccines ◽  
Essential Contribution ◽  
Anthrax Vaccines

ABSTRACT Anthrax is caused by Bacillus anthracis, a gram-positive spore-forming bacterium. Septicemia and toxemia rapidly lead to death in infected mammal hosts. Currently used acellular vaccines against anthrax consist of protective antigen (PA), one of the anthrax toxin components. However, in experimental animals such vaccines are less protective than live attenuated strains. Here we demonstrate that the addition of formaldehyde-inactivated spores (FIS) of B. anthracis to PA elicits total protection against challenge with virulent B. anthracis strains in mice and guinea pigs. The toxin-neutralizing activities of sera from mice immunized with PA alone or PA plus FIS were similar, suggesting that the protection conferred by PA plus FIS was not only a consequence of the humoral response to PA. A PA-deficient challenge strain was constructed, and its virulence was due solely to its multiplication. Immunization with FIS alone was sufficient to protect mice partially, and guinea pigs totally, against infection with this strain. This suggests that spore antigens contribute to protection. Guinea pigs and mice had very different susceptibilities to infection with the nontoxigenic strain, highlighting the importance of verifying the pertinence of animal models for evaluating anthrax vaccines.

Sign in / Sign up

Export Citation Format

Share Document