title: Isolation and chimerization of a highly neutralizing antibody conferring passive protection against lethal Bacillus anthracis infection.
abstract: Several studies have demonstrated that the passive transfer of protective antigen (PA)-neutralizing antibodies can protect animals against Bacillus anthracis infection. The standard protocol for the isolation of PA-neutralizing monoclonal antibodies is based upon a primary selection of the highest PA-binders by ELISA, and usually yields only few candidates antibodies. We demonstrated that by applying a PA-neutralization functionality-based screen as the primary criterion for positive clones, it was possible to isolate more than 100 PA-neutralizing antibodies, some of which exhibited no measurable anti-PA titers in ELISA. Among the large panel of neutralizing antibodies identified, mAb 29 demonstrated the most potent activity, and was therefore chimerized. The variable region genes of the mAb 29 were fused to human constant region genes, to form the chimeric 29 antibody (cAb 29). Guinea pigs were fully protected against infection by 40LD(50)B. anthracis spores following two separate administrations with 10 mg/kg of cAb 29: the first administration was given before the challenge, and a second dose was administered on day 4 following exposure. Moreover, animals that survived the challenge and developed endogenous PA-neutralizing antibodies with neutralizing titers above 100 were fully protected against repeat challenges with 40LD(50) of B. anthracis spores. The data presented here emphasize the importance of toxin neutralization-based screens for the efficient isolation of protective antibodies that were probably overlooked in the standard screening protocol. The protective activity of the chimeric cAb 29 demonstrated in this study suggest that it may serve as an effective immunotherapeutic agent against anthrax.
authors.affiliation: Department of Biochemistry, Israel Institute for Biological Research, Ness-Ziona, Israel.
authors.forename: Ronit
authors.initials: R
authors.lastname: Rosenfeld
authors.forename: Hadar
authors.initials: H
authors.lastname: Marcus
authors.forename: Einat
authors.initials: E
authors.lastname: Ben-Arie
authors.forename: Bat-El
authors.initials: BE
authors.lastname: Lachmi
authors.forename: Adva
authors.initials: A
authors.lastname: Mechaly
authors.forename: Shaul
authors.initials: S
authors.lastname: Reuveny
authors.forename: Orit
authors.initials: O
authors.lastname: Gat
authors.forename: Ohad
authors.initials: O
authors.lastname: Mazor
authors.forename: Arie
authors.initials: A
authors.lastname: Ordentlich
mesh_terms: D000818:Animals; D000881:Anthrax / Q000276:immunology / Q000382:microbiology / Q000517:prevention & control*; D000907:Antibodies, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D001408:Bacillus anthracis / Q000276:immunology* / Q000502:physiology; D001483:Base Sequence; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D006168:Guinea Pigs; D007928:Lethal Dose 50; D009500:Neutralization Tests; D016133:Polymerase Chain Reaction; D013171:Spores, Bacterial; D020349:Surface Plasmon Resonance
chemical_list: D000907:Antibodies, Bacterial; D017931:DNA Primers