Limits...
Isolation and chimerization of a highly neutralizing antibody conferring passive protection against lethal Bacillus anthracis infection.

Rosenfeld R, Marcus H, Ben-Arie E, Lachmi BE, Mechaly A, Reuveny S, Gat O, Mazor O, Ordentlich A - PLoS ONE (2009)

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Israel Institute for Biological Research, Ness-Ziona, Israel.

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.

Show MeSH

Related in: MedlinePlus

Protection of guinea pigs against anthrax infection.(A) Female Hartley guinea pigs (n = 6 for each group) were i.m. administered with the indicated doses of cAb 29, followed by s.c. challenge with 40LD50 of B. anthracis Vollum spores 14 hrs later. (B) 30 days after the first challenge, surviving animals from groups 20 and 10+10 mg/kg were re-challenged with the same dose (40LD50), and survival was then monitored for another 14 days.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2710523&req=5

pone-0006351-g006: Protection of guinea pigs against anthrax infection.(A) Female Hartley guinea pigs (n = 6 for each group) were i.m. administered with the indicated doses of cAb 29, followed by s.c. challenge with 40LD50 of B. anthracis Vollum spores 14 hrs later. (B) 30 days after the first challenge, surviving animals from groups 20 and 10+10 mg/kg were re-challenged with the same dose (40LD50), and survival was then monitored for another 14 days.

Mentions: To test the ability of cAb 29 to protect the animals against the anthrax infection, antibody doses of either 5, 10 or 20 mg/kg were i.m. administered 14 hrs prior to challenge with 40LD50 of B. anthracis Vollum spores. Under these challenge conditions, control mice (no treatment) died within 3 days post infection, with MTTD of 2.5 days (Fig. 6A; Table 3). When treated with antibody at a dose of 5 mg/kg, 50% of the animals survived the challenge with a delayed MTTD of 7 days (Fig. 6A; Table 3), significantly longer (P<0.005) than the respective control group value. Increasing the antibody dose to 10 and 20 mg/kg resulted in a minor improvement in animal survival (Fig. 6A; Table 3).


Isolation and chimerization of a highly neutralizing antibody conferring passive protection against lethal Bacillus anthracis infection.

Rosenfeld R, Marcus H, Ben-Arie E, Lachmi BE, Mechaly A, Reuveny S, Gat O, Mazor O, Ordentlich A - PLoS ONE (2009)

Protection of guinea pigs against anthrax infection.(A) Female Hartley guinea pigs (n = 6 for each group) were i.m. administered with the indicated doses of cAb 29, followed by s.c. challenge with 40LD50 of B. anthracis Vollum spores 14 hrs later. (B) 30 days after the first challenge, surviving animals from groups 20 and 10+10 mg/kg were re-challenged with the same dose (40LD50), and survival was then monitored for another 14 days.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2710523&req=5

pone-0006351-g006: Protection of guinea pigs against anthrax infection.(A) Female Hartley guinea pigs (n = 6 for each group) were i.m. administered with the indicated doses of cAb 29, followed by s.c. challenge with 40LD50 of B. anthracis Vollum spores 14 hrs later. (B) 30 days after the first challenge, surviving animals from groups 20 and 10+10 mg/kg were re-challenged with the same dose (40LD50), and survival was then monitored for another 14 days.
Mentions: To test the ability of cAb 29 to protect the animals against the anthrax infection, antibody doses of either 5, 10 or 20 mg/kg were i.m. administered 14 hrs prior to challenge with 40LD50 of B. anthracis Vollum spores. Under these challenge conditions, control mice (no treatment) died within 3 days post infection, with MTTD of 2.5 days (Fig. 6A; Table 3). When treated with antibody at a dose of 5 mg/kg, 50% of the animals survived the challenge with a delayed MTTD of 7 days (Fig. 6A; Table 3), significantly longer (P<0.005) than the respective control group value. Increasing the antibody dose to 10 and 20 mg/kg resulted in a minor improvement in animal survival (Fig. 6A; Table 3).

Bottom Line: 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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Israel Institute for Biological Research, Ness-Ziona, Israel.

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.

Show MeSH
Related in: MedlinePlus