Isolation and Characterization of Human Monoclonal Antibodies which Neutralize Botulinum Neurotoxin

摘要

To produce neutralizing antibodies to Botulinum neurotoxin type A (boNT/A), single chain Fv (scFv) phage antibody libraries were constructed from the immunoglobulin VH and VL genes of: 1) mice immunized with recombinant binding domain (Hc) of BoNT/A and boosted with BoNT/A; 2) mice immunized with BoNT/A HC; 3) a human immunized with pentavalent BoNT toxoid, and 4) Non-immune humans. A total of 130 unique scFv were selected from the different phage antibody libraries which bound the different Botulinum serotypes. This included 51 scFv from immunized mice, 51 scFv from immunized humans and 28 scFv from the non-immune antibody library. Subsequent characterization focused primarily on those scFv which bound BoNT/A Hc since immunization with this domain has been shown to be protective in mice. A total of 44 unique scFv were identified that bound 15 different epitopes on BoNT/A Hc. scFv binding to each of these epitopes were evaluated in an in vitro toxin neutralization assay. scFv binding three of the epitopes were found to have toxin neutralizing capacity. Two of these neutralize toxin in vivo with enhanced potency when co-administered. The results indicate an approach to therapeutic toxin neutralization.