The Receptor Binding Domain of Botulinum Neurotoxin Serotype A (BoNT/A) Inhibits BoNT/A and BoNT/E Intoxications In Vivo

Abstract

The receptor binding domain of botulinum neurotoxin (BoNT), also designated the C terminus of the heavy chain (H C ), is a promising vaccine candidate against botulism. In this study, a highly efficient expression system for the protein was developed in Escherichia coli, which provided yields that were 1 order of magnitude higher than those reported to date (350 mg H C per liter). The product was highly immunogenic, protecting mice from a challenge with 10 5 50% lethal dose (LD 50 ) after a single vaccination and generating a neutralizing titer of 49.98 IU/ml after three immunizations. In addition, a single boost with H C increased neutralizing titers by up to 1 order of magnitude in rabbits hyperimmunized against toxoid. Moreover, we demonstrate here for the first time in vivo inhibition of BoNT/A intoxication by H C /A, presumably due to a blockade of the neurotoxin protein receptor SV2. Administration of H C /A delayed the time to death from 10.4 to 27.3 h in mice exposed to a lethal dose of BoNT/A (P ‫؍‬ 0.0005). Since BoNT/A and BoNT/E partially share SV2 isoforms as their protein receptors, the ability of H C /A to cross-inhibit BoNT/E intoxication was evaluated. The administration of H C /A together with BoNT/E led to 50% survival and significantly delayed the time to death for the nonsurviving mice (P ‫؍‬ 0.003). Furthermore, a combination of H C /A and a subprotective dose of antitoxin E fully protected mice against 850 mouse LD 50 of BoNT/E, suggesting complementary mechanisms of protection consisting of toxin neutralization by antibodies and receptor blocking by H C /A.

Antigenic sites on the HN domain of botulinum neurotoxin A stimulate protective antibody responses against active toxin

“…It is very hard to compare protective activity of anti-H N 519–593 with that of anti-H C Abs. Those studies 10 29 30 31 32 33 measured toxin challenge doses in LD 50 units, whereas the present work used toxin doses in 1.05 × LD 100 units. We strongly believe that a construct comprising the two domains would provide a more efficient vaccine than either alone.…”

Engineering of bacterial toxins for research and medicine

“…Overall, this work shows the potential of recombinant R domains as toxoids and the importance of conformational epitopes in generating a protective antitoxin immune response. It has been confirmed recently by other teams with successful immunization of mice with R domains of botulinum neurotoxins [20,21].…”

A mutated recombinant subunit vaccine protects mice and guinea pigs against botulinum type A intoxication

“…showed that the recombinant HCR of BoNT/A protected mice from BoNT/A intoxication. 22 In fact, the most advanced vaccine candidate for botulinum is a recombinant HCR subunit that is currently undergoing clinical trials. Although recombinant HCRs can protect animals, they can also enter neurons that are similar to BoNT/A via synaptic vesicle protein 2.…”