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Vaccine Detail

BoNT/A(Hc)
Vaccine Information
  • Vaccine Name: BoNT/A(Hc)
  • Target Pathogen: Clostridium botulinum
  • Target Disease: Botulism
  • Vaccine Ontology ID: VO_0004074
  • Type: Subunit vaccine
  • SCFV single-chain Fv fragment gene engineering:
    • Type: Protein
    • Description:
    • Detailed Gene Information: Click Here.
  • Adjuvant: Alhydrogel vaccine adjuvant
    • Adjuvant name: Alhydrogel vaccine adjuvant
    • VO adjuvant ID: VO_0001241
    • Description: The botulinum neurotoxins (BoNT) are the causative agents of botulism and represent a family of seven structurally similar but antigenically distinct serotypes (A to G). These toxins exert their action by blocking release of the neurotransmitter acetylcholine at the neuromuscular junction. BoNT are usually expressed in as a single polypeptide chain and then post-translationally nicked, forming a dichain consisting of a 100-kDa heavy chain and a 50-kDa light chain held together by a single disulfide bond. Topologically, these neurotoxins are composed of three domains: binding, translocation, and catalytic, each of which is believed to play a role in intoxication. The carboxy-terminal portion of the heavy chain is responsible for binding nerve cell receptor(s). After toxin binding, it is thought to be internalized into an endosome through receptor-mediated endocytosis. It is believed that the 50-kDa amino-terminal domain of the heavy chain possesses channel-forming capabilities when in the acidic environment of the endosome, allowing internalization of the toxin. The final step in the mechanism involves zinc-dependent proteolysis by the catalytic domain of key cytosolic substrates necessary for neurotransmitter release (Byrne et al., 1998).
  • Preparation: After fermentation and cell disruption, BoNT/A(Hc) was purified by using a three-step chromatographic process consisting of expanded-bed chromatography, Mono S cation-exchange chromatography, and hydrophobic interaction chromatography (Byrne et al., 1998).
  • Virulence: (Byrne et al., 1998)
  • Description: The botulinum neurotoxins (BoNT) are the causative agents of botulism and represent a family of seven structurally similar but antigenically distinct serotypes (A to G). These toxins exert their action by blocking release of the neurotransmitter acetylcholine at the neuromuscular junction. BoNT are usually expressed in as a single polypeptide chain and then post-translationally nicked, forming a dichain consisting of a 100-kDa heavy chain and a 50-kDa light chain held together by a single disulfide bond. Topologically, these neurotoxins are composed of three domains: binding, translocation, and catalytic, each of which is believed to play a role in intoxication. The carboxy-terminal portion of the heavy chain is responsible for binding nerve cell receptor(s). After toxin binding, it is thought to be internalized into an endosome through receptor-mediated endocytosis. It is believed that the 50-kDa amino-terminal domain of the heavy chain possesses channel-forming capabilities when in the acidic environment of the endosome, allowing internalization of the toxin. The final step in the mechanism involves zinc-dependent proteolysis by the catalytic domain of key cytosolic substrates necessary for neurotransmitter release (Byrne et al., 1998).
Host Response

Mouse Response

  • Host Strain: Cr1:CD-1 (ICR) mice (Charles River).
  • Vaccination Protocol: Each of 10 mice per group was injected one to three times with 0.01, 0.1, 0.5, 1.0, or 2.0 µg FPLC-purified BoNT/A(Hc). Multiple injections were given 14 days apart. Two days before challenge, mice were bled retro-orbitally for ELISA and serum neutralization testing (Byrne et al., 1998).
  • Persistence: ELISA titers for individual mice successfully predicted survival. When the titers were at least 1600, 98.8% of the mice survived. When the titers were 100 or less, mice had only a 14.3% survival rate (Byrne et al., 1998).
  • Side Effects: None noted (Byrne et al., 1998).
  • Challenge Protocol: Mice were challenged 21 days after the last injection with 1,000 mouse i.p. LD50 of BoNT/A toxin complex diluted in GPB in a total volume of 100 µl per mouse. Mice were observed daily, and deaths were recorded 5 days post-challenge (Byrne et al., 1998).
  • Efficacy: In general, multiple injections protected better than one, with complete or nearly complete protection realized at doses of 0.5 µg/mouse (Byrne et al., 1998).
  • Description: Inhibition of BoNT action at a key step of the process could abolish the onset of botulism. One approach to developing a vaccine against botulism would be to construct and express a gene encoding only the binding domain of BoNT [BoNT(Hc)] and purify the translated product. This material, when administered to an organism, would not cause botulism because it lacks the enzyme and should not be able to enter the nerve cell without the translocation domain. Antibodies toward the product which neutralize BoNT serotype A (BoNT/A) toxicity when the host is directly challenged could be produced. Currently, a toxoid vaccine against BoNT serotypes A to E is used. However, there are inherent problems with the toxoid. The product consists of a crude extract of clostridial proteins. The material is dangerous to produce, and there is a high cost associated with preparing the toxoid vaccine. The toxoid also contains formalin, which is very painful for the recipient. Finally, only five of the seven serotypes are represented in the formulation. Thus, the aim of the present work was to develop a process for isolating a highly immunogenic recombinant BoNT(Hc) which could protect animals against a direct challenge of BoNT and that would be cheaper and less dangerous to produce. Ultimately, the developed process will be licensed as a vaccine (Byrne et al., 1998).
References
Byrne et al., 1998: Byrne MP, Smith TJ, Montgomery VA, Smith LA. Purification, potency, and efficacy of the botulinum neurotoxin type A binding domain from Pichia pastoris as a recombinant vaccine candidate. Infection and immunity. 1998 Oct; 66(10); 4817-22. [PubMed: 9746584].