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Vaccine Detail

DNA vaccine encoding PA83 and LF
Vaccine Information
  • Vaccine Name: DNA vaccine encoding PA83 and LF
  • Target Pathogen: Bacillus anthracis
  • Target Disease: Anthrax
  • Vaccine Ontology ID: VO_0000521
  • Type: DNA vaccine
  • Antigen: B. anthracis PA and LF (Hermanson et al., 2004)
  • PagA from B. anthracis str. 'Ames Ancestor' gene engineering:
    • Type: DNA vaccine construction
    • Description: The PA construct is chemically synthesized to include an amino terminal human tissue plasminogen activator (hTPA) leader peptide fused to a PA83 sequence (amino acids 30–764) with the furin cleavage site deleted (SRKKRS, amino acids 192–197). This construct, designated PA83 furin, is cloned into the mammalian expression vector VR1012 (Hermanson et al., 2004).
    • Detailed Gene Information: Click Here.
  • Lef from B. anthracis str. A2012 gene engineering:
    • Type: Protein
    • Description: contains Bacillus anthracis lethal factor x-ray crystal structure derived domain I; LF; vaccine construct
    • Detailed Gene Information: Click Here.
  • DNA vaccine plasmid: pVR1012 DNA vaccine plasmid
  • Preparation: The PA construct is chemically synthesized to include an amino terminal human tissue plasminogen activator (hTPA) leader peptide fused to a PA83 sequence (amino acids 30–764) with the furin cleavage site deleted (SRKKRS, amino acids 192–197). This construct, designated PA83 furin, is cloned into the mammalian expression vector VR1012. The LF coding sequences are derived from the B. anthracis LF93 protein sequence, codon-optimized, and chemically synthesized as above to include the hTPA leader peptide. The LF domain I–III is PCR amplified from this clone by using a forward and reverse primer pair to amplify the 1,740-bp fragment encoding the hTPA leader peptide fused to LF amino acids 34–583. The LF domain I is also derived from the LF93 plasmid by PCR amplification using forward and reverse primer pairs to amplify an 876-bp fragment encoding an hTPA leader peptide fused to LF amino acids 34–295. Both LF genes are cloned into the VR1012 vector (Hermanson et al., 2004).
  • Virulence: The virulence of B. anthracis in rabbits, non-human primates, and humans is primarily the result of a multicomponent toxin secreted by the organism. The toxin consists of three separate gene products, designated protective antigen (PA), lethal factor (LF), and edema factor (EF), that are encoded on a 184-kb plasmid designated pXO1 (Hermanson et al., 2004).
  • Description: DNA vaccines provide an attractive technology platform against bioterrorism agents due to their safety record in humans and ease of construction, testing, and manufacture. Monovalent and bivalent anthrax plasmid DNA (pDNA) vaccines encoding genetically detoxified protective antigen (PA) and lethal factor (LF) proteins have been designed and tested for their immunogenicity and ability to protect rabbits from an aerosolized inhalation spore challenge. Immune responses after two or three injections of cationic lipid-formulated PA, PA + LF, or LF pDNAs were at least equivalent to two doses of anthrax vaccine adsorbed (AVA) (Hermanson et al., 2004).
Host Response

Rabbit Response

  • Host Strain: New Zealand White
  • Vaccination Protocol: Plasmid DNA was prepared from overnight cultures of transformed XL-2 Blue bacteria in Terrific Broth plus 50 µg/ml kanamycin sulfate and processed by using Endo-free Giga kits. One milliliter of sterile water for irrigation (SWFI) was added to a vial containing a dried film of 3.75 µmol each of a 1:1 mixture of cationic lipid and colipid and vortex mixed for 5 minutes. The liposome suspension was diluted to 1.5 mM with SWFI and added to an equal volume of pDNA and vortex mixed briefly. The final molar ratio of all formulations was 4:1, DNA/cationic lipid (Hermanson et al., 2004). Two- to five-kilogram female New Zealand White rabbits were injected bilaterally in the quadriceps muscles with 1 ml of pDNA formulated with Vaxfectin or DMRIE/DOPE (0.5 ml per leg). Rabbits vaccinated with PA, LF, or vector received 1 mg of that pDNA whereas rabbits co-injected with PA + LF pDNAs received a mixture of 0.5 mg of each plasmid. Groups of rabbits receiving three doses were injected on days 0, 28, and 56; rabbits receiving only two doses were injected on study days 0 and 28. Rabbits immunized with AVA were injected unilaterally with 50 µl of AVA diluted to 0.5 ml in PBS on days 28 and 56. Prebleeds and biweekly postvaccination bleeds were taken for all groups for analysis of serum antibodies (Hermanson et al., 2004).
  • Persistence: Spore challenge induced a significant increase in the Letx neutralization titer in group 4 rabbits, suggesting that there was limited spore germination after challenge in these animals. This post-challenge increase in Letx neutralization titer, however, was smaller than the increase seen in AVA- or LF pDNA-vaccinated rabbits challenged at week 12 (Hermanson et al., 2004).
  • Immune Response: Both the PA and the LF pDNAs generate anti-PA and anti-LF antibody responses, respectively, when injected alone or co-injected. Furthermore, co-injection of PA and LF pDNAs does not cause detectable interference in the immunogenicity of either of the pDNAs (Hermanson et al., 2004). Immune responses after two or three injections of cationic lipid-formulated PA, PA + LF, or LF pDNAs were at least equivalent to two doses of anthrax vaccine adsorbed (AVA). High titers of anti-PA, anti-LF, and neutralizing antibody to lethal toxin were achieved in all rabbits.
  • Side Effects: none reported (Hermanson et al., 2004)
  • Challenge Protocol: Eight or nine animals in each group were challenged with 100x LD50 of aerosolized anthrax spores 5 or 9 weeks after vaccination. An additional 10 animals vaccinated with PA pDNA were challenged over 7 months post-vaccination.
  • Efficacy: All animals receiving PA or PA + LF pDNA vaccines were protected. In addition, 5/9 animals receiving LF pDNA survived, and the time to death was significantly delayed in the others. Groups receiving three immunizations with PA or PA + LF pDNA showed no increase in anti-PA, anti-LF, or Letx neutralizing antibody titers post-challenge, suggesting little or no spore germination (Hermanson et al., 2004).
References
Hermanson et al., 2004: Hermanson G, Whitlow V, Parker S, Tonsky K, Rusalov D, Ferrari M, Lalor P, Komai M, Mere R, Bell M, Brenneman K, Mateczun A, Evans T, Kaslow D, Galloway D, Hobart P. A cationic lipid-formulated plasmid DNA vaccine confers sustained antibody-mediated protection against aerosolized anthrax spores. Proceedings of the National Academy of Sciences of the United States of America. 2004 Sep 14; 101(37); 13601-6. [PubMed: 15342913].