Plant-made subunit vaccine against pneumonic and bubonic plague is orally immunogenic in mice

Alvarez, M. Lucrecia; Pinyerd, Heidi L; Crisantes, Jason D; Rigano, Maria Manuela; Pinkhasov, Julia; Walmsley, Amanda Maree; Mason, Hugh S.; Cardineau, Guy A.

doi:10.1016/j.vaccine.2005.12.057

Cited by 111 publications

(114 citation statements)

References 47 publications

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“…Numerous expression and purification strategies for F1-V have been published ranging from traditional prokaryotic systems [5,6,19,20] to transgenic tomatoes [21] and the tobacco-like Nicotiana benthamiana [22]. Regardless of the ultimate expression strategy employed, the final F1-V fusion protein will retain a tendency to multimerize because of its subunit composition.…”

Section: Discussionmentioning

confidence: 99%

Purification and protective efficacy of monomeric and modified Yersinia pestis capsular F1-V antigen fusion proteins for vaccination against plague

Goodin

Nellis

Powell

et al. 2007

Protein Expression and Purification

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The F1-V vaccine antigen, protective against Yersinia pestis, exhibits a strong tendency to multimerize that affects larger-scale manufacture and characterization. In this work, the sole F1-V cysteine was replaced with serine by site-directed mutagenesis for characterization of F1-V noncovalent multimer interactions and protective potency without participation by disulfide-linkages. F1-V and F1-V C424S proteins were over-expressed in Escherichia coli, recovered using mechanical lysis/pH-modulation and purified from urea-solubilized soft inclusion bodies, using successive ionexchange, ceramic hydroxyapatite, and size-exclusion chromatography. This purification method resulted in up to 2 mg per gram of cell paste of 95% pure, mono-disperse protein having ≤ 0.5 endotoxin units per mg by a kinetic chromogenic limulus amoebocyte lysate reactivity assay. Both F1-V and F1-V C424S were monomeric at pH 10.0 and progressively self-associated as pH conditions decreased to pH 6.0. Solution additives were screened for their ability to inhibit F1-V self-association at pH 6.5. An L-arginine buffer provided the greatest stabilizing effect. Conversion to >500-kDa multimers occurred between pH 6.0 and 5.0. Conditions for efficient F1-V adsorption to the cGMPcompatible Alhydrogel® adjuvant were optimized. Side-by-side evaluation for protective potency against subcutaneous plague infection in mice was conducted for F1-V C424S monomer; cysteinecapped F1-V monomer; cysteine-capped F1-V multimer; and a F1-V standard reported previously. After a two-dose vaccination with 2 × 20 µg of F1-V, respectively, 100, 80, 80, and 70% of injected mice survived a subcutaneous lethal plague challenge with 10 8 LD 50 Y. pestis CO92. Thus, vaccination with F1-V monomer and multimeric forms resulted in significant, and essentially equivalent, protection.

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Section: Discussionmentioning

confidence: 99%

Purification and protective efficacy of monomeric and modified Yersinia pestis capsular F1-V antigen fusion proteins for vaccination against plague

Goodin

Nellis

Powell

et al. 2007

Protein Expression and Purification

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“…In addition to the potential for natural infections, Y. pestis is generally considered to be among the top five potential biological weapons (19). Recent efforts to create a safe and effective pneumonic plague vaccine have focused on the development of recombinant subunit vaccines that elicit antibodies against two well-characterized Y. pestis antigens, the F1 capsule and the virulence protein LcrV (2,8,40,53). A plague vaccine based on live attenuated Y. pestis provides the theoretical advantage of simultaneously priming against many antigens, thereby greatly enhancing the likelihood of broadbased protection.…”

mentioning

confidence: 99%

Yersinia pestiswith Regulated Delayed Attenuation as a Vaccine Candidate To Induce Protective Immunity against Plague

et al. 2010

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Two mutant strains of Yersinia pestis KIM5؉, a ⌬crp mutant and a mutant with arabinose-dependent regulated delayed-shutoff crp expression (araC P BAD crp), were constructed, characterized in vitro, and evaluated for virulence, immunogenicity, and protective efficacy in mice. Both strains were highly attenuated by the subcutaneous (s.c.) route. The 50% lethal doses (LD 50 s) of the ⌬crp and araC P BAD crp mutants were approximately 1,000,000-fold and 10,000-fold higher than those of Y. pestis KIM5؉, respectively, indicating that both strains were highly attenuated. Mice vaccinated s.c. with 3.8 ؋ 10 7 CFU of the ⌬crp mutant developed high anti-Y. pestis and anti-LcrV serum IgG titers, both with a strong Th2 bias, and induced protective immunity against subcutaneous challenge with virulent Y. pestis (80% survival) but no protection against pulmonary challenge. Mice vaccinated with 3.0 ؋ 10 4 CFU of the araC P BAD crp mutant also developed high anti-Y. pestis and anti-LcrV serum IgG titers but with a more balanced Th1/Th2 response. This strain induced complete protection against s.c. challenge and partial protection (70% survival) against pulmonary challenge. Our results demonstrate that arabinose-dependent regulated crp expression is an effective strategy to attenuate Y. pestis while retaining strong immunogenicity, leading to protection against the pneumonic and bubonic forms of plague.Bubonic and pneumonic plague are zoonotic diseases endemic in many parts of the world, including the United States, and have resulted in over 200 million deaths over the course of human history (51). The etiological agent of plague is Yersinia pestis. Although the number of confirmed plague cases that occur worldwide has stabilized over the last 50 years at an average of about 1,700 per year, plague remains a serious public health threat in some regions of the world and outbreaks still occur (19). In addition to the potential for natural infections, Y. pestis is generally considered to be among the top five potential biological weapons (19). Recent efforts to create a safe and effective pneumonic plague vaccine have focused on the development of recombinant subunit vaccines that elicit antibodies against two well-characterized Y. pestis antigens, the F1 capsule and the virulence protein LcrV (2,8,40,53). A plague vaccine based on live attenuated Y. pestis provides the theoretical advantage of simultaneously priming against many antigens, thereby greatly enhancing the likelihood of broadbased protection. In the past, live attenuated strains were generated by selection, rather than precise genetic manipulation, thus raising concern about their genetic composition and stability. The live EV76 vaccine is an apparent pgm mutant that has been used in some countries (49). However, a concern is that the EV76 vaccine strain can cause disease in primates, raising questions about its suitability as a human vaccine (29). Nevertheless, as recently as 2002, USAMRIID researchers noted, "Despite their drawbacks, there is ample evidence that live-a...

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“…Freeze-dried plant material could be stored for long time and consumed without cooking; in addition, this technique could allow to standardize and concentrate the plant-made vaccine. Several studies applied this technique to vaccine produced in transgenic tomato and demonstrated that freeze-dried produced stable formulations for oral delivery (Alvarez et al, 2006;Salyaev et al, 2010;Shchelkunov et al, 2006;Soria-Guerra et al, 2011;Zhang et al, 2006).…”

Section: Production Of Pharmaceutical Proteinsmentioning

confidence: 99%

Genetic Transformation in Tomato: Novel Tools to Improve Fruit Quality and Pharmaceutical Production

Matteo¹,

Rigano²,

Sacco³

et al. 2011

Genetic Transformation

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Plant-made subunit vaccine against pneumonic and bubonic plague is orally immunogenic in mice

Cited by 111 publications

References 47 publications

Purification and protective efficacy of monomeric and modified Yersinia pestis capsular F1-V antigen fusion proteins for vaccination against plague

Purification and protective efficacy of monomeric and modified Yersinia pestis capsular F1-V antigen fusion proteins for vaccination against plague

Yersinia pestiswith Regulated Delayed Attenuation as a Vaccine Candidate To Induce Protective Immunity against Plague

Genetic Transformation in Tomato: Novel Tools to Improve Fruit Quality and Pharmaceutical Production

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