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Influenza Virus-Like Particles Produced by Transient Expression in Nicotiana Benthamiana Induce a Protective Immune Response against a Lethal Viral Challenge in Mice

Medicago Inc., 1020 Route de l'Eglise, Bureau 600, Québec, QC, Canada, G1V 3V9.
Plant Biotechnology Journal (Impact Factor: 5.68). 01/2009; 6(9):930-40. DOI: 10.1111/j.1467-7652.2008.00384.x
Source: PubMed

ABSTRACT A strain-specific vaccine represents the best possible response to the threat of an influenza pandemic. Rapid delivery of such a vaccine to the world's population before the peak of the first infection wave seems to be an unattainable goal with the current influenza vaccine manufacturing capacity. Plant-based transient expression is one of the few production systems that can meet the anticipated surge requirement. To assess the capability of plant agroinfiltration to produce an influenza vaccine, we expressed haemagglutinin (HA) from strains A/Indonesia/5/05 (H5N1) and A/New Caledonia/20/99 (H1N1) by agroinfiltration of Nicotiana benthamiana plants. Size distribution analysis of protein content in infiltrated leaves revealed that HA was predominantly assembled into high-molecular-weight structures. H5-containing structures were purified and examination by transmission electron microscopy confirmed virus-like particle (VLP) assembly. High-performance thin layer chromatography analysis of VLP lipid composition highlighted polar and neutral lipid contents comparable with those of purified plasma membranes from tobacco plants. Electron microscopy of VLP-producing cells in N. benthamiana leaves confirmed that VLPs accumulated in apoplastic indentations of the plasma membrane. Finally, immunization of mice with two doses of as little as 0.1 microg of purified influenza H5-VLPs triggered a strong immune response against the homologous virus, whereas two doses of 0.5 microg of H5-VLPs conferred complete protection against a lethal challenge with the heterologous A/Vietnam/1194/04 (H5N1) strain. These results show, for the first time, that plants are capable of producing enveloped influenza VLPs budding from the plasma membrane; such VLPs represent very promising candidates for vaccination against influenza pandemic strains.

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    • "Medicago, Inc., has also developed their own CPMV vector-based technology which produces virus-like particles (VLPs) carrying influenza virus antigens [24]. These VLPs express a lipid-anchored recombinant HA and are fully protected against lethal viral challenge in both mice and ferrets. "
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    03/2014; 2014:785382. DOI:10.1155/2014/785382
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    • "However, biosafety issues and difficulties with growth and yield of certain isolates may cause constraints during production. Besides whole virus vaccines, subunit vaccines comprising hemagglutinin (HA) and virus-like particles (VLPs) are considered as alternative strategies and have been shown to successfully induce neutralizing immune response [4] [5] [6] [7] [8] [9] [10] [11] [12] [13]. "
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    • "Protein assays are frequently described in the literature for the quantification of purified VLPs as illustrated in Table 1. D’Aoust et al. demonstrated 80% purity of plant derived VLP produced purified by size exclusion chromatography [29]. Additionally, protein assays such as the bicinchoninic acid (BCA) assay [17] or the Quant-IT assay kit with bovine serum albumin (BSA) [12] were used to quantify the total protein content in purified samples. "
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