Puumala virus full-length M segment-based DNA vaccines
Inventors
Assignees
United States Department of the Army
Publication Number
US-8852598-B2
Publication Date
2014-10-07
Expiration Date
2028-02-12
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Abstract
The invention contemplates a new synthetic, codon-optimized Puumala virus (PUUV) full-length M gene open reading frame (ORF) that encodes a unique consensus amino acid sequence. The PUUV ORF was cloned into a plasmid to form the first stable PUUV full-length M gene that elicits neutralizing antibodies. The gene can be engineered into a molecular vaccine system, and is useful to protect mammals against infection with Puumala virus.
Core Innovation
The invention discloses a synthetic, codon-optimized Puumala virus (PUUV) full-length M gene open reading frame (ORF) encoding a unique consensus amino acid sequence. This PUUV ORF was cloned into a plasmid to form the first stable PUUV full-length M gene that elicits neutralizing antibodies. The gene can be engineered into molecular vaccine systems, including DNA vaccine plasmids or virus-vectored vaccines, and is useful to protect mammals against infection with Puumala virus.
The background identifies the problem that there is currently no Food and Drug Administration (FDA) approved vaccine or specific drug to prevent or treat hemorrhagic fever with renal syndrome (HFRS) or hantavirus pulmonary syndrome (HPS) caused by PUUV. Previous attempts to develop molecular vaccines eliciting neutralizing antibodies against PUUV have been unsuccessful due at least in part to instability of the full-length M gene sequence in plasmid systems and ineffective immunogenicity in animal models. Earlier PUUV M gene-based DNA vaccine constructs elicited insufficient or no neutralizing antibodies, were not immunogenic in certain animal models, and were unstable during plasmid amplification in E. coli.
The summary describes overcoming these issues by creating a synthetic, codon-optimized full-length M gene ORF with five amino acid alterations relative to the original unstable sequence. This codon-optimized gene eliminates sequences that destabilize the plasmid in E. coli, facilitating stable high-yield production. The resulting plasmid, designated pWRG/PUU-M(s2), effectively expresses full-length functional G1 and G2 glycoproteins and elicits strong neutralizing antibody responses in animals including hamsters and nonhuman primates, demonstrating protection against PUUV infection and cross-neutralization of geographically distinct PUUV strains. This is reported as the first molecular vaccine candidate to achieve such neutralizing antibody responses against PUUV.
Claims Coverage
The patent includes one independent claim directed to a passive vaccine composition, with subsequent dependent claims detailing additional features. The main inventive features relate to the composition of polyclonal antibodies derived from vaccinees immunized with a DNA vaccine plasmid expressing the synthetic Puumala virus M gene.
Passive vaccine composition comprising polyclonal antibodies from DNA vaccine recipients
A passive vaccine composition that includes polyclonal antibodies obtained from a population vaccinated with a Puumala virus DNA vaccine composed of a plasmid expressing the synthetic full-length M gene open reading frame (SEQ ID NO:1).
Use of a stable DNA vaccine plasmid expressing synthetic M gene sequence
The plasmid used for vaccination is characterized by comprising the stable, codon-optimized full-length PUUV M gene sequence with five amino acid corrections (pWRG/PUU-M(s2)).
Effectiveness against multiple Puumala virus strains
The passive vaccine is effective against distinct Puumala virus strains including Sotkamo, K27, and P360, demonstrating cross-strain protection potential.
Plasmid composition features
The plasmid comprises a vector and the DNA fragment comprising the nucleic acid sequence set forth in SEQ ID NO:1 operably linked to a promoter sequence functional in mammalian cells, wherein the vector can be an expression vector such as an adenovirus, alphavirus replicon, or vesicular stomatitis virus vector.
The claims cover passive vaccines comprising polyclonal antibodies derived from recipients vaccinated with a stable, synthetic, codon-optimized DNA plasmid encoding the Puumala virus full-length M gene, effective against multiple virus strains, and include details on the plasmid vector compositions.
Stated Advantages
Does not require the use of live Puumala virus, improving safety by avoiding a BSL-3 agent.
Eliminates the need to grow virus in rodent brains, simplifying manufacturing.
Avoids extensive testing for inactivation required in killed virus vaccines.
Does not require formulation with alum adjuvant.
Gene-based molecular vaccine approach enables production of glycoproteins within vaccinee cells, presenting them in a manner resembling natural infection.
Elicits neutralizing antibodies for the first time against PUUV, which were not produced by prior molecular vaccines.
Stable plasmid production with high yield DNA, overcoming previous instability issues in E. coli.
Cross-neutralizing antibodies against geographically distinct PUUV strains are induced.
Documented Applications
DNA vaccines to protect mammals, including humans, against infection with Puumala virus.
Use of pWRG/PUU-M(s2) or derivatives thereof in molecular vaccine systems, such as DNA plasmids, virus-vectored vaccines, and pseudotyped viruses.
Production of polyclonal or monoclonal antibodies for passive immunization or therapeutic treatments against Puumala virus infection.
Diagnostic methods for Puumala virus infection utilizing antibodies generated from the described DNA vaccine products.
Use in serologic assays or gene therapy delivery systems via production of pseudotyped viruses incorporating PUUV glycoproteins.
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