Nucleic acids containing a synthetic codon-optimized Sin Nombre virus full-length M gene
Inventors
Assignees
US Army Medical Research Institute of Infectious Diseases • United States Department of the Army
Publication Number
US-9315826-B2
Publication Date
2016-04-19
Expiration Date
2031-01-31
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Abstract
The invention contemplates a new synthetic, codon-optimized Sin Nombre virus (SNV) full-length M gene open reading frame (ORF) that encodes a unique consensus amino acid sequence. The SNV ORF was cloned into a plasmid to form the first stable recombinant SNV full-length M gene that elicits neutralizing antibodies. The gene can be engineered into a vaccine system, and is useful to protect mammals against infection with Sin Nombre virus.
Core Innovation
The invention describes a novel synthetic, codon-optimized Sin Nombre virus (SNV) full-length M gene open reading frame (ORF) that encodes a unique consensus amino acid sequence. This synthetic M gene has been engineered into a plasmid-based vaccine system, designated pWRG/SN-M(opt), which is capable of eliciting good neutralizing antibody responses against Sin Nombre virus. The vaccine construct can be delivered using gene gun or muscle electroporation and is the first to produce convincing levels of neutralizing antibodies against SNV in animals.
The problem addressed by the invention arises from the severe and often fatal hantavirus pulmonary syndrome (HPS) caused by SNV, which has a high case-fatality rate of 35-40% and no specific therapeutics or vaccines currently available. Prior efforts and vaccine attempts, including use of Andes virus M gene-based vaccines, failed to consistently elicit protective neutralizing antibodies against SNV. SNV infections progress rapidly, affecting healthy individuals, and there is an alarming lack of medical countermeasures for prevention or treatment. The invention overcomes prior limitations through a specifically optimized SNV M gene, providing an effective vaccine construct that elicits potent neutralizing antibodies to protect against SNV infection.
Claims Coverage
The patent includes 8 claims, with 4 independent claims defining the core inventive features related to nucleic acid sequences, DNA constructs, and cassettes.
Isolated nucleic acid sequences of the codon-optimized SNV M gene
An isolated nucleic acid sequence set forth in SEQ ID NO:1, including SEQ ID NO:2 and SEQ ID NO:3, representing the synthetic, codon-optimized full-length M gene open reading frame and flanking sequences of Sin Nombre virus.
Recombinant DNA constructs with promoter-linked SNV M gene sequences
A recombinant DNA construct comprising (i) a vector and (ii) the nucleic acid sequence set forth in SEQ ID NO:1, SEQ ID NO:2, or SEQ ID NO:3 operably linked to a promoter sequence, enabling expression of the SNV M gene product. The promoter may be the cytomegalovirus promoter with Intron A, beta-actin, or SV40 promoter, and the vector may be plasmid pWRG/SN-M(opt), recombinant adenovirus, recombinant vesicular stomatitis virus, or alphavirus replicon.
Use of specific promoters operably linked to SNV M gene in constructs
The recombinant DNA constructs utilize specific promoters operably linked to the SNV M gene nucleic acid fragment for effective expression, including the cytomegalovirus promoter operably linked to Intron A, beta-actin promoter, or SV40 promoter.
DNA cassettes comprising SNV M gene sequences linked to eukaryotic promoters
A DNA cassette comprising either SEQ ID NO:2 or SEQ ID NO:3 linked to a promoter operable in a eukaryotic expression system.
The claims cover isolated codon-optimized nucleic acid sequences of the SNV full-length M gene, recombinant constructs containing these sequences operably linked to mammalian or other promoters enabling expression in suitable vectors, and DNA cassettes comprising these sequences linked to eukaryotic promoters, constituting the invention's molecular basis for recombinant SNV vaccines and related uses.
Stated Advantages
The SNV M gene DNA vaccine plasmid pWRG/SN-M(opt) elicits high-titer neutralizing antibodies against SNV, representing the first vaccine to achieve this.
The codon optimization increases mRNA stability and plasmid stability, improving immunogenicity and potency compared to earlier versions.
The vaccine can be delivered efficiently via gene gun or muscle electroporation, enabling potent immune responses with fewer vaccinations.
Combination vaccines including SNV and other hantavirus M genes can elicit broad neutralizing antibodies, supporting multivalent vaccine approaches.
The vaccine approach avoids the need to culture pathogenic SNV, offering safety advantages and facilitating vaccine development.
Documented Applications
Use as a vaccine to protect mammals against infection with Sin Nombre virus by eliciting neutralizing antibodies.
Inclusion in multivalent hantavirus DNA vaccines to induce immunity against multiple HPS-associated hantaviruses or combined HPS and HFRS hantaviruses.
Production of polyclonal and monoclonal antibodies for therapeutic or prophylactic treatment of SNV infection or for passive immunization.
Diagnostic methods for detecting Sin Nombre virus infection using antibodies generated with the synthetic SNV M gene.
Production of pseudotyped viruses expressing SNV glycoproteins for serologic assays and targeted gene delivery to endothelial cells.
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