Attenuation of human respiratory syncytial virus by genome scale codon-pair deoptimization
Inventors
Collins, Peter L. • LE NOUEN, Cyril • Brock, Linda G. • Buchholz, Ursula J. • DiNapoli, Joshua Marc • Mueller, Steffen • Wimmer, Eckard
Assignees
Research Foundation of the State University of New York • US Department of Health and Human Services
Publication Number
US-12054749-B2
Publication Date
2024-08-06
Expiration Date
2034-02-07
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Abstract
Described herein are RSV polynucleotide sequences that make use of multiple codons that are containing silent nucleotide substitutions engineered in multiple locations in the genome, wherein the substitutions introduce a numerous synonymous codons into the genome. Due to the large number of defects involved, the attenuated viruses disclosed herein provide a means of producing attenuated, live vaccines against RSV.
Core Innovation
Described herein are recombinant polynucleotide sequences of respiratory syncytial virus (RSV) that include multiple silent nucleotide substitutions engineered at various locations in the viral genome, introducing numerous synonymous codons without changing the amino acid sequences of the encoded proteins. This codon-pair deoptimization affects parameters such as codon bias, codon pair bias, RNA secondary structure, CpG dinucleotide content, and others, creating numerous defects in the virus genome. Because of these multiple defects, the resulting viruses are stably attenuated and serve as candidates for live RSV vaccines.
The background identifies human RSV as a significant pathogen causing high morbidity and mortality worldwide, with no commercially available vaccines despite ongoing development efforts. The problem addressed is the need for novel, effective, and stable attenuated RSV vaccine candidates. Traditional vaccine development struggled with achieving sufficient attenuation while maintaining immunogenicity and genetic stability.
The invention uses synthetic attenuated virus engineering (SAVE) to recode the RSV genome by rearranging synonymous codons to reduce codon pair bias relative to wild-type virus, thus attenuating virus replication but preserving wild-type protein sequences and antigenic epitopes. The attenuation can be adjusted by the extent of codon-pair deoptimization (CPD). The viruses constructed include codon-pair deoptimized versions of multiple RSV proteins such as NS1, NS2, N, P, M, SH, G, F, and L proteins, either individually or in combinations, creating viable, genetically stable, temperature-sensitive viruses with reduced replication in vitro and in vivo.
Claims Coverage
The claims include one independent claim directed to a recombinant polynucleotide encoding RSV proteins NS1 and/or NS2 with reduced codon pair bias and related claims covering recombinant viruses, compositions, methods of production, and uses.
Recombinant polynucleotide with reduced codon pair bias encoding RSV NS1 and/or NS2 proteins
A recombinant nucleotide sequence encoding the RSV NS1 protein, the RSV NS2 protein, or both, wherein the codon pair bias of the recombinant nucleotide sequence is reduced by at least 0.05 compared to the wildtype RSV sequence, while maintaining the corresponding amino acid sequence identical or with up to four substitutions, additions, or deletions.
Recombinant polynucleotide sequences with defined identity ranges to parental sequences
Recombinant nucleotide sequences encoding RSV NS1 and/or NS2 proteins with nucleotide sequence identity ranging from about 75% to about 95% to corresponding regions of a wildtype RSV, including specific references to nucleotide segments of SEQ ID NO: 1 and SEQ ID NO: 5.
Recombinant genome or antigenome incorporating the recombinant polynucleotide sequences
A recombinant RSV genome or antigenome comprising the codon pair deoptimized RSV NS1 and/or NS2 coding sequences described.
Production and use of recombinant RSV containing codon pair deoptimized sequences
Methods of producing recombinant viruses by expressing the recombinant RSV genome or antigenome in cells, recombinant viruses comprising the codon pair deoptimized genome, modified to possibly include previously characterized RSV mutations or deletions, compositions comprising such viruses with acceptable carriers or adjuvants, and methods of inducing immune responses in animals (including humans) by administering these recombinant viruses through various routes.
The claims cover recombinant RSV nucleotide sequences with reduced codon pair bias encoding NS1 and/or NS2 proteins, recombinant viruses and genomes comprising these sequences, methods of production and administration, and vaccine compositions, all aimed at stably attenuated RSV vaccines maintaining immunogenic protein sequences.
Stated Advantages
The attenuated viruses are viable yet more attenuated than previous mutants.
They exhibit enhanced genetic stability in vivo compared to prior attenuated RSV strains.
The vaccine candidates maintain the wild-type protein sequences, preserving antigenic epitopes for effective immune response.
Replication of the viruses can be titrated by adjusting the extent of codon pair deoptimization.
Documented Applications
Production of live, attenuated RSV vaccines for human and veterinary use.
Use of the attenuated recombinant RSV viruses for inducing protective immune responses against RSV infection by administration via injection, aerosol, nasal spray, oral or topical routes.
Use of recombinant RSV viruses in vaccine compositions including carriers and adjuvants.
Use in immunizing susceptible mammals, including humans, to prevent or reduce severity of RSV disease.
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