Attenuation of human respiratory syncytial virus by genome scale codon-pair deoptimization
Inventors
Collins, Peter L. • Le Nouën, 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-9957486-B2
Publication Date
2018-05-01
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
Human respiratory syncytial virus (RSV) is a significant pathogen responsible for extensive pediatric infections and deaths worldwide, causing considerable morbidity and mortality. Despite ongoing vaccine and antiviral drug development, no commercially available vaccines exist. There is a need for new vaccine candidates that are effective and safe.
The invention describes recombinant RSV polynucleotide sequences that use multiple codons containing silent nucleotide substitutions engineered in multiple genomic locations. These substitutions introduce numerous synonymous codons into the viral genome without altering the amino acid sequence of viral proteins. This codon pair deoptimization alters parameters such as codon bias, codon pair bias, RNA secondary structure, CpG dinucleotide content, and others.
Because of the large number of defects introduced by this widespread synonymous codon substitution, the resulting RSV viruses are stably attenuated and suitable for producing live vaccines. The viruses retain the same antigenic features as wild-type RSV, thereby likely inducing appropriate cellular and humoral immunity to the same epitopes as wild-type virus. This approach aims to provide stably attenuated, live vaccines against RSV with improved safety and genetic stability.
Claims Coverage
The patent contains one independent claim focused on a recombinant polynucleotide encoding RSV with specific codon pair deoptimization in the RSV L protein gene, and related features affecting replication and immune response.
Recombinant polynucleotide with codon pair deoptimized RSV L protein sequence
A recombinant polynucleotide encoding RSV wherein the nucleotide sequence encoding the RSV L protein comprises a codon pair deoptimized sequence that is about 70% to about 90% identical to the corresponding wild-type sequence (SEQ ID NO:1), resulting in an attenuated virus.
Incorporation of specific amino acid changing mutations in viral proteins
The recombinant polynucleotide further includes nucleotide changes causing amino acid substitutions at specific positions in RSV proteins N, P, and M2-1, namely position 136 in N, 114 in P, 88 and/or 73 in M2-1, which may affect viral phenotype.
Methods for producing recombinant RSV and eliciting immune response
Methods of producing recombinant RSV by expressing the codon pair deoptimized polynucleotide in a cell; and methods of inducing an immune response in mammals, including humans, by administering such recombinant RSV.
The independent claim covers a recombinant RSV polynucleotide with codon pair deoptimization in the L gene and includes embodiments with additional site-specific mutations, as well as methods for producing this virus and using it to generate an immune response.
Stated Advantages
The attenuated viruses produced are stably attenuated and genetically stable in vivo, capable of replication sufficient to induce immune responses but with reduced pathogenicity compared to wild-type RSV.
Viruses maintain identical amino acid sequences of viral proteins, preserving antigenicity and likely inducing both cellular and humoral immune responses targeting wild-type epitopes.
Codon pair deoptimization allows titratable attenuation by adjusting the extent of deoptimization, enabling control over virus attenuation levels.
These attenuated viruses can be propagated efficiently in cell culture at permissive temperatures, allowing sufficient vaccine manufacture yields.
The approach can be combined with known attenuating mutations to enhance vaccine safety and immunogenicity.
Documented Applications
Production of live, attenuated RSV vaccines for human use, particularly for immunization against RSV infection and disease.
Use in immunogenic compositions to induce protective immune responses in mammals, including humans.
Methods for producing recombinant RSV vaccines by expression of codon pair deoptimized polynucleotides in cell culture.
Use of attenuated RSV strains in susceptible hosts such as infants, children, and other at-risk populations for prevention of RSV disease.
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