Vaccine candidates for human respiratory syncytial virus (RSV) having attenuated phenotypes
Inventors
LENOUEN, Cyril • Buchholz, Ursula J. • Collins, Peter L. • Mueller, Steffen
Assignees
Usa Secretary Dhhs • Codagenix Inc • US Department of Health and Human Services
Publication Number
US-11390651-B2
Publication Date
2022-07-19
Expiration Date
2037-09-22
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Abstract
Reported herein are presumptively de-attenuating mutations that are useful, either individually or in combinations that may include other known mutations, in producing recombinant strains of human respiratory syncytial virus (RSV) exhibiting attenuation phenotypes. Also described herein is a novel RSV construct, Min_L-NPM2-1(N88K)L, which exhibits an attenuated phenotype, is stable and is as immunogenic as wild type RSV. The recombinant RSV strains described here are suitable for use as live-attenuated RSV vaccines. Exemplary vaccine candidates are described. Also provided are polynucleotide sequences capable of encoding the described viruses, as well as methods for producing and using the viruses.
Core Innovation
The invention provides recombinant strains of human respiratory syncytial virus (RSV) exhibiting attenuated phenotypes suitable for use as live-attenuated RSV vaccines. These strains include mutations that are presumptively de-attenuating in vitro but confer attenuation in vivo. A novel RSV construct, Min_L-NPM2-1[N88K]L, is described, which exhibits an attenuated phenotype, is genetically stable, and induces an immune response comparable to wild-type RSV.
The problem addressed is the challenge in RSV vaccine development, including the lack of commercially available vaccines or antiviral drugs, risks of disease enhancement with inactivated or subunit vaccines, inefficient immune protection due to replication in respiratory airway luminal cells, the balance required between attenuation and immunogenicity, limited replication of RSV in animal models, instability of attenuating mutations leading to reversion to virulence, and difficulties in identifying attenuating mutations that allow for efficient replication in vitro and durable attenuation in vivo.
Claims Coverage
The claims include a single independent claim covering an isolated polynucleotide encoding a recombinant RSV variant with an attenuated phenotype defined by specific mutations. The inventive features focus on particular mutations in several viral open reading frames (ORFs), codon-pair deoptimization, and vaccine compositions and methods.
Specific mutation combination conferring attenuation
An isolated polynucleotide encoding a recombinant RSV variant, wherein the RSV genome or antigenome contains a mutation in the L ORF at a position corresponding to T1166 of the L protein, further modified by at least two mutations selected from mutations at positions corresponding to N88 or A73 of M2-1, K136 of N, and E114 of P proteins.
Particular sets of additional mutations in M2-1, N, and P
The RSV genome or antigenome further modified by mutation in M2-1 at position N88 (N88K), in N at K136 (K136R), and in P at E114 (E114V); or alternatively, mutation in M2-1 at A73 (A73S), in N at K136, and in P at E114.
Specific amino acid substitutions in key viral proteins
The RSV genome or antigenome comprises mutations corresponding to T1166I in L; and either N88K or A73S in M2-1; K136R in N; and E114V in P proteins.
Inclusion of deletions in accessory proteins for attenuation
The RSV genome or antigenome optionally includes deletions in at least one of the proteins M2-2, NS1, and NS2.
Codon pair deoptimization as an attenuation strategy
The RSV genome or antigenome or specifically the L ORF is codon-pair deoptimized to confer attenuation.
Vectors and cells comprising the polynucleotides
Vectors and cells comprising the isolated polynucleotide encoding the recombinant RSV variants are claimed.
Pharmaceutical compositions and methods of vaccination
Pharmaceutical compositions comprising immunologically effective amounts of the recombinant RSV variants, methods of vaccinating subjects, inducing immune responses, and different routes of administration are described.
Live attenuated RSV vaccines and methods of production
Live attenuated RSV vaccines comprising the recombinant RSV variants and methods of making such vaccines by expressing the isolated polynucleotides are included.
Nucleotide sequence identity claims
Isolated polynucleotide molecules with at least about 95% identity to SEQ ID NO:14, including the specific sequence of SEQ ID NO:14, vectors, cells, and pharmaceutical compositions comprising same, are claimed along with their uses in vaccination and immune response induction.
The claims collectively cover recombinant RSV variants attenuated by defined mutations in the L, M2-1, N, and P ORFs; codon pair deoptimization; deletions in accessory genes; their nucleic acid sequences; vectors and cells harboring them; pharmaceutical compositions; live vaccines; and methods of vaccination and vaccine production employing these recombinant viruses.
Stated Advantages
The codon-pair deoptimized RSV strains with multiple CPD ORFs (e.g., Min_FLC) are genetically and phenotypically stable under temperature stress and maintain attenuation.
The novel RSV construct Min_L-NPM2-1[N88K]L is more attenuated in vivo than Min_L yet induces an immune response as strong as wild-type RSV, representing a substantial improvement over prior candidates.
Min_L-NPM2-1[N88K]L shows increased replication in Vero cells facilitating vaccine manufacture, while maintaining genetic stability under conditions promoting de-attenuation in previous strains.
Specific mutations in the M2-1 protein improve polymerase L gene transcription efficiency, enhancing replication without compromising attenuation.
Documented Applications
Use as live-attenuated RSV vaccines for immunizing human subjects, including neonates, infants, and adults vulnerable to RSV infection.
Administration of the vaccine compositions via injection, aerosol delivery, nasal spray, or nasal droplets, primarily intranasally.
Inducing an immune response against RSV in subjects susceptible to RSV infection.
Incorporation of these recombinant RSV strains in pharmaceutical compositions and methods for making live attenuated vaccines for prophylactic immunization.
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