Recombinant respiratory syncytial virus strains comprising NS1 and NS2 gene shifts
Inventors
Collins, Peter L. • McCarty, Thomas Charles
Assignees
US Department of Health and Human Services
Publication Number
US-11918638-B2
Publication Date
2024-03-05
Expiration Date
2036-12-12
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Abstract
Reported herein are novel recombinant respiratory syncytial viruses (RSV) having an attenuated phenotype in which the native positions of the NS1 and/or NS2 genes in the RSV genome are shifted to a higher position, that is at positions that are more distal to the promoter. The changes in the gene positions may be present in combination with mutations at other loci to achieve desired levels of attenuation and immunogenicity. The recombinant RSV strains described here are suitable for use as live-attenuated RSV vaccines. Also provided are polynucleotide sequences capable of encoding the described viruses, as well as methods for producing and using the viruses.
Core Innovation
Reported herein are novel recombinant respiratory syncytial viruses (RSV) having an attenuated phenotype in which the position of the NS1 and/or NS2 gene in the RSV genome or antigenome is shifted to a position that is more distal to the promoter. The changes in the gene positions may be present in combination with mutations at other loci to achieve desired levels of attenuation and immunogenicity. The recombinant RSV strains described here are suitable for use as live-attenuated RSV vaccines.
Human respiratory syncytial virus (RSV) infects nearly everyone worldwide early in life and is responsible for considerable mortality and morbidity, including 75,000-125,000 hospitalizations yearly in the United States and millions of pediatric infections and deaths worldwide. Severe infection in infancy frequently leads to lingering airway dysfunction and exacerbates or may initiate asthma. The development of RSV vaccines has been complicated by the risk of enhanced disease with inactivated and subunit vaccines, the inefficiency of immune protection due to replication in superficial respiratory cells, and the challenge of balancing attenuation with sufficient replication for immunogenicity.
The recombinant RSV disclosed comprise the native RSV genome modified by shifting the NS1 and/or NS2 genes from their native promoter-proximal positions 1 and 2 to higher gene positions more distal to the promoter, such as positions 7 and 8 or 9 and 10, independently or in tandem. This gene shift results in decreased transcription and expression of NS1 and NS2, reducing their inhibition of host interferon responses, thereby attenuating the virus while retaining replication efficiency in interferon-incompetent cells such as Vero cells. Deletions of NS1 and NS2 severely restrict replication, but the gene-shift mutants achieve attenuation without over-restriction, providing a novel approach to control attenuation levels.
Claims Coverage
The patent includes one independent claim detailing recombinant RSV with specific gene shifts and related features. The main inventive features encompass genetic modifications leading to attenuation through NS1 and NS2 gene repositioning and associated genome characteristics.
Shift of NS1 and NS2 genes from native to distal positions in RSV genome
A recombinant RSV modified by shifting the NS1 and NS2 genes from gene positions 1 and 2 to gene positions 7 and 8, respectively, resulting in attenuation.
Additional genomic modifications with deletion of M2-2 gene
The RSV genome may further comprise deletion of all or part of the M2-2 gene to influence viral properties.
Inclusion of reporter gene encoding GFP
The genome may comprise a reporter gene such as Green Fluorescent Protein for visualization purposes in research.
Reduced expression and transcription of NS1 and NS2 genes
The recombinant RSV exhibits reduced expression and transcription of NS1 and/or NS2 genes compared to wild-type positioning.
Reduced inhibition of host interferon response
The modified RSV shows reduced inhibition of host interferon response compared to wild-type RSV, increasing susceptibility to interferon-mediated restriction.
Efficiency of replication in interferon-incompetent cells retained
The recombinant RSV maintains replication efficiency in cultured cells unable to produce interferons, such as Vero cells, supporting vaccine manufacture.
Applicability to RSV subtypes with infectious, attenuated, self-replicating phenotype
The recombinant RSV can be subtype A or B, maintaining infectivity, attenuated phenotype, and self-replication suitable for vaccines.
The claim coverage centers on recombinant RSV strains attenuated by shifting the NS1 and NS2 genes distally in the genome, optionally combined with other deletions and reporter genes, leading to reduced NS gene expression, reduced interferon antagonism, sustained replication in certain cell types, and applicability as live-attenuated RSV vaccine candidates.
Stated Advantages
The recombinant RSV strains replicate efficiently in Vero cells, the preferred substrate for vaccine manufacture, unlike NS-deletion mutants that exhibit growth restriction.
The gene shift provides a controllable magnitude of attenuation by incremental repositioning of NS1 and NS2 genes, avoiding over-attenuation associated with gene deletion.
Reduced inhibition of the host interferon response may enhance immunogenicity through increased interferon expression and apoptosis, potentially improving vaccine efficacy.
The mutations render recombinant RSV increasingly susceptible to restriction in interferon-producing cells while retaining replication capacity in interferon-deficient cells, balancing safety and immunogenicity.
Documented Applications
Use of the recombinant RSV strains as live-attenuated RSV vaccines for immunizing human subjects, including infants between 1 and 18 months of age.
Vaccine formulations comprising the recombinant RSV, optionally with adjuvants, administered intranasally, by injection, aerosol, nasal spray, or nasal droplets.
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