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-10953083-B2
Publication Date
2021-03-23
Expiration Date
2036-12-12
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
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 native positions of the NS1 and/or NS2 genes in the RSV genome are shifted to higher gene positions that are more distal to the promoter. The recombinant RSV strains described may include, in combination with gene position shifts, mutations at other loci to achieve desired levels of attenuation and immunogenicity. These recombinant RSV strains are suitable for use as live-attenuated RSV vaccines.
Human RSV causes significant mortality and morbidity worldwide, including severe infection in infancy often followed by prolonged airway dysfunction, and complicating asthma. Developing RSV vaccines has been challenging due to issues including disease enhancement following inactivated RSV vaccination, inefficiency of immune protection at the superficial respiratory epithelium, balancing replication and attenuation to maintain immunogenicity, moderate RSV titers and viral instability during manufacturing, and difficulty identifying stable attenuating mutations. There is a need for live attenuated RSV strains that replicate efficiently in vitro, are maximally immunogenic, satisfactorily attenuated, and are refractory to de-attenuation.
Shifting the NS1 and NS2 genes from their native promoter-proximal positions to higher gene positions results in decreased transcription and expression, thereby decreasing the antagonistic effect of NS1 and NS2 against host interferon responses. Unlike gene deletion mutants that cause over-attenuation and poor replication in vitro, the gene shifted RSV mutants maintain replication efficiency in cell lines such as Vero cells that lack interferon response, while exhibiting attenuation in interferon competent cells. This gene shift strategy enables incremental adjustment of attenuation without compromising manufacturing yield, providing a novel means to balance virus replication, attenuation, and immunogenicity for vaccine development.
Claims Coverage
The patent contains one independent claim that discloses a recombinant RSV with specific modifications in the NS1 and NS2 gene positions in the genome, and associated features and uses. The main inventive features focus on gene position shifts, attenuated virus phenotypes, and vaccine compositions.
NS1 and NS2 gene position shift
A recombinant RSV with the NS1 and NS2 genes shifted from native gene positions 1 and 2 to gene positions 9 and 10 in the RSV genome, resulting in attenuation.
Combination with gene deletions
The RSV genome may further comprise deletions of all or part of the NS1 gene, NS2 gene, or the M2-2 gene.
Defined genome sequence with gene shifts
The RSV genome comprises a positive-sense nucleotide sequence denoted by SEQ ID NO: 4 (6120/NS12Ltr) representing the gene shifts.
Reduced NS1/NS2 gene expression and transcription
The recombinant RSV exhibits reduced expression and transcription of NS1 and/or NS2 genes compared to RSV with native gene positions 1 and 2.
Reduced inhibition of host interferon response
The recombinant RSV shows reduced inhibition of host interferon response as compared to RSV with NS1 and NS2 in native gene positions.
Replication efficiency in interferon incompetent cells
The recombinant RSV retains replication efficiency in cultured cells lacking interferon production, e.g., Vero cells.
Infectious, attenuated, self-replicating RSV
The recombinant RSV is characterized as infectious, attenuated, and self-replicating, applicable to subtype A or B RSV.
Polynucleotide and vector compositions
Isolated polynucleotide molecules encoding the recombinant RSV genome or antigenome and vectors comprising such polynucleotides are claimed.
Methods for producing and using recombinant RSV
Methods include transfecting permissive cells with vectors to produce recombinant RSV, purifying the virus, and administering pharmaceutical compositions containing the recombinant RSV to subjects to elicit immune responses, including intranasal administration.
The claims cover recombinant RSV characterized by shifting NS1 and NS2 genes to more distal gene positions (specifically 9 and 10), optionally combined with deletions of NS1, NS2, or M2-2 genes, exhibiting reduced NS1 and NS2 expression and host interferon inhibition, maintaining replication in interferon incompetent cells, and useful as vaccine compositions with specified methods of production and administration.
Stated Advantages
The recombinant RSV strains retain efficient replication in Vero cells used for vaccine manufacture despite reduced NS1 and NS2 expression, overcoming replication restrictions seen in deletion mutants.
Gene shifting of NS1 and NS2 provides tunable incremental attenuation phenotypes while maintaining viral immunogenicity.
Reduced NS1 and NS2 expression leads to decreased inhibition of host interferon response, potentially increasing viral immunogenicity due to adjuvant effects of interferon.
Documented Applications
Use of the recombinant RSV strains as live-attenuated vaccines for immunizing human subjects, including infants (e.g., between 1 and 6 months of age).
Pharmaceutical compositions comprising the recombinant RSV and optionally an adjuvant for vaccine administration.
Methods of eliciting an immune response in subjects by administering an immunogenically effective amount of the recombinant RSV pharmaceutical compositions by intranasal, injection, aerosol, nasal spray, or nasal droplet routes.
Interested in licensing this patent?