Conformationally stabilized RSV pre-fusion F proteins
Inventors
Marshall, Christopher Patrick • McLellan, Jason Scott • Alff, Peter Joseph • Bertuccioli, Claudio • Mariani, Roberto
Assignees
Calder Biosciences Inc • US Department of Health and Human Services
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Publication Number
US-11993633-B2
Publication Date
2024-05-28
Expiration Date
2034-07-24
Abstract
In some embodiments, the present invention provides respiratory syncytial virus (RSV) F proteins, polypeptides and protein complexes that comprise one or more cross-links to stabilize the protein, polypeptide or protein complex in its pre-fusion conformation. In some embodiments the present invention provides RSV F proteins, polypeptides and protein complexes comprising one or more mutations to facilitate such cross-linking. In some embodiments the present invention provides compositions comprising such proteins, polypeptides or protein complexes, including vaccine compositions, and methods of making and using the same.
Core Innovation
The invention provides respiratory syncytial virus (RSV) F proteins, polypeptides, and protein complexes that are cross-linked to stabilize the protein in its pre-fusion conformation. The invention includes RSV F proteins with mutations to facilitate cross-linking, compositions containing such proteins including vaccines, and methods of making and using these proteins.
The problem addressed is the instability of the RSV F protein in its pre-fusion conformation, which is highly unstable and tends to transition to the post-fusion conformation, limiting its use as a vaccine immunogen. Current prophylactic treatments are limited and costly, creating a need for more effective and cost-efficient vaccines.
The invention solves this by providing specific amino acid sites in the RSV F protein where cross-links, especially di-tyrosine cross-links, can be introduced to stabilize the pre-fusion conformation. The invention also provides engineered RSV F proteins with point mutations to tyrosine at these sites to facilitate such cross-linking, alone or in combination with other stabilizing mutations or domains like foldon trimerization domains.
Claims Coverage
The claims include one independent claim focused on an RSV F polypeptide, protein, or protein complex with specific mutations and domains. The main inventive features relate to mutation, cross-linking, trimerization domains, and nucleic acid sequences encoding such proteins.
Point mutation to tyrosine at position 226
An RSV F polypeptide, protein or protein complex comprising a point mutation to tyrosine at amino acid position 226 or a corresponding position determined by alignment to SEQ ID NO.1.
Inclusion of cavity-filling hydrophobic substitutions
The RSV F polypeptide, protein or protein complex includes one or more cavity-filling hydrophobic amino acid substitutions.
Presence of a trimerization domain
The RSV F polypeptide, protein or protein complex includes a trimerization domain, specifically a foldon domain in some claims.
Di-tyrosine cross-link between tyrosines at positions 226 and 198
The RSV F polypeptide, protein or protein complex further comprises a di-tyrosine cross-link between the tyrosine at position 226 and the tyrosine at position 198 or their corresponding aligned positions.
Nucleic acid encoding RSV F proteins with the above features
Nucleic acid molecules encoding RSV F polypeptides, proteins or protein complexes that include the point mutation at position 226, optionally the foldon domain, and optionally the di-tyrosine cross-link between positions 226 and 198.
Compositions comprising the engineered RSV F proteins
Pharmaceutical or vaccine compositions comprising the engineered RSV F polypeptides, proteins or protein complexes and optionally comprising an adjuvant.
The independent claim covers engineered RSV F proteins featuring a tyrosine substitution at position 226, hydrophobic cavity-filling substitutions, a trimerization domain, and optionally di-tyrosine cross-links between residues 226 and 198, as well as nucleic acids encoding these proteins and compositions comprising them.
Stated Advantages
Stabilization of the RSV F protein in its pre-fusion conformation, increasing its stability and usefulness as a vaccine immunogen.
Improved ability to maintain key neutralizing epitopes such as antigenic site ∅.
Introduced cross-links such as di-tyrosine bonds provide stable, zero-length, physiologically stable covalent bonds that prevent transition to post-fusion form.
Cross-linking enhances the ability of RSV F proteins to bind neutralizing antibodies and elicit protective immune responses in vivo.
Targeted cross-linking avoids protein aggregation and improves manufacturability and storage stability of vaccine immunogens.
Documented Applications
Use of the stabilized RSV F polypeptides, proteins, and protein complexes as vaccine immunogens to elicit protective immune responses against RSV infection.
Methods of vaccinating subjects, including humans less than 24 months old or greater than 50 years old, with compositions comprising the engineered RSV F proteins.
Use as research and diagnostic tools for studying and measuring anti-RSV F antibodies.
Production of therapeutic, research, chimeric, humanized or fully-human monoclonal or polyclonal antibodies by immunization of animals using the RSV F polypeptides, proteins, or complexes.
Pharmaceutical compositions comprising the engineered RSV F proteins for treatment or prevention of RSV infection.
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