Influenza hemagglutinin proteins and methods of use thereof
Inventors
Marshall, Christopher Patrick • Alff, Peter Joseph • Bertuccioli, Claudio • Yondola, Mark Andrew
Assignees
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Publication Number
US-9393297-B2
Publication Date
2016-07-19
Expiration Date
Abstract
In some embodiments the present invention provides influenza hemagglutinin (“HA”) polypeptides, proteins, and protein complexes that comprise a stalk domain that is engineered to facilitate maintenance of its native trimeric conformation, even if the head domain of the HA protein is removed or disrupted. In some embodiments, the present invention provides compositions comprising such polypeptides, proteins, and protein complexes, and methods of use of such proteins and compositions, for example as vaccine immunogens.
Core Innovation
The invention relates to influenza hemagglutinin (HA) polypeptides, proteins, or protein complexes comprising an amino acid sequence having at least 65% sequence identity to amino acid residues 229 to 519 of SEQ ID NO: 1 and including point mutations introducing one or more tyrosines at specified amino acid positions, including positions 403, 406, 411, 422, 429, 432, 433, and 435, and corresponding residues. The approach uses di-tyrosine (DT) cross-linking as a zero-length, irreversible, non-disruptive stabilization method for influenza HA polypeptides and protein complexes.
Engineered tyrosine point mutations introduce tyrosyl side chains at targeted HA stalk positions, and DT bond formation conformationally locks the native HA stalk trimer to preserve stalk and quaternary neutralizing epitopes. The DT stabilization concept is directed to maintaining the native trimeric stalk conformation and supporting the generation of headless HA immunogens.
The invention further describes headless influenza HA polypeptides, proteins, or protein complexes that preserve stalk quaternary neutralizing epitopes while including protease recognition motifs or engineered cleavage sites in the head for proteolytic head removal. The framework ties the tyrosine substitutions and DT cross-links to an influenza HA trimeric stalk complex context and includes optional removal of undesired from-tyrosine residues, foldon trimerization domains, and engineered cleavage sites.
Claims Coverage
Two independent claims are identified. The claim set covers sequence-identity-defined influenza HA variants with specified tyrosine point mutations at selected HA positions, including headless HA variants, with dependent claim refinements directed to di-tyrosine cross-linking in an influenza HA trimeric stalk complex context and selected paired tyrosine residues.
Sequence-identity-defined influenza HA with specified tyrosine point mutations
An influenza hemagglutinin (HA) polypeptide, protein or protein complex comprising an amino acid sequence having at least 65% sequence identity to amino acid residues 229 to 519 of SEQ ID NO: 1, wherein the amino acid sequence comprises a point mutation to tyrosine at one or more of amino acid positions 403, 406, 429, 432, 433, and 435, or an amino acid residue corresponding thereto.
Headless influenza HA with specified tyrosine point mutations
A headless influenza hemagglutinin (HA) polypeptide, protein or protein complex that comprises an amino acid sequence having at least 65% sequence identity to amino acid residues 229 to 519 of SEQ ID NO: 1, wherein the amino acid sequence comprises a point mutation to tyrosine at one or more of amino acid positions 403, 406, 411, 422, 429, 432, 433, and 435, or an amino acid residue corresponding thereto.
Overall, independent claim coverage is directed to influenza HA, including headless HA, sequences defined by at least 65% identity to residues 229 to 519 of SEQ ID NO: 1 and incorporating tyrosine point mutations at specified stalk positions. Dependent claim refinements further restrict these variants by requiring di-tyrosine cross-links within an influenza HA trimeric stalk complex and by specifying paired tyrosine residue positions.
Stated Advantages
Provides zero-length, irreversible, non-disruptive stabilization of influenza HA polypeptides and protein complexes.
Conformationally locks the native HA stalk trimer to preserve stalk and quaternary neutralizing epitopes.
Supports generation of headless HA immunogens while preserving stalk and quaternary neutralizing epitope features.
Stabilizes the conserved stalk domain in a native trimeric conformation, including when the HA head is removed (headless).
Binds to stalk-specific broadly neutralizing antibodies.
Elicits protective immune responses.
Documented Applications
Generation of DT-locked headless influenza HA immunogens that preserve stalk quaternary neutralizing epitope features for preclinical evaluation concepts.
Use of protease recognition motifs or engineered cleavage sites in headless influenza HA polypeptides and protein complexes to enable proteolytic head removal while maintaining DT-locked stalk conformation.
Vaccine immunogens and antibody targets using the engineered influenza HA (including headless HA) and related nucleic acids encoding HA.
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