Influenza virus vaccines and uses thereof
The present disclosure provides influenza hemagglutinin stem domain polypeptides comprising (a) an influenza hemagglutinin HA1 domain that comprises an HA1 N-terminal stem segment, covalently linked by a linking sequence of 0-50 amino acid residues to an HA1 C-terminal stem segment, and (b) an influenza hemagglutinin HA2 domain, wherein on or more amino acids in the HA2 domain have been mutated. Also provided are nucleic acids encoding the polypeptides, compositions comprising the polypeptides and/or nucleic acid molecules, as well as methods of their use, in particular in the detection, prevention and/or treatment of influenza.
1. A polypeptide comprising:
(a) an influenza hemagglutinin HA1 domain that comprises an HA1 N-terminal stem segment comprising amino acids 1 to 52 of HA1, covalently linked by a linking sequence of 0 to 10 amino acid residues to an HA1 C-terminal stem segment comprising amino acids 321 to the end of HA1, wherein amino acids 53 to 320 are deleted, and
(b) an influenza hemagglutinin HA2 domain,
wherein the C-terminal amino acid residue of the HA1 C-terminal segment is any amino acid other than arginine (R) or lysine (K),
wherein one or more amino acids at position 406, 409, 413 and 416 are mutated into an amino acid selected from the group consisting of S, T, N, Q, R, H, K, D, E and G, and wherein the polypeptides comprise a disulphide bridge between the amino acids at position 324 and 436,
wherein the HA1 domain and HA2 domain are derived from an influenza A virus H1 subtype,
and wherein the numbering is based on the numbering of amino acids of the H1N1 influenza strain A/Brisbane/59/2007 (SEQ ID NO: 1).
2. The polypeptide of claim 1 , wherein the polypeptide is glycosylated.
3. The polypeptide of claim 1 , wherein the polypeptide comprises one or more further mutations in the HA1 domain and/or the HA2 domain.
4. The polypeptide of claim 1 , wherein the polypeptide does not comprise a signal sequence.
5. The polypeptide of claim 1 , wherein the polypeptide does not comprise the intracellular and transmembrane sequence of HA.
6. The polypeptide of claim 5 , wherein the polypeptide does not comprise the C-terminal part of the influenza H1 HA2 domain spanning from amino acid residue 520, 521, 522, 523, 524, 525, 526, 527, 528, 529 or 530 to the C-terminal amino acid of H1 HA2.
7. A polypeptide produced by a method comprising:
providing an influenza HA0 amino acid sequence;
removing the cleavage site between HA1 and HA2 by mutating the C-terminal amino acid of HA1 into an amino acid other than arginine (R) or lysine (K);
removing the amino acid sequence comprising the amino acid residues from position 53 to and including the amino acid residue on position 320 of the globular head domain from the HA0 sequence and joining the remaining parts of the sequence directly, or by introducing a linker sequences of 1 to 10 amino acids in length;
introducing one or more mutations in the amino acid sequence connecting the C terminal residue of helix A to the N terminal residue of helix CD; and
introducing at least a disulfide bridge in the HA stem domain polypeptide between amino acids 324 and 436 (numbering according to SEQ ID NO: 1).
8. An immunogenic composition comprising the polypeptide of claim 1 .
9. An immunogenic composition comprising the polypeptide of claim 1 in an amount sufficient to induce an immune response in a subject.
10. The polypeptide of claim 1 , wherein the C-terminal amino acid residue of the HA1 C-terminal stem segment is glutamine (Q).
11. The polypeptide of claim 1 , wherein the polypeptide comprises hemagglutinin stem domains from or based on HA of the influenza virus A/Brisbane/59/2007 (SEQ ID NO: 1).
12. The polypeptide of claim 1 , wherein the polypeptide comprises SEQ ID NO: 84 at position 419-433.
13. The polypeptide of claim 1 , wherein the polypeptide selectively binds to the antibodies CR6261 and/or CR9114, and does not bind to the antibody CR8057.