Influenza immunogen and vaccine
AssigneesAcambis Inc., Vlaams Interuniversitair Institutuut voor Biotechnologie
InventorsAshley J. Birkett, Walter Fiers
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Abstract
A chimeric, carboxy-terminal truncated hepatitis B virus nucleocapsid (HBc) protein is disclosed that contains an immunogen for inducing the production of antibodies to the influenza M2 protein. An immunogenic influenza sequence in two to four copies is preferably expressed at or near the N-terminus or in the HBc immunogenic loop sequence. The HBc chimer preferably contains an influenza-specific T cell epitope and is preferably engineered for both enhanced stability of self-assembled particles and enhanced yield of those chimeric particles. Methods of making and using the chimers are also disclosed.
Claims45
A recombinant hepatitis B virus core (HBc) protein chimer molecule with a length of about 150 to about 375 amino acid residues that contains four peptide-linked amino acid residue sequence domains from the N-terminus that are denominated Domains I, II, III and IV, wherein
- (a) Domain I comprises (i) about 75 to about 160 amino acid residues whose sequence includes at least the sequence of the residues of position 4 through about position 75 of HBc, (ii) one to three Cysteine residues present at a position in the chimer molecule of about one to about −55 relative to the N-terminus of HBc of SEQ ID NO:1 [N-terminal Cysteine residue(s)], said one or more N-terminal Cysteine residues being present within a sequence other than that of the pre-core sequence of HBc, and (iii) includes two to four sequences of about 6 to about 24 residues of the influenza A M2 polypeptide of SEQ ID NO:9 that is peptide-bonded to or within about 15 residues of the N-terminus of the HBc sequence;
- (b) Domain II comprises about zero to about 60 amino acid residues peptide-bonded to about residue 75 of which (i) zero to all of the sequence of HBc is present from position 76 through 85 and (ii) an optional sequence of 6 to about 48 residues that constitute one or more repeats of 6 to about 24 residues of an influenza A M2 polypeptide of SEQ ID NO:9;
- (c) Domain III is an HBc sequence from about position 86 through about position 135 peptide-bonded to about residue 85;
- and
- d) Domain IV comprises (i) the residues of about positions 136 through 140 plus up to sixteen residues of an HBc amino acid residue sequence from position 141 through 156 peptide-bonded to the residue of about position 135 of Domain III, (ii) zero to three Cysteine residues, and (iii) up to about 100 amino acid residues in a sequence heterologous to HBc from position 156 to the HBc C-terminus;
- said chimer molecule (i) containing no more than 10 percent conservatively substituted amino acid residues in the HBc sequence, (ii) self-assembling into particles that are substantially free of binding to nucleic acids on expression in a host cell, and said particles being more stable on formation than are particles formed from an otherwise identical HBc chimer that lacks said N-terminal Cysteine residue(s) or in which an N-terminal Cysteine residue present in the chimer molecule is replaced by another residue.
A recombinant hepatitis B virus core (HBc) protein chimer molecule with a sequence of about 150 to about 235 amino acid residues that contains four peptide-linked domains from the N-terminus that are denominated Domains I, II, III and IV, wherein
- (a) Domain I comprises (i) about 95 to about 140 amino acid residues whose sequence includes at least the sequence of the residues of position 4 through about position 75 of HBc, (ii) one to three Cysteine residues present at a position in the chimer molecule of about one to about −55 relative to the N-terminus of HBc of SEQ ID NO:1 [N-terminal Cysteine residue(s)], said one or more N-terminal Cysteine residues being present within a sequence other than that of the pre-core sequence of HBc, and (iii) includes two to four sequences of about 6 to about 24 residues of the influenza A M2 polypeptide of SEQ ID NO:9 that is peptide-bonded to or within about 15 residues of the N-terminus of the HBc sequence;
- (b) Domain II comprises about zero to about 60 amino acid residues peptide-bonded to about residue 75 of which (i) zero to all of the sequence of HBc is present from position 76 through 85 and (ii) an optional sequence of 6 to about 48 residues that constitute one or more repeats of 6 to about 24 residues of an influenza A M2 polypeptide of SEQ ID NO:
- 9;
- (c) Domain III consists essentially of the HBc sequence from about position 86 through about position 135;
- and
- d) Domain IV comprises (i) the residues of about positions 136 through 140 plus up to sixteen residues of an HBc amino acid residue sequence from position 141 through 156 peptide-bonded to the residue of position 135 of Domain III, (ii) zero or one Cysteine residue, and (iii) up to about 50 amino acid residues in a sequence heterologous to HBc from position 156 to the HBc C-terminus;
- said chimer molecule (i) containing no more than 10 percent conservatively substituted amino acid residues in the HBc sequence, (ii) self-assembling into particles that are substantially free of binding to nucleic acids on expression in a host cell, and said particles being more stable on formation than are particles formed from an otherwise identical HBc chimer that lacks said N-terminal Cysteine residue(s) or in which an N-terminal Cysteine residue present in the chimer molecule is replaced by another residue.
Dependencies are inferred from the claim text.