Stabilised viral fusion proteins
AssigneeOxford University Innovation Limited
InventorsAlexander DOUGLAS, Sofiya FEDOSYUK
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Abstract
The invention relates to stabilised pre-fusion conformation Class III fusion proteins. The inve ntion also provides vaccine compositions for immunising a subject against viral infections.
Claims33
A stable pre-fusion Class III fusion protein, or an immunogenic fragment thereof, comprising one or more mutations that prevent formation of the central extended helix in the post-fusion conformation trimerisation interface.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein said central extended helix in the post-fusion conformation trimerisation interface is defined as extending, at its N-terminal end, up to the 32nd amino acid residue N-terminal to the conserved cysteine residue of the Class III fusion protein and, at its C-terminal end, up to the 16th amino acid residue C-terminal to the conserved cysteine of the Class III fusion protein;
- wherein optionally the conserved cysteine residue corresponds to or aligns with:
- (a) amino acid residue 283 of the RVG sequence according to SEQ ID NO:
- 3;
- (b) amino acid residue 284 of the VSVG sequence according to SEQ ID NO:
- 5;
- (c) amino acid residue 484 of the EBV gB sequence according to SEQ ID NO:
- 6;
- (d) amino acid residue 507 of the cytomegalovirus glycoprotein B (CMV gB) sequence according to SEQ ID NO:
- 8;
- and/or
- (e) amino acid residue 529 of the herpes simplex virus glycoprotein B (HSV-1 gB) sequence according to SEQ ID NO:
- 10.
- (f) amino acid residue 526 of the herpes simplex virus glycoprotein B (HSV-2 gB) sequence according to SEQ ID NO:
- 32.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein said central extended helix of the post-fusion conformation trimerisation interface corresponds to, or aligns with amino acid residues:
- (a) 262 to 293 of the RVG sequence of SEQ ID NO:
- 3;
- (b) 263 to 294 of the VSVG sequence of SEQ ID NO:
- 5;
- (c) 458 to 500 of the EBV gB sequence of SEQ ID NO:
- 6;
- (d) 481 to 520 of the CMV gB sequence of SEQ ID NO:
- 8;
- and/or
- (e) 503 to 545 of the HSV-1 gB sequence of SEQ ID NO:
- 10.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein the Class III fusion protein is a:
- (a) RVG;
- (b) VSVG;
- or
- (c) herpesvirus glycoprotein B, wherein optionally said herpesvirus is selected from a cytomegalovirus, Epstein- Barr virus, herpes simplex virus-1 and/or herpes simplex virus-2.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein said one or more mutation is an amino acid substitution.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein said amino acid substitution is a non-conservative amino acid substitution.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein said one or more mutation is:
- (a) a helix-disrupting mutation and said amino acid substitution is a substitution by proline;
- and/or
- (b) an amino acid substitution by an amino acid with increased hydrophobicity compared with the amino acid being substituted, wherein said amino acid substitution is independently selected from a substitution by leucine, alanine, isoleucine or valine.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein said one or more mutation is at an amino acid corresponding to, or aligning with, position 270, 271, 272 and/or 273 of the RVG sequence of SEQ ID NO:
- 3.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein the amino acid corresponding to position:
- (a) 270 of the RVG sequence of SEQ ID NO:
- 3 is substituted by proline;
- (b) 271 of the RVG sequence of SEQ ID NO:
- 3 is substituted by proline;
- (c) 272 of the RVG sequence of SEQ ID NO:
- 3 is substituted by proline;
- and/or
- (d) 273 of the RVG sequence of SEQ ID NO:
- 3 is substituted by proline.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein the Class III fusion protein is a RVG and wherein said protein comprises one or more amino acid substitutions selected from H270P, L271P, V272P and/or V273P.
The protein or immunogenic fragment thereof of any one of the preceding claims, wherein the protein or immunogenic fragment thereof is RVG comprising an amino acid sequence selected from SEQ ID NOs:
- 11, 12, 13 or 14.
A stable pre-fusion Class III fusion protein, or an immunogenic fragment thereof, comprising:
- (a) one or more non-conservative amino acid substitutions within its pre-fusion conformation central helix;
- and/or
- (b) one or more non-conservative amino acid substitutions within amino acid residues corresponding to or aligning with amino acid residues:
- (i) 262 to 293 of the RVG sequence of SEQ ID NO:
- 3;
- (ii) 263 to 294 of the VSVG sequence of SEQ ID NO:
- 5;
- (iii) 458 to 500 of the EBV gB sequence of SEQ ID NO:
- 6;
- (iv) 481 to 520 of the CMV gB sequence of SEQ ID NO:
- 8;
- and/or (v) 503 to 545 of the HSV-1 gB sequence of SEQ ID NO:
- 10;
- wherein preferably the one or more non-conservative amino acid substitution is substitution by proline.
The protein or immunogenic fragment thereof of any one of the preceding claims, which further comprises one or more additional mutations in a region other than the central extended helix of the post-fusion conformation trimerisation interface.
The protein or immunogenic fragment thereof of any one of the preceding claims, which induces neutralising antibodies against one or more epitope of the pre-fusion Class III fusion protein.
The protein or immunogenic fragment thereof of any one of the preceding claims, for use in a vaccine.
Dependencies are inferred from the claim text.