Mutant fragments of OspA and methods and uses relating thereto

Patent metadata

Field	Value
Publication number	US-11208439-B2
Application number	US-201815906618-A
Country	US
Kind code	B2
Filing date	Feb 27, 2018
Priority date	Jul 6, 2012
Publication date	Dec 28, 2021
Grant date	Dec 28, 2021

Abstract

The present invention relates to a polypeptide comprising a mutant fragment of an outer surface protein A (OspA), a nucleic acid coding the same, a pharmaceutical composition (particularly for use as a medicament of in a method of treating or preventing a Borrelia infection) comprising the polypeptide and/or the nucleic acid, a method of treating or preventing a Borrelia infection and a method of immunizing a subject.

First claim

What is claimed is:

1. A polypeptide comprising a mutant fragment of a Borrelia outer surface protein A (OspA), and wherein the mutant OspA fragment comprises a C-terminal domain of an OspA of Borrelia afzelii, B. burgdorferi s.s., B. bavariensis , or B. garinii which lacks at least the first 70 N-terminal amino acid residues; and wherein the mutant OspA fragment differs from the corresponding wild-type B. afzelii, B. burgdorferi s.s., B. bavariensis , or B. garinii OspA fragment sequence at least by the introduction of at least one disulfide bond that is formed by the introduction of
i) one cysteine, which forms said disulfide bond with a cysteine residue present within the wild-type fragment; or
ii) two cysteines, which together form said disulfide bond, wherein said two cysteines are substituted for amino acids at
one amino acid at any of positions 182+/−3 and one amino acid at any of positions 269+/−3;
one amino acid at any of positions 182+/−3 and one amino acid at any of positions 272+/−3;
one amino acid at any of positions 244+/−3 and one amino acid at any of positions 259+/−3;
one amino acid at any of positions 141+/−3 and one amino acid at any of positions 241+/−3; and/or
one amino acid at any of positions 165+/−3 and one amino acid at any of positions 265+/−3,
wherein the numbering of the cysteine substitutions corresponds to amino acid positions in the full-length OspA of B. afzelii , K78 as defined by SEQ ID NO: 19 or the homologous amino acids of an OspA from a Borrelia other than B. afzelii.

2. The polypeptide of claim 1 , wherein the mutant OspA fragment has a difference in protective capacity (Δpc) in a Borrelia Challenge Method of at least 50% compared to a placebo control.

3. The polypeptide of claim 1 , wherein the mutant OspA fragment has a difference in protective capacity (Δpc) in a Borrelia Challenge Method of at least 60% compared to a placebo control.

4. The polypeptide of claim 1 , wherein the mutant OspA fragment has a difference in protective capacity (Δpc) in a Borrelia Challenge Method of at least 70% compared to a placebo control.

5. The polypeptide of claim 1 , wherein the mutant OspA fragment has a difference in protective capacity (Δpc) in a Borrelia Challenge Method of at least 80% compared to a placebo control.

6. The polypeptide of claim 1 , wherein the mutant OspA fragment has a difference in protective capacity (Δpc) in a Borrelia Challenge Method of at least 90% compared to a placebo control.

7. The polypeptide of claim 1 , wherein the introduction of the at least one disulfide bond in the OspA C-terminal domain increases the protective capacity (Δpc) of the polypeptide comprising the mutant OspA fragment relative to a polypeptide comprising the respective OspA C-terminal domain without a disulfide bond(s) by at least 10%, when measured in a Borrelia Challenge Method.

8. The polypeptide of claim 1 , wherein said OspA C-terminal domain is from B. burgdorferi s.s., strain B31, OspA serotype 1; wherein the wild-type sequence of OspA from B. burgdorferi s.s., strain B31 is defined by SEQ ID NO: 20.

9. The polypeptide of claim 1 , wherein said mutant OspA fragment is from the OspA C-terminal domain as defined by SEQ ID NO: 179, and comprises at least one introduced disulfide bond between
a) the amino acid at position 182 and the amino acid at position 269;
b) the amino acid at position 182 and the amino acid at position 272;
c) the amino acid at position 244 and the amino acid at position 259;
d) the amino acid at position 141 and the amino acid at position 241; and/or
e) the amino acid at position 165 and the amino acid at position 265;
wherein the numbering corresponds to amino acid positions in the full-length OspA protein as defined by SEQ ID NO: 20.

10. The polypeptide according to claim 1 , wherein said polypeptide comprises one or more features of the group consisting of
a) an E. coli -derived Ipp lipidation signal as defined by MKATKLVLGAVILGSTLLAG (SEQ ID NO: 16);
b) absence of the amino acid sequence GYVLEGTLTAE (SEQ ID NO: 17);
c) a linker peptide led by an N-terminal cysteine residue as a site for lipidation;
d) presence of between 140 and 152 of the most C-terminal contiguous amino acids of the OspA protein; and
e) absence of the N-terminal β-sheet of OspA.

11. The polypeptide of claim 1 , wherein said mutant OspA fragment is from SEQ ID NO: 179 and comprises an introduced disulfide bond between the amino acid at position 182 and the amino acid at position 269, wherein the numbering corresponds to amino acid positions in the full-length OspA protein as defined by SEQ ID NO: 20.

12. A polypeptide comprising at least two mutant OspA fragments as defined in claim 1 .

13. The polypeptide of claim 12 , wherein the polypeptide comprises or consists of a heterodimer selected from the group consisting of Lip-S1D4-S2D4 (SEQ ID NO: 185), Lip-S1D1-S2D1 (SEQ ID NO: 186), Lip-S3D4-S4D4 (SEQ ID NO: 187), Lip-S3D1-S4D1 (SEQ ID NO: 188), Lip-S5D4-S6D4 (SEQ ID NO: 189), Lip-S5D1-S6D1 (SEQ ID NO: 190), Lip-S2D4-S1D4 (SEQ ID NO: 191), Lip-S2D1-S1D1 (SEQ ID NO: 192), Lip-S4D4-53D4 (SEQ ID NO: 193), Lip-S4D1-S3D1 (SEQ ID NO: 194), Lip-S6D4-S5D4 (SEQ ID NO: 195), Lip-S6D1-S5D1 (SEQ ID NO: 196), Lip-S1D4-S2D1 (SEQ ID NO: 197), Lip-S1D1-S2D4 (SEQ ID NO: 198), S3D4-S4D1 (SEQ ID NO: 199), S3D1-S4D4 (SEQ ID NO: 200), S5D4-S6D1 (SEQ ID NO: 201), S5D1-S6D4 (SEQ ID NO: 202), S2D4-S1D1 (SEQ ID NO: 203), S2D1-S1D4 (SEQ ID NO: 204), S4D4-S3D1 (SEQ ID NO: 205), S4D1-S3D4 (SEQ ID NO: 206), S6D4-S5D1 (SEQ ID NO: 207) and Lip-S6D1-S5D4 (SEQ ID NO: 208).

14. A pharmaceutical composition comprising a polypeptide according to claim 1 and optionally a pharmaceutically acceptable carrier or excipient.

15. A pharmaceutical composition comprising at least one of the polypeptides according to claim 13 and optionally a pharmaceutically acceptable carrier or excipient.

16. The pharmaceutical composition according to claim 14 , wherein said pharmaceutically acceptable excipient is L-methionine.

17. The pharmaceutical composition according to claim 14 , wherein said immunostimulatory substance is selected from the group consisting of polycationic polymers, polycationic peptides, immunostimulatory oligodeoxynucleotides (ODNs), oligo(dIdC)13 (SEQ ID NO: 32), peptides containing at least two LysLeuLys motifs, peptide KLKLLLLLKLK (SEQ ID NO: 33), a combination of KLKLLLLLKLK (SEQ ID NO: 33) and oligo(dIdC)13 (SEQ ID NO: 32), polyarginine, neuroactive compounds, human growth hormone, aluminium hydroxide, aluminium phosphate, Freund's complete adjuvant, and Freund's incomplete adjuvant.

18. The polypeptide of claim 1 , wherein said OspA C-terminal domain comprises no more than 152 residues of the most C-terminal contiguous amino acid sequence of the OspA.

19. The polypeptide according to claim 8 , wherein said OspA C-terminal domain further differs from the corresponding wild-type sequence by the replacement of the amino acid sequence provided by SEQ ID NO: 17 with the amino acid sequence provided by SEQ ID NO: 18.

Assignees

• Valneva Austria Gmbh

Inventors

• Comstedt Pär
• Hanner Markus
• Lundberg Urban
• Meinke Andreas
• Schüler Wolfgang
• Wizel Benjamin

Classifications

• Y02A50/30

  Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change · CPC title

• A61K39/02

  Bacterial antigens · CPC title

• A61K2039/55505

  Inorganic adjuvants · CPC title

• C07K14/20 Primary

  from Spirochaetales (O), e.g. Treponema, Leptospira · CPC title

• C12N5/06

  Animal cells or tissues; Human cells or tissues (preservation of excised living parts A01N1/10) · CPC title