# US 10,766,931 B2 · Mutant fragments of OspA and methods and uses relating thereto

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- Number: US 10,766,931 B2
- Document ID: US10766931B2
- Status: Granted
- Kind: B2
- Application number: 15/944,835
- Filed: 2018-04-04
- Published: 2018-12-20
- Granted: 2020-09-08
- Earliest claimed: 2014-01-09
- Assignees: Valneva Austria GmbH
- Applicants: Valneva Austria GmbH
- Inventors: Urban Lundberg; Wolfgang Schüler
- Web page: https://patentgrep.com/patent/US10766931B2

## CPC

- C07K 14/20: from Spirochaetales (O), e.g. Treponema, Leptospira
- A61K 39/0225: {Spirochetes, e.g. Treponema, Leptospira, Borrelia}
- A61K 39/39: characterised by the immunostimulating additives, e.g. chemical adjuvants
- A61P 31/04: Antibacterial agents
- A61P 31/12: Antivirals
- C07K 16/1207: {from Spirochaetales (O), e.g. Treponema, Leptospira}
- A61K 2039/55516: {Proteins; Peptides}
- A61K 2039/70: {Multivalent vaccine}
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/33: Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity
- C07K 2319/00: Fusion polypeptide
- C07K 2319/21: containing a His-tag
- Y02A 50/30: Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- Y02A: TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE

## Abstract

The present invention relates to compositions and methods for the prevention and treatment of Borrelia infection. Particularly, the present invention relates to a polypeptide comprising a hybrid C-terminal fragment of an outer surface protein A (OspA), a nucleic acid coding the same, an antibody specifically binding the same, a pharmaceutical composition (particularly for use as a medicament or in a method of treating or preventing a Borrelia infection) comprising the polypeptide and/or the nucleic acid and/or the antibody, a method of treating or preventing a Borrelia infection and a method of immunizing a subject.

## Claims

### Claim 1

A polypeptide comprising a first disulfide bond-stabilized C-terminal fragment of an outer surface protein (OspA), wherein said first disulfide bond-stabilized C-terminal fragment is a hybrid C-terminal OspA fragment consisting of, from the N- to C-terminal direction, a fusion of a first and a second OspA portion from two different Borrelia strains, wherein said polypeptide induces an immune response protective against a Borrelia infection and wherein
i) said first OspA portion consists of amino acids 125-176 or amino acids 126-175 of OspA from a Borrelia strain that is not B. garinii, strain PBr, and
ii) said second OspA portion consists of amino acids 176-274 or amino acids 177-274 of OspA from B. garinii, strain PBr (SEQ ID NO: 8), wherein the second OspA portion differs from the corresponding wild-type sequence at least by the substitution of the wild-type amino acid at position 182+/−3 of SEQ ID NO: 8 by a cysteine and by the substitution of the wild-type amino acid at position 269+/−3 of SEQ ID NO: 8 by a cysteine and wherein a disulfide bond between the introduced cysteines is present; and
wherein the numbering of the amino acids and of the cysteine substitutions is according to the numbering of corresponding amino acids of the full length OspA of B. burgdorferi s.s., strain B31 (SEQ ID NO: 5).

### Claim 2 (depends on 1)

The polypeptide according to claim 1, wherein said hybrid C-terminal OspA fragment consists of amino acids 125-176 from B. valaisiana, strain VS116, and the disulfide bond-stabilized amino acids 177-274 from Borrelia garinii, strain PBr, as defined by SEQ ID NO: 1.

### Claim 3 (depends on 1)

The polypeptide according to claim 1, wherein said hybrid C-terminal OspA fragment consists of amino acids 126-175 from B. spielmanii and the disulfide bond-stabilized amino acids 177-274 from Borrelia garinii, strain PBr, as defined by SEQ ID NO: 51.

### Claim 4 (depends on 1)

The polypeptide according to claim 1, wherein said second OspA portion comprises a substitution of the threonine residue at amino acid 233 of wild-type OspA of Borrelia garinii, strain PBr, with a proline residue, as defined by SEQ ID NO: 7.

### Claim 5 (depends on 1)

The polypeptide according to claim 1, further comprising a second disulfide bond-stabilized C-terminal OspA fragment; wherein said second disulfide bond-stabilized C-terminal OspA fragment consists of a C-terminal domain of an OspA from B. burgdorferi s.s., B. afzelii, B. bavariensis, or B. garinii, which differs from the corresponding wild-type OspA sequence at least by the introduction of at least one disulfide bond, and wherein said second disulfide bond-stabilized C-terminal OspA fragment is not a hybrid C-terminal OspA fragment.

### Claim 6 (depends on 5)

The polypeptide according to claim 5, wherein said at least one disulfide bond is formed by the substitution of the amino acid at position 182+/−3 of the wild-type sequence by a cysteine and by the substitution of the amino acid at position 269+/−3 of the wild-type sequence by a cysteine; and wherein the numbering of said amino acids is according to the numbering of corresponding amino acids of the full length OspA of B. burgdorferi s.s., strain B31 (SEQ ID NO: 5).

### Claim 7 (depends on 1)

The polypeptide according to claim 1,
i) wherein the polypeptide is lipidated or wherein the polypeptide comprises an E. coli-derived lpp lipidation signal as defined by MKATKLVLGAVILGSTLLAG (SEQ ID NO: 15); and/or
ii) wherein the polypeptide comprises a lipidation site peptide led by an N-terminal cysteine residue as a site for lipidation, wherein said lipidation site peptide is defined by amino acids CSS; and/or
iii) wherein the polypeptide comprises a linker between the hybrid C-terminal OspA fragment and the second C-terminal OspA fragment, wherein said linker comprises GTSDKNNGSGSKEKNKDGKYS (SEQ ID NO: 16).

### Claim 8 (depends on 1)

The polypeptide according to claim 1, further comprising a second hybrid C-terminal fragment of OspA.

### Claim 9 (depends on 1)

The polypeptide according to claim 1, wherein said polypeptide consists of the heterodimer of Lip-S4D1-S3hybD1 (SEQ ID NO: 27).

### Claim 10 (depends on 1)

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

### Claim 11 (depends on 10)

The pharmaceutical composition according to claim 10, wherein said composition additionally comprises Lip-S1D1-S2D1 (SEQ ID NO: 29) and/or Lip-S5D1-S6D1 (SEQ ID NO: 33).

### Claim 12 (depends on 10)

The pharmaceutical composition according to claim 10, wherein said pharmaceutically acceptable excipient is L-methionine and/or alum.

### Claim 13 (depends on 1)

A method of treating or protecting against a Borrelia infection in a subject in need thereof comprising the step of administering to the subject a therapeutically-effective amount of a polypeptide according to claim 1.

### Claim 14 (depends on 10)

A method of treating or protecting against a Borrelia infection in a subject in need thereof comprising the step of administering to the subject a therapeutically-effective amount of a pharmaceutical composition according to claim 10.

### Claim 15 (depends on 13)

The method according to claim 13, wherein the Borrelia is selected from the group consisting of B. burgdorferi, B. garinii, B. afzelii, B. andersoni, B. bavariensis, B. bissettii, B. valaisiana, B. lusitaniae, B. spielmanii, B. japonica, B. tanukii, B. turdi, and B. sinica.

### Claim 16 (depends on 14)

The method according to claim 14, wherein the Borrelia is selected from the group consisting of B. burgdorferi, B. garinii, B. afzelii, B. andersoni, B. bavariensis, B. bissettii, B. valaisiana, B. lusitaniae, B. spielmanii, B. japonica, B. tanukii, B. turdi, and B. sinica.

### Claim 17 (depends on 5)

The polypeptide according to claim 5, wherein said second disulfide bond-stabilized C-terminal OspA fragment consists of the amino acids from position 125, 126, 130, or 131 to position 273 of wild-type OspA of B. afzelii strain K78, serotype 2 (SEQ ID NO: 6) or the homologous fragments of OspA from B. burgdorferi s.s., B. garinii or B. bavariensis, differing from the corresponding wild-type OspA by the introduction of at least one disulfide bond.

### Claim 18 (depends on 5)

The polypeptide according to claim 5, wherein said second disulfide bond-stabilized C-terminal OspA fragment from B. burgdorferi s.s. does not comprise the human leukocyte function-associated (hLFA-1)-like sequence, defined by the amino acid sequence GYVLEGTLTAE (SEQ ID NO: 17).

### Claim 19 (depends on 18)

The polypeptide according to claim 18, wherein said hLFA-1-like sequence is replaced by the homologous sequence from B. afzelii as defined by SEQ ID NO: 18.