# US 2022/0033482 A1 · Alpha-synuclein single domain antibodies

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- Number: US 2022/0033482 A1
- Document ID: US20220033482A1
- Status: Published application
- Kind: A1
- Application number: 16/971,157
- Filed: 2019-02-19
- Published: 2022-02-03
- Earliest claimed: 2018-02-19
- Assignees: New York University
- Applicants: New York University
- Inventors: Einar M. SIGURDSSON
- Web page: https://patentgrep.com/patent/US20220033482A1

## CPC

- C07K 16/18: against material from animals or humans
- G01N 33/6896: {Neurological disorders, e.g. Alzheimer's disease}
- C07K 2317/21: from primates, e.g. man
- C07K 2317/22: from camelids, e.g. camel, llama or dromedary
- C07K 2317/24: containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
- C07K 2317/569: Single domain, e.g. dAb, sdAb, VHH, VNAR or nanobody®
- G01N 2800/28: Neurological disorders
- G01N 2800/56: Staging of a disease; Further complications associated with the disease
- 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)
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

The present disclosure is directed to single domain antibodies that bind to α-synuclein and the use of these antibodies for the treatment and diagnosis of α-synucleinopathies The present disclosure is also directed to polynucleotides encoding the α-synuclein single domain antibodies, therapeutic vectors comprising these polynucleotides and methods of administering these therapeutic vectors for the treatment of α-synucleinopathies.

## Claims

### Claim 1

An antibody or fragment thereof, said antibody or binding fragment thereof comprising a heavy chain variable region, wherein said heavy chain variable region comprises:
a complementarity-determining region 1 (H-CDR1) comprising an amino acid sequence of any one of SEQ ID NOs: 1-36, 218, 222, 226, 230, 234, 238, 340 or a modified amino acid sequence of any one of SEQ ID NOs: 1-36, 218, 222, 226, 230, 234, 238, and 340 said modified sequence containing 1, 2, or 3 amino acid residue modifications as compared to any one of SEQ ID NOs: 1-36, 218, 222, 226, 230, 234, 238, and 340;
a complementarity-determining region 2 (H-CDR2) comprising an amino acid sequence of any one of SEQ ID NOs: 37-72, 219, 223, 227, 231, 235, 239, 341 or a modified amino acid sequence of any one of SEQ ID NOs: 37-72, 219, 223, 227, 231, 235, 239, and 341 said modified sequence containing 1, 2, or 3 amino acid residue modifications as compared to any one of SEQ ID NOs: 37-72, 219, 223, 227, 231, 235, 239, and 341; and
a complementarity-determining region 3 (H-CDR3) comprising an amino acid sequence of any one of SEQ ID NOs: 73-108, 220, 224, 228, 232, 236, 240, 342 or a modified amino acid sequence of any one of SEQ ID NOs: 73-108, 220, 224, 228, 232, 236, 240, and 342 said modified sequence containing 1, 2, or 3 amino acid residue modifications as compared to any one of SEQ ID NOs: 73-108, 220, 224, 228, 232, 236, 240, 342.

### Claim 2

(canceled)

### Claim 3 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said antibody or binding fragment thereof binds to human α-synuclein protein.

### Claim 4 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said antibody or binding fragment thereof is a camelid antibody or binding fragment thereof.

### Claim 5 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said antibody or binding fragment thereof is a monoclonal antibody or binding fragment thereof.

### Claim 6 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said heavy chain variable region comprises:
the H-CDR1 comprising the amino acid sequence of SEQ ID NO: 18, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid residue modifications as compared to SEQ ID NO: 18;
the H-CDR2 comprising the amino acid sequence of SEQ ID NO: 54, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2, or 3 amino acid residue modifications as compared to SEQ ID NO: 54; and
the H-CDR3 comprising the amino acid sequence of SEQ ID NO: 90, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid modifications as compared to SEQ ID NO: 90.

### Claim 7 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said heavy chain variable region comprises:
the H-CDR1 comprising the amino acid sequence of SEQ ID NO: 33, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid residue modifications as compared to SEQ ID NO: 33;
the H-CDR2 comprising the amino acid sequence of SEQ ID NO: 69, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2, or 3 amino acid residue modifications as compared to SEQ ID NO: 69; and
the H-CDR3 comprising the amino acid sequence of SEQ ID NO: 105, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid modifications as compared to SEQ ID NO: 105.

### Claim 8 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said heavy chain variable region comprises:
the H-CDR1 comprising the amino acid sequence of SEQ ID NO: 36, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid residue modifications as compared to SEQ ID NO: 36;
the H-CDR2 comprising the amino acid sequence of SEQ ID NO: 72, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2, or 3 amino acid residue modifications as compared to SEQ ID NO: 72; and
the H-CDR3 comprising the amino acid sequence of SEQ ID NO: 108, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid modifications as compared to SEQ ID NO: 108.

### Claim 9 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said heavy chain variable region comprises:
the H-CDR1 comprising the amino acid sequence of SEQ ID NO: 218, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid residue modifications as compared to SEQ ID NO: 218;
the H-CDR2 comprising the amino acid sequence of SEQ ID NO: 219, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2, or 3 amino acid residue modifications as compared to SEQ ID NO: 219; and
the H-CDR3 comprising the amino acid sequence of SEQ ID NO: 220, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid modifications as compared to SEQ ID NO: 220.

### Claim 10 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said heavy chain variable region further comprises human or a humanized immunoglobulin heavy chain framework regions.

### Claim 11 (depends on 10)

The antibody or binding fragment thereof of claim 10, wherein the heavy chain variable region comprises an amino acid sequence that is at least 80% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 109-145, 221, 225, 229, 233, 237, 241, and 343.

### Claim 12

(canceled)

### Claim 13 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 109-145, 221, 225, 229, 233, 237, 241, and 343.

### Claim 14 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said antibody or binding fragment thereof is selected from the group consisting of: a single domain antibody (Fv), a minibody, a scFv, and a bispecific antibody.

### Claim 15

An antibody or fragment thereof, said antibody or binding fragment thereof comprising a heavy chain variable region, wherein said heavy chain variable region comprises:
a complementarity-determining region 1 (H-CDR1) comprising an amino acid sequence of any one of SEQ ID NOs: 146-160, or a modified amino acid sequence of any one of SEQ ID NOs: 146-160, said modified sequence containing 1, 2, or 3 amino acid residue modifications as compared to any one of SEQ ID NOs: 146-160;
a complementarity-determining region 2 (H-CDR2) comprising an amino acid sequence of any one of SEQ ID NOs: 161-175, or a modified amino acid sequence of any one of SEQ ID NOs: 161-175, said modified sequence containing 1, 2, or 3 amino acid residue modifications as compared to any one of SEQ ID NOs: 161-175; and
a complementarity-determining region 3 (H-CDR3) comprising an amino acid sequence of any one of SEQ ID NOs: 176-190, or a modified amino acid sequence of any one of SEQ ID NOs: 176-190, said modified sequence containing 1, 2, or 3 amino acid residue modifications as compared to any one of SEQ ID NOs: 176-190.

### Claim 16

(canceled)

### Claim 17 (depends on 15)

The antibody or binding fragment thereof of claim 15, wherein said antibody or binding fragment thereof binds to human α-synuclein protein.

### Claim 18 (depends on 15)

The antibody or binding fragment thereof of claim 15, wherein said antibody or binding fragment thereof is a camelid antibody or binding fragment thereof.

### Claim 19 (depends on 15)

The antibody or binding fragment thereof of claim 15, wherein said antibody or binding fragment thereof is a monoclonal antibody or binding fragment thereof.

### Claim 20 (depends on 15)

The antibody or binding fragment thereof of claim 15, wherein said heavy chain variable region comprises:
the H-CDR1 comprising the amino acid sequence of SEQ ID NO: 147, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid residue modifications as compared to SEQ ID NO: 147;
the H-CDR2 comprising the amino acid sequence of SEQ ID NO: 162, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2, or 3 amino acid residue modifications as compared to SEQ ID NO: 162; and
the H-CDR3 comprising the amino acid sequence of SEQ ID NO: 177, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid modifications as compared to SEQ ID NO: 177.

### Claim 21 (depends on 15)

The antibody or binding fragment thereof of claim 15, wherein said heavy chain variable region comprises:
the H-CDR1 comprising the amino acid sequence of SEQ ID NO: 151, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid residue modifications as compared to SEQ ID NO: 151;
the H-CDR2 comprising the amino acid sequence of SEQ ID NO: 166, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2, or 3 amino acid residue modifications as compared to SEQ ID NO: 166; and
the H-CDR3 comprising the amino acid sequence of SEQ ID NO: 181, or a modified amino acid sequence thereof, said modified amino acid sequence containing 1, 2 or 3 amino acid modifications as compared to SEQ ID NO: 181.

### Claim 22 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said heavy chain variable region further comprises human or a humanized immunoglobulin heavy chain framework regions.

### Claim 23 (depends on 22)

The antibody or binding fragment thereof of claim 22, wherein the heavy chain variable region comprises an amino acid sequence that is at least 80% identical to an amino acid sequence selected from the group consisting of SEQ ID NOs: 191-217.

### Claim 24

(canceled)

### Claim 25 (depends on 15)

The antibody or binding fragment thereof of claim 15, wherein the heavy chain variable region comprises an amino acid sequence selected from the group consisting of SEQ ID NOs: 191-217.

### Claim 26 (depends on 1)

The antibody or binding fragment thereof of claim 1, wherein said antibody or binding fragment thereof is selected from the group consisting of: a single domain antibody (Fv), a minibody, a scFv, and a bispecific antibody.

### Claim 27 (depends on 1)

A polyclonal antibody composition comprising two or more antibodies or fragments thereof of claim 1.

### Claim 28 (depends on 1)

An isolated polynucleotide encoding the antibody or binding fragment thereof of claim 1.

### Claim 29 (depends on 28)

A vector comprising the isolated polynucleotide of claim 28.

### Claim 30 (depends on 29)

The vector of claim 29, wherein said vector is an adeno-associated viral vector.

### Claim 31 (depends on 29)

A host cell comprising the vector of claim 29.

### Claim 32 (depends on 1)

A pharmaceutical composition comprising:
the antibody or binding fragment thereof of claim 1 and
a pharmaceutical carrier.

### Claim 33 (depends on 32)

A method of inhibiting onset of one or more symptoms of an α-synucleinopathy in a subject, said method comprising:
administering to the subject the pharmaceutical composition of claim 32, wherein said composition is administered in an amount effective to inhibit onset of one or more symptoms of the α-synucleinopathy in the subject.

### Claim 34 (depends on 32)

A method of treating an α-synucleinopathy in a subject, said method comprising:
administering to the subject the pharmaceutical composition of claim 32, wherein said composition is administered in an amount effective to treat the α-synucleinopathy in the subject.

### Claim at position 35

-36. (canceled)

### Claim 37 (depends on 1)

A method of diagnosing an α-synucleinopathy in a subject, said method comprising:
detecting, in the subject, the presence of accumulated α-synuclein protein or peptide using the antibody or binding fragment thereof of claim 1, and
diagnosing the α-synucleinopathy in the subject based on said detecting.

### Claim at position 37

-40. (canceled)

### Claim 41 (depends on 1)

A method of monitoring the progression of an α-synucleinopathy in a subject, said method comprising:
detecting, in the subject, the presence of α-synuclein protein or peptide using the antibody or binding fragment thereof of claim 1;
repeating said detecting periodically; and
monitoring the progression of the α-synucleinopathy in the subject based on said repeated detecting.

### Claim at position 39

-45. (canceled)

### Claim 46 (depends on 1)

A diagnostic kit comprising:
the antibody or binding fragment thereof of claim 1 and
a detectable label.