# US 9,518,132 B2 · Protein complexes for antigen binding and methods of use

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- Number: US 9,518,132 B2
- Document ID: US9518132B2
- Status: Granted
- Kind: B2
- Application number: 13/883,282
- Filed: 2011-11-09
- Published: 2013-09-05
- Granted: 2016-12-13
- Earliest claimed: 2010-11-09
- Assignees: Altimab Therapeutics, Inc.
- Applicants: Zijuan Li; Chun Wu; Jun Bao; Bo Chen
- Inventors: Zijuan Li; Chun Wu; Jun Bao; Bo Chen
- Web page: https://patentgrep.com/patent/US9518132B2

## CPC

- C07K 16/468: {Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies}
- A61P 35/00: Antineoplastic agents
- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- C07K 16/248: {IL-6}
- C07K 16/2803: {against the immunoglobulin superfamily}
- C07K 16/2809: {against the T-cell receptor (TcR)-CD3 complex}
- C07K 16/2878: {against the NGF-receptor/TNF-receptor superfamily, e.g. CD27, CD30, CD40, CD95}
- C07K 16/30: from tumour cells
- C07K 16/40: against enzymes
- C07K 16/46: Hybrid immunoglobulins (hybrids of an immunoglobulin with a peptide not being an immunoglobulin C07K19/00)
- G01N 33/6854: {Immunoglobulins}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61K 2039/505: {comprising antibodies}
- C07K 2317/30: characterized by aspects of specificity or valency
- C07K 2317/31: multispecific
- C07K 2317/522: CH1 domain
- C07K 2317/524: CH2 domain
- C07K 2317/526: CH3 domain
- C07K 2317/53: Hinge
- C07K 2317/66: comprising a swap of domains, e.g. CH3-CH2, VH-CL or VL-CH1
- C07K 2317/73: Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
- 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)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- 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)
- 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)

## Abstract

Provided herein in certain embodiments are polypeptide complexes capable of binding to an antigen. Pharmaceutical compositions, method of using the polypeptide complexes are also provided.

## Claims

### Claim 1

A polypeptide complex comprising:
a first polypeptide comprising from N-terminus to C-terminus: a first antigen-binding domain, a first protein monomer, a first thiol residue-containing peptide linker and a second antigen-binding domain, and
a second polypeptide comprising from N-terminus to C-terminus: a third antigen-binding domain, a second protein monomer, a second thiol residue-containing peptide linker and a fourth antigen-binding domain,
wherein the first protein monomer forms a dimer with the second protein monomer, and each of the first and the second protein monomers comprises a CH3 domain from an immunoglobulin;
wherein the first peptide linker and the second peptide linker form a disulfide bond, and each of the first thiol peptide linker and the second peptide linker independently comprises a sequence selected from the group consisting of: SEQ ID NO: 68, SEQ ID NO: 69, and SEQ ID NO: 70;
wherein the first antigen-binding domain and the third antigen-binding domain are capable of binding to a first antigen, and the second antigen-binding domain and the fourth antigen-binding domain are capable of binding to a second antigen, and one of the first and the second antigens is human CD3 and the other is human EpCAM or human CD19, and
wherein each of the first, second, third, and fourth antigen-binding domains is independently selected from the group consisting of VH, VL, Fab, and scFv.

### Claim 2 (depends on 1)

The polypeptide complex of claim 1 wherein the disulfide bond reduces the simultaneous binding of the first antigen-binding domain and the second antigen-binding domain to the first antigen, and of the third antigen-binding domain and the fourth antigen-binding domain to the second antigen.

### Claim 3 (depends on 1)

The polypeptide complex of claim 1 wherein the first protein monomer is the same as the second protein monomer.

### Claim 4 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen-binding domain is the same as the second antigen-binding domain, and/or the third antigen-binding domain is the same as the fourth antigen-binding domain.

### Claim 5 (depends on 1)

The polypeptide complex of claim 1 wherein the first protein monomer and/or the second protein monomer further comprise a CH2 domain from the immunoglobulin and the C-terminal of the CH2 domain is covalently linked to the N-terminal of the CH3 domain.

### Claim 6 (depends on 1)

The polypeptide complex of claim 1 wherein the immunoglobulin is selected from the group consisting of IgA, IgD, IgE, IgG, and IgM.

### Claim 7 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen-binding domain or the second antigen-binding domain is Fab comprising a first light chain fragment which is disulfidely bonded to a first heavy chain fragment.

### Claim 8 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen-binding domain or the second antigen-binding domain is Fab comprising a second heavy chain fragment which is disulfidely bonded to a second light chain fragment.

### Claim 9 (depends on 1, 2, 3, 4, 5, 6, 7, 8)

A composition comprising the polypeptide complex of any of claims 1, 2-4, 5, 6, 7-8 and one or more conjugates, wherein the polypeptide complex is linked to the one or more conjugates.

### Claim 10 (depends on 1, 2, 3, 4, 5, 6, 7, 8)

A pharmaceutical composition comprising the polypeptide complex of any of claims 1, 2-4, 5, 6, 7-8, and a pharmaceutically acceptable carrier.

### Claim 11 (depends on 1)

The polypeptide complex of claim 1 wherein the first and second antigen-binding domains are Fab and the third and fourth binding domains are scFv.

### Claim 12 (depends on 1)

The polypeptide complex of claim 1 wherein the third and fourth antigen-binding domains are scFv and the first and second binding domains are Fab.

### Claim 13 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen is human EpCAM or human CD19, and the second antigen is human CD3.

### Claim 14 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen is human CD3, and the second antigen is human EpCAM or human CD19.

### Claim 15 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen-binding domain is anti-CD3 Fab, and the second antigen-binding domain is anti-CD19 scFv.

### Claim 16 (depends on 1)

The polypeptide complex of claim 1 wherein the first antigen-binding domain is anti-CD3 Fab, and the second antigen-binding domain is anti-EpCAM scFv.