# US 12,760,124 B2 · Trispecific and/or trivalent binding proteins

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- Number: US 12,760,124 B2
- Document ID: US12760124B2
- Status: Granted
- Kind: B2
- Application number: 18/433,295
- Filed: 2024-02-05
- Published: 2024-08-22
- Granted: 2026-10-13
- Earliest claimed: 2016-04-13
- Assignees: Sanofi
- Applicants: Sanofi
- Inventors: Zhi-Yong Yang; Gary J. Nabel; Lan Wu; Edward Seung; Ronnie Wei; Jochen Beninga; Ercole Rao; Wulf Dirk Leuschner; Christian Beil; Christian Lange; Carsten Corvey
- Web page: https://patentgrep.com/patent/US12760124B2

## CPC

- C07K 16/468: {Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies}
- A61K 39/3955: {against proteinaceous materials, e.g. enzymes, hormones, lymphokines}
- A61K 39/39558: {against tumor tissues, cells, antigens}
- A61K 45/06: Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
- C07K 1/22: Affinity chromatography or related techniques based upon selective absorption processes
- C07K 16/241: {Tumor Necrosis Factors}
- C07K 16/244: {Interleukins [IL]}
- C07K 16/247: {IL-4}
- C07K 16/2803: {against the immunoglobulin superfamily}
- C07K 16/2809: {against the T-cell receptor (TcR)-CD3 complex}
- C07K 16/2818: {against CD28 or CD152}
- C07K 16/2887: {against CD20}
- C07K 16/2896: {against molecules with a "CD"-designation, not provided for elsewhere}
- C07K 16/32: against translation products of oncogenes
- A61K 2039/505: {comprising antibodies}
- C07K 2317/31: multispecific
- C07K 2317/35: Valency
- C07K 2317/52: Constant or Fc region; Isotype
- C07K 2317/526: CH3 domain
- C07K 2317/56: variable (Fv) region, i.e. VH and/or VL
- C07K 2317/565: Complementarity determining region [CDR]
- C07K 2317/64: comprising a combination of variable region and constant region components
- C07K 2317/73: Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
- C07K 2317/75: Agonist effect on antigen
- C07K 2317/76: Antagonist effect on antigen, e.g. neutralization or inhibition of binding
- C07K 2317/92: Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
- C07K 2317/94: Stability, e.g. half-life, pH, temperature or enzyme-resistance
- C07K 2319/00: Fusion polypeptide
- 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)

## Abstract

The disclosure provides trispecific and/or trivalent binding proteins comprising four polypeptide chains that form three antigen binding sites that specifically bind one or more target proteins, wherein a first pair of polypeptides forming the binding protein possess dual variable domains having a cross-over orientation and wherein a second pair of polypeptides forming the binding protein possess a single variable domain. The disclosure also provides methods for making trispecific and/or trivalent binding proteins and uses of such binding proteins.

## Claims

### Claim 1

A trivalent and/or trispecific binding protein comprising four polypeptide chains that form three antigen binding sites that specifically bind one or more target proteins, wherein a first polypeptide chain of the binding protein comprises a structure represented by the formula:
and a second polypeptide chain of the binding protein comprises a structure represented by the formula:
and a third polypeptide chain of the binding protein comprises a structure represented by the formula:
and a fourth polypeptide chain of the binding protein comprises a structure represented by the formula:
wherein:
VL1 is a first immunoglobulin light chain variable domain;
VL2 is a second immunoglobulin light chain variable domain;
VL3 is a third immunoglobulin light chain variable domain;
VH1 is a first immunoglobulin heavy chain variable domain;
VH2 is a second immunoglobulin heavy chain variable domain;
VH3 is a third immunoglobulin heavy chain variable domain;
CL is an immunoglobulin light chain constant domain;
CH1 is an immunoglobulin CH1 heavy chain constant domain;
CH2 is an immunoglobulin CH2 heavy chain constant domain;
CH3 is an immunoglobulin CH3 heavy chain constant domain;
hinge is an immunoglobulin hinge region connecting the CH1 and CH2 domains; and
L1, L2, L3 and L4 are each independently amino acid linkers or zero amino acids in length;
wherein the polypeptide of formula I and the polypeptide of formula II form a cross-over light chain-heavy chain pair of the binding protein;
wherein VH1 and VL1 form a binding pair and a first antigen binding site, VH2 and VL2 form a binding pair and a second antigen binding site, and VH3 and VL3 form a binding pair and a third antigen binding site; and
wherein the first antigen binding site specifically binds human IL4 or IL13, and the second antigen binding site specifically binds human IL4 or IL13, the first and second antigen binding sites being different.

### Claim 2 (depends on 1)

An isolated nucleic acid molecule comprising a nucleotide sequence encoding the binding protein of claim 1.

### Claim 3 (depends on 2)

An expression vector comprising the nucleic acid molecule of claim 2.

### Claim 4 (depends on 2)

An isolated host cell comprising the nucleic acid molecule of claim 2.

### Claim 5 (depends on 3)

An isolated host cell comprising the expression vector of claim 3.

### Claim 6 (depends on 1)

A pharmaceutical composition comprising the binding protein of claim 1 and a pharmaceutically acceptable carrier.

### Claim 7 (depends on 6)

A method of treating cancer in a patient comprising administering to the patient a therapeutically effective amount of the pharmaceutical composition of claim 6, wherein the binding protein comprises one antigen binding site that specifically
binds a T-cell surface protein and another antigen binding site that specifically binds a target protein.

### Claim 8 (depends on 7)

The method of claim 7, wherein the patient is a human.

### Claim 9 (depends on 6)

A method of treating an inflammatory disease or disorder in a patient comprising administering to the patient a therapeutically effective amount of the pharmaceutical composition of claim 6, wherein the binding protein comprises three antigen binding sites that each specifically bind a cytokine target protein.

### Claim 10 (depends on 9)

The method of claim 9, wherein the patient is a human.