# US 2017/0247475 A1 · Bispecific binding molecules binding to vegf and ang2

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- Number: US 2017/0247475 A1
- Document ID: US20170247475A1
- Status: Published application
- Kind: A1
- Application number: 15/350,364
- Filed: 2016-11-14
- Published: 2017-08-31
- Earliest claimed: 2011-04-01
- Assignees: Boehringer Ingelheim International GmbH
- Applicants: Boehringer Ingelheim International GmbH
- Inventors: Andreas GSCHWIND; Rene Georg OTT; Joachim BOUCNEAU; Marie-Ange BUYSE; Erik DEPLA
- Web page: https://patentgrep.com/patent/US20170247475A1

## CPC

- A61P 5/00: Drugs for disorders of the endocrine system
- C07K 16/468: {Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies}
- A61P 9/00: Drugs for disorders of the cardiovascular system
- A61P 27/02: Ophthalmic agents
- A61P 35/00: Antineoplastic agents
- C07K 16/18: against material from animals or humans
- C07K 16/22: against growth factors {; against growth regulators}
- C07K 2317/22: from camelids, e.g. camel, llama or dromedary
- C07K 2317/31: multispecific
- C07K 2317/33: Crossreactivity, e.g. for species or epitope, or lack of said crossreactivity
- C07K 2317/565: Complementarity determining region [CDR]
- C07K 2317/567: Framework region [FR]
- C07K 2317/569: Single domain, e.g. dAb, sdAb, VHH, VNAR or nanobody®
- C07K 2317/62: comprising only variable region components
- C07K 2317/73: Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
- 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
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- 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)

## Abstract

Bispecifc binding molecules binding to both VEGF and Ang2, preferably in the form of immunoglobulin single variable domains like VHHs and domain antibodies, pharmaceutical compositions containing the same and their use in the treatment of diseases that are associated with VEGF- and/or Ang2-mediated effects on angiogenesis are disclosed. Further, nucleic acids encoding bispecific binding molecules, host cells and methods for preparing same are also described.

## Claims

### Claim at position 1

1-26. (canceled)

### Claim 27

An Ang2-binding VHH with an amino acid sequence selected from acid sequences shown in SEQ ID NOs: 214, 215, 216, 217, 218, 219, 220, 221, 222, or 223.

### Claim 28 (depends on 27)

An immunoglobulin single variable domain which has been obtained by humanization of a VHH defined in claim 27.

### Claim at position 4

29-37. (canceled)

### Claim 38

A method of treating a disease that is associated with VEGF- and/or Ang2-mediated effects on angiogenesis comprising administering to a patient an effective amount of a pharmaceutical composition comprising at least one binding molecule as the active ingredient and a pharmaceutically acceptable carrier, diluent or excipient, wherein the said binding molecule comprises:
at least one VEGF-binding component,
at least one serum albumin binding component, and
at least one Angiopoietin-2 (Ang2) binding component,
wherein said VEGF-, serum albumin and Ang2-binding components are immunoglobulin single variable domains, each immunoglobulin single variable domain consisting of four framework regions and three complementarity determining regions (CDRs), and
wherein:
said VEGF binding immunoglobulin single variable domain, comprises the following CDR sequences:
(SEQ ID NO: 450)
CDR1: SYSMG
(SEQ ID NO: 465)
CDR2: AISKGGYKYDAVSLEG
(SEQ ID NO: 4)
CDR3: SRAYGSSRLRLADTYEY
said serum albumin binding immunoglobulin single variable domain, comprises the following CDR sequences:
(SEQ ID NO: 255)
CDR1: SFGMS
(SEQ ID NO: 256)
CDR2: SISGSGSDTLYADSVKG
(SEQ ID NO: 257)
CDR3: GGSLSR;
and
said Ang2-binding immunoglobulin single variable domain, comprises the following CDR sequences:
(SEQ ID NO: 248)
CDR1: DYAIG
(SEQ ID NO: 49)
CDR2: AIRSSGGSTYYADSVKG
(SEQ ID NO: 250)
CDR3: VPAGRLRYGEQWYPIYEYDA.

### Claim 39 (depends on 38)

The method of claim 38 wherein the disease is selected from cancer and cancerous diseases.

### Claim 40 (depends on 38)

The method of claim 38 wherein the disease is eye diseases.

### Claim 41

An isolated nucleic acid molecule encoding a bispecific binding molecule, wherein said binding molecule comprises:
at least one VEGF-binding component,
at least one serum albumin binding component, and
at least one Angiopoietin-2 (Ang2) binding component,
wherein said VEGF-, serum albumin and Ang2-binding components are immunoglobulin single variable domains, each immunoglobulin single variable domain consisting of four framework regions and three complementarity determining regions (CDRs), and
wherein:
said VEGF binding immunoglobulin single variable domain, comprises the following CDR sequences:
(SEQ ID NO: 450)
CDR1: SYSMG
(SEQ ID NO: 465)
CDR2: AISKGGYKYDAVSLEG
(SEQ ID NO: 4)
CDR3: SRAYGSSRLRLADTYEY
said serum albumin binding immunoglobulin single variable domain, comprises the following CDR sequences:
(SEQ ID NO: 255)
CDR1: SFGMS
(SEQ ID NO: 256)
CDR2: SISGSGSDTLYADSVKG
(SEQ ID NO: 257)
CDR3: GGSLSR;
and
said Ang2-binding immunoglobulin single variable domain, comprises the following CDR sequences:
(SEQ ID NO: 248)
CDR1: DYAIG
(SEQ ID NO: 249)
CDR2: AIRSSGGSTYYADSVKG
(SEQ ID NO: 250)
CDR3: VPAGRLRYGEQWYPIYEYDA.

### Claim 42

An isolated nucleic acid molecule encoding a bispecific binding molecule, wherein said bispecific binding molecule comprises:
a VEGF-binding VHH, comprising the following CDR sequences:
(SEQ ID NO: 450)
CDR1: SYSMG
(SEQ ID NO: 465)
CDR2: AISKGGYKYDAVSLEG
(SEQ ID NO: 4)
CDR3: SRAYGSSRLRLADTYEY
a serum albumin binding VHH, comprising the following CDR sequences:
(SEQ ID NO: 255)
CDR1: SFGMS
(SEQ ID NO: 256)
CDR2: SISGSGSDTLYADSVKG
(SEQ ID NO: 257)
CDR3: GGSLSR;
and
an Ang2-binding VHH, comprising the following CDR sequences:
(SEQ ID NO: 248)
CDR1: DYAIG
(SEQ ID NO: 249)
CDR2: AIRSSGGSTYYADSVKG
(SEQ ID NO: 250)
CDR3: VPAGRLRYGEQWYPIYEYDA.

### Claim 43 (depends on 42)

The isolated nucleic acid molecule of claim 42, wherein said VEGF-binding VHH comprised in said bispecific binding molecule has the amino acid sequence shown in SEQ ID NO: 57.

### Claim 44 (depends on 42)

The isolated nucleic acid molecule of claim 42, wherein said serum albumin binding VHH comprised in said bispecific binding molecule has the amino acid sequence shown in SEQ ID NO: 254.

### Claim 45 (depends on 42)

The isolated nucleic acid molecule of claim 42, wherein said Ang2-binding VHH comprised in said bispecific binding molecule has the amino acid sequence shown in SEQ ID NO: 222.

### Claim 46 (depends on 42)

The isolated nucleic acid molecule of claim 42, wherein said serum albumin binding VHH has the amino acid sequence shown in SEQ ID NO: 254 and wherein the said Ang2-binding VHH has the amino acid sequence shown in SEQ ID NO: 222.

### Claim 47

An isolated nucleic acid molecule encoding a bispecific binding molecule comprising the amino acid sequence shown in SEQ ID NO: 207.

### Claim 48 (depends on 41)

An expression vector containing a nucleic acid molecule of claim 41.

### Claim 49 (depends on 48)

A host cell containing an expression vector of claim 48.

### Claim 50 (depends on 49)

A method of manufacturing a bispecific binding molecule comprising the steps of:
culturing a host cell according to claim 49 under conditions that allow expression of said bispecific binding molecule; and
recovering or isolating said bispecific binding molecule expressed by said host cell from the culture.

### Claim 51 (depends on 50)

The method of claim 50, additionally comprising the step of:
further purifying or modifying or formulating said bispecific binding molecule.