MULTISPECIFIC ANTIBODIES

• US 20170349669A1
• Filed: 05/15/2017
• Published: 12/07/2017
• Est. Priority Date: 07/29/2014
• Status: Abandoned Application

• Current Assignee: Hoffmann-La Roche Incorporated (Roche Holding AG)
• Original Assignee: Hoffmann-La Roche Incorporated (Roche Holding AG)
• Inventors: IMHOF-JUNG, SABINE, KLEIN, CHRISTIAN, KLOSTERMANN, STEFAN, REGULA, JOERG THOMAS, SCHAEFER, WOLFGANG
• Application Number:
  US15/595,325
• Publication Number:
  US 20170349669A1
• Time in Patent Office: Days
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 16/00 Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies antibodies with enzymatic activity, e.g. abzymes C12N9/0002
• CPC Class Codes: C07K 16/22 against growth factors ; against growth regulators
• CPC Class Codes: C07K 16/244 Interleukins [IL]
• CPC Class Codes: C07K 16/2863 against receptors for growth factors, growth regulators
• CPC Class Codes: C07K 16/2875 against the NGF/TNF superfamily, e.g. CD70, CD95L, CD153, CD154 against NGF C07K16/22, against TNF C07K16/241
• CPC Class Codes: C07K 16/32 against translation products of oncogenes
• CPC Class Codes: C07K 16/468 Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/31 multispecific
• CPC Class Codes: C07K 2317/35 Valency
• CPC Class Codes: C07K 2317/41 Glycosylation, sialylation, or fucosylation
• CPC Class Codes: C07K 2317/515 : Complete light chain, i.e. VL + CL
• CPC Class Codes: C07K 2317/522 : CH1 domain
• CPC Class Codes: C07K 2317/526 : CH3 domain
• CPC Class Codes: C07K 2317/56 : variable (Fv) region, i.e. VH and/or VL
• CPC Class Codes: C07K 2317/66 : comprising a swap of domains, e.g. CH3-CH2, VH-CL or VL-CH1
• CPC Class Codes: C07K 2317/92 : Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• CPC Class Codes: C07K 2317/94 : Stability, e.g. half-life, pH, temperature or enzyme-resistance

Abstract

The present invention relates to multispecific antibodies, methods for their production, pharmaceutical compositions containing said antibodies and uses thereof.

Claims

1. A multispecific antibody, comprising

a first light chain and a first heavy chain derived from a first antibody which specifically binds to a first antigen;

and

a second light chain and a second heavy chain derived from a second antibody which specifically binds to a second antigen, wherein;

in the second light chain, the variable domain VL is replaced by the variable domain VH of the second heavy chain and the constant domain CL is replaced by the constant domain CH1 of the second heavy chain; and

in the second heavy chain the variable domain VH is replaced by the variable domain VL of the second light chain and the constant domain CH1 is replaced by the constant domain CL of the second light chain; and

wherein in the constant domain CL of the first light chain the amino acids at position 124 and 123 (numbering according to Kabat) are substituted independently from each other by an amino acid selected from lysine (K), arginine (R) and histidine (H); and

wherein in the constant domain CH1 of the first heavy chain the amino acids at position 147 and 213 (numbering according to EU index of Kabat) are substituted independently from each other by an amino acid selected from glutamic acid (E) or aspartic acid (D);

or

wherein in the constant domain CL of the second heavy chain the amino acids at position 124 and 123 (numbering according to Kabat) are substituted independently from each other by an amino acid selected from lysine (K), arginine (R) and histidine (H); and

wherein in the constant domain CH1 of the second light chain the amino acids at position 147 and 213 (numbering according to EU index of Kabat) are substituted independently from each other by an amino acid selected from glutamic acid (E) or aspartic acid (D).

2. The multispecific antibody according to claim 1,

wherein in the constant domain CL of the first light chain the amino acids at position 124 and 123 (numbering according to Kabat) are substituted independently from each other by an amino acid selected from lysine (K), arginine (R) and histidine (H), and wherein in the constant domain CH1 of the first heavy chain the amino acids at position 147 and 213 (numbering according to EU index of Kabat) are substituted independently from each other by an amino acid selected from glutamic acid (E) or aspartic acid (D), and wherein in the constant domain CL of the second heavy chain the amino acid at position 124 (numbering according to Kabat) is substituted by an amino acid selected from glutamic acid (E) or aspartic acid (D);

or

wherein in the constant domain CL of the second heavy chain the amino acids at position 124 and 123 (numbering according to Kabat) are substituted independently from each other by an amino acid selected from lysine (K), arginine (R) and histidine (H), and wherein in the constant domain CH1 of the second light chain the amino acids at position 147 and 213 (numbering according to EU index of Kabat) are substituted independently from each other by an amino acid selected from glutamic acid (E) or aspartic acid (D), and wherein

in the constant domain CL of the first light chain the amino acid at position 124 (numbering according to Kabat) is substituted by an amino acid selected from glutamic acid (E) or aspartic acid (D).

3. The multispecific antibody according to claim 1,

wherein in the variable domain VL of the first light chain the amino acid at position 38 (numbering according to Kabat) is substituted by an amino acid selected from lysine (K), arginine (R) and histidine (H);

and

wherein in the variable domain VH of the first heavy chain the amino acid at position 39 (numbering according to Kabat) is substituted by an amino acid selected from glutamic acid (E) or aspartic acid (D); and

wherein in the variable domain VL of the second heavy chain the amino acid at position 38 (numbering according to Kabat) is substituted by an amino acid selected from glutamic acid (E) or aspartic acid (D); and

wherein in the variable domain VH of the second light chain the amino acid at position 39 (numbering according to Kabat) is substituted by an amino acid selected from lysine (K), arginine (R) and histidine (H).

4. The multispecific antibody according to claim 1, wherein

in the tertiary structure of the antibody the CH3 domain of the first heavy chain and the CH3 domain of the second heavy chain form an interface that is located between the respective antibody CH3 domains,

wherein the respective amino acid sequences of the CH3 domain of the first heavy chain and the CH3 domain of the second heavy chain each comprise a set of amino acids that is located within said interface in the tertiary structure of the antibody,

wherein from the set of amino acids that is located in the interface in the CH3 domain of one heavy chain at least one amino acid residue is substituted by an amino acid residue having a larger side chain volume than the original amino acid residue,

thereby generating a protuberance within the interface, wherein the protuberance is located in the CH3 domain of the one heavy chain, and wherein the protuberance is positionable in a cavity located in the CH3 domain of the other heavy chain within the interface;

and

wherein from the set of amino acids that is located in the interface in the CH3 domain of the other heavy chain at least one amino acid residue is substituted by an amino acid residue having a smaller side chain volume than the original amino acid residue,

thereby generating a cavity within the interface, wherein the cavity is located in the CH3 domain of the other heavy chain, and wherein in the cavity the protuberance within the interface located in the CH3 domain of the one heavy chain is positionable.

5. The multispecific antibody according to claim 4, wherein

said amino acid residue having a larger side chain volume than the original amino acid residue is selected from arginine (R), phenylalanine (F), tyrosine (Y) and tryptophan (W);

and

wherein said amino acid residue having a smaller side chain volume than the original amino acid residue is selected from alanine (A), serine (S), threonine (T) and valine (V).

6. The multispecific antibody according to claim 4, wherein:

from the set of amino acids that is located in the interface in the CH3 domain of the one heavy chain a first amino acid is substituted by cysteine (C); and

from the set of amino acids that is located in the interface in the CH3 domain of the other heavy chain a second amino acid is substituted by cysteine (C), wherein the second amino acid is facing the first amino acid within the interface;

such that a disulfide bridge between the CH3 domain of the one heavy chain and the CH3 domain of the other heavy chain can be formed via the introduced cysteine residues.

7. The multispecific antibody according to claim 1 that specifically binds to human Angiopoietin-2 and human VEGF, wherein:

the antibody comprises a variable heavy chain domain (VH) according to SEQ ID NO;

32 and a variable light chain domain (VL) according to SEQ ID NO;

33; and

the antibody comprises a variable heavy chain domain (VH) according to SEQ ID NO;

31 and a variable light chain domain (VL) according to SEQ ID NO;

30.

8. The multispecific antibody according to claim 1 that specifically binds to human TWEAK and human IL17, wherein:

the antibody comprises a variable heavy chain domain (VH) according to SEQ ID NO;

36 and a variable light chain domain (VL) according to SEQ ID NO;

37; and

the antibody comprises a variable heavy chain domain (VH) according to SEQ ID NO;

35 and a variable light chain domain (VL) according to SEQ ID NO;

34.

9. The multispecific antibody according to claim 1 that specifically binds to human Her1, human Her3 and human cMet, wherein:

the antibody comprises a variable heavy chain domain (VH) according to SEQ ID NO;

40 and a variable light chain domain (VL) according to SEQ ID NO;

41; and

the antibody comprises a variable heavy chain domain (VH) according to SEQ ID NO;

39 and a variable light chain domain (VL) according to SEQ ID NO;

38.

10. A method for the preparation of a multispecific antibody according to claim 1, comprising the steps of

a) transforming a host cell with vectors comprising nucleic acids encoding:

i) the first light chain as defined in claim 1 derived from a first antibody which specifically binds to a first antigen;

ii) the first heavy chain as defined in claim 1 derived from a first antibody which specifically binds to a first antigen;

iii) the second light chain as defined in claim 1 derived from a second antibody which specifically binds to a second antigen; and

iv) the second heavy chain as defined in claim 1 derived from a second antibody which specifically binds to a second antigen;

b) culturing said host cell under conditions that allow synthesis of said multispecific antibody; and

c) recovering said multispecific antibody from said host cell culture.

11. A nucleic acid encoding the amino acid sequence of

a first light chain as defined in claim 1,

a second light chain as defined in claim 1,

a first heavy chain as defined in claim 1;

or

a second heavy chain as defined in claim 1.

12. A vector comprising a nucleic acid according to claim 11, wherein the vector is capable of expressing said nucleic acid in a host cell.

13. A host cell comprising

one or a plurality of vectors comprising:

a) a first polynucleotide encoding the first light chain as defined in claim 1 derived from a first antibody which specifically binds to a first antigen;

b) a second polynucleotide encoding the first heavy chain as defined in claim 1 derived from a first antibody which specifically binds to a first antigen;

c) a third polynucleotide encoding the second light chain as defined in claim 1 derived from a second antibody which specifically binds to a second antigen; and

d) a fourth polynucleotide encoding the second heavy chain as defined in claim 1 derived from a second antibody which specifically binds to a second antigen.

14. A pharmaceutical composition comprising a multispecific antibody according to claim 1 in combination with at least one pharmaceutically acceptable carrier.

16. A method of treatment administering an effective amount of the bispecific antibody of claim 1 to an individual in need thereof.

17. A multispecific antibody obtained by a method according to claim 10.

15. (canceled)