Tri- or tetraspecific antibodies

• US 8,796,424 B2
• Filed: 05/27/2010
• Issued: 08/05/2014
• Est. Priority Date: 05/27/2009
• Status: Active Grant

• Current Assignee: Hoffmann-La Roche Incorporated (Roche Holding AG)
• Original Assignee: Hoffmann-La Roche Incorporated (Roche Holding AG)
• Inventors: Croasdale, Rebecca, Klein, Christian, Schaefer, Wolfgang, Schanzer, Juergen Michael
• Primary Examiner(s): Aeder, Sean
• Assistant Examiner(s): WU, JULIE ZHEN QIN
• Application Number:
  US12/788,967
• Publication Number:
  US 20100322935A1
• Time in Patent Office: 1,531 Days
• Field of Search: None
• US Class Current: 530/387.1
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/22 against growth factors ; against growth regulators
• CPC Class Codes: C07K 16/2863 against receptors for growth factors, growth regulators
• CPC Class Codes: C07K 2317/31 multispecific
• CPC Class Codes: C07K 2317/35 Valency
• CPC Class Codes: C07K 2317/526 CH3 domain
• CPC Class Codes: C07K 2317/64 comprising a combination of variable region and constant region components
• 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

Abstract

The present invention relates to tri- or tetraspecific antibodies, their manufacture and use.

Claims

1. A trispecific or tetraspecific antibody, comprising:

a) a light chain comprising a light chain variable domain VL and a light chain constant domain CL, and a heavy chain comprising a heavy chain variable domain VH and heavy chain constant domains CH1, CH2, and CH3, wherein the light chain and the heavy chain specifically bind to a first antigen; and

b) a modified light chain and a modified heavy chain that specifically bind to a second antigen, wherein;

(i) the modified light chain comprises in N-terminal to C-terminal direction VL, CH1, and the modified heavy chain comprises in N-terminal to C-terminal direction VH, CL, CH2, and CH3;

(ii) the modified light chain comprises in N-terminal to C-terminal direction VH, CL, and the modified heavy chain comprises in N-terminal to C-terminal direction VL, CH1, CH2, and CH3;

or

(iii) the modified light chain comprises in N-terminal to C-terminal direction VH, CH1, and the modified heavy chain comprises in N-terminal to C-terminal direction VL, CL, CH2, and CH3;

and

c) one to four antigen binding peptides fused via a peptide connector to the C- or N-terminus of the light chains or heavy chains of a) and/or b) wherein said antigen binding peptides specifically bind one or two further antigens.

2. The antibody according to claim 1, wherein the antigen binding peptides comprise one or two antigen binding peptides which specifically bind to one or two further antigens.

3. The antibody according to claim 1, wherein the antigen binding peptides comprise one or two antigen binding peptides which specifically bind to a third antigen.

4. The antibody according to claim 1, wherein the antigen binding peptides comprise two identical antigen binding peptides which specifically bind to a third antigen.

5. The antibody according to claim 1, wherein the antigen binding peptides comprise one antigen binding peptide which specifically binds to a third antigen and one other antigen binding peptide which specifically binds to a fourth antigen.

6. The antibody according to claim 1, wherein the antigen binding peptides are selected from the group consisting of a scFv fragment and a scFab fragment.

7. The antibody according to claim 1, wherein the antigen binding peptides are scFv fragments.

8. The antibody according to claim 1, wherein the antigen binding peptides are scFab fragments.

9. The antibody according to claim 1, wherein the antigen binding peptides are fused to the C-terminus of the heavy chains of a) and/or b).

10. The antibody according to claim 1, wherein the CH3 domain of the heavy chain of a) and the CH3 domain of the modified heavy chain of b) meet at an interface which has been altered to promote the formation of the trispecific or tetraspecific antibody, wherein the interface of one CH3 domain comprises a protuberance which is positionable in a cavity in the interface of the other CH3 domain, and wherein

said interface is altered such that:

i) the CH3 domain of one heavy chain is altered, so that within the original interface the CH3 domain of the one heavy chain that meets the original interface of the CH3 domain of the other heavy chain, an amino acid residue is replaced with an amino acid residue having a larger side chain volume, thereby generating a protuberance within the interface of the CH3 domain of the one heavy chain, said protuberance being positionable in a cavity within the interface of the CH3 domain of the other heavy chain; and

ii) the CH3 domain of the other heavy chain is altered, so that within the original interface of the CH3 domain of the other heavy chain that meets the original interface of the CH3 domain of the one heavy chain, an amino acid residue is replaced with an amino acid residue having a smaller side chain volume, thereby generating a cavity within the interface of the CH3 domain of the other heavy chain, said cavity being positionable in a protuberance within the interface of the CH3 domain of the one heavy chain.

11. The antibody according to claim 10, wherein said amino acid residue having a larger side chain volume is selected from the group consisting of arginine (R), phenylalanine (F), tyrosine (Y), tryptophan (W) and said amino acid residue having a smaller side chain volume is selected from the group consisting of alanine (A), serine (S), threonine (T), valine (V).

12. The antibody of claim 10 wherein the protuberance comprises an introduced arginine (R) residue.

13. The antibody of claim 10 wherein the protuberance comprises an introduced phenylalanine (F) residue.

14. The antibody of claim 10 wherein the protuberance comprises an introduced tyrosine (Y) residue.

15. The antibody of claim 10 wherein the protuberance comprises an introduced tryptophan (W) residue.

16. The antibody of claim 10 wherein the cavity is formed by an introduced alanine (A) residue.

17. The antibody of claim 10 wherein the cavity is formed by an introduced serine (S) residue.

18. The antibody of claim 10 wherein the cavity is formed by an introduced threonine (T) residue.

19. The antibody of claim 10 wherein the cavity is formed by an introduced valine (V) residue.

20. The antibody according to claim 10, characterized in that both CH3 domains are further altered by the introduction of a cysteine (C) residue in the corresponding positions of each CH3 domain such that a disulfide bridge between both CH3 domains can be formed.

21. A composition, comprising the antibody according to claim 1.

22. The composition according to claim 21 wherein the composition is a pharmaceutical or diagnostic composition.

23. A pharmaceutical composition comprising an antibody according to claim 1 and at least one pharmaceutically acceptable excipient.

24. The antibody according to claim 1, wherein the modified light chain comprises in N-terminal to C-terminal direction VL, CH1, and the modified heavy chain comprises in N-terminal to C-terminal direction VH, CL, CH2, and CH3.

25. The antibody according to claim 1, wherein the modified light chain comprises in N-terminal to C-terminal direction VH, CL, and the modified heavy chain comprises in N-terminal to C-terminal direction VL, CH1, CH2, and CH3.

26. The antibody according to claim 1, wherein the modified light chain comprises in N-terminal to C-terminal direction VH, CH1, and the modified heavy chain comprises in N-terminal to C-terminal direction VL, CL, CH2, and CH3.