Expression technology for proteins containing a hybrid isotype antibody moiety

• US 20060263856A1
• Filed: 07/19/2006
• Published: 11/23/2006
• Est. Priority Date: 03/07/2001
• Status: Active Grant

• Current Assignee: Merck Patent GmbH (Merck & Co., Inc.)
• Original Assignee: EMD Lexigen Research Center Corp.
• Inventors: Gillies, Stephen D., Lo, Kin-Ming, Way, Jeffrey
• Granted Patent: US 8,066,994 B2
• Time in Patent Office: Days
• US Class Current: 435/69.700
• CPC Class Codes: C07K 16/30 from tumour cells
• CPC Class Codes: C07K 2317/52 Constant or Fc region; Isotype
• CPC Class Codes: C07K 2317/53 Hinge
• CPC Class Codes: C07K 2319/00 Fusion polypeptide
• CPC Class Codes: C07K 2319/30 Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto

Abstract

Disclosed are methods and compositions for efficiently expressing antibody fusion proteins. Antibody fusion proteins of the invention include a hybrid antibody moiety containing sequences from more than one type of antibody and/or mutant antibody sequences. Hybrid antibody fusion proteins of the invention may be produced at high levels and may combine functional properties characteristic of different antibody types in addition to functional properties of a non-antibody moiety.

Claims

1-16. -16. (canceled)

17. An antibody or Ig fusion protein comprising an IgG2 antibody moiety in which the IgG2 hinge has been mutated to contain two cysteines.

20. A nucleic acid encoding a fusion protein of claim 17, or 29.

21. A stably transfected cell expressing a protein of claim 17, or 29.

22. A method for improving the assembly of an antibody or Ig fusion protein, the method comprising the steps of:

a) replacing the hinge region of said protein with a different hinge region; and

b) testing the assembly of the altered protein.

23. The method of claim 22, wherein said protein and said altered protein are produced by genetic engineering techniques.

24. (canceled)

26. (canceled)

27. A method for improving the production of an antibody or Ig fusion protein from a cell, the method comprising the steps of:

a) replacing the hinge region of said protein with a different hinge region; and

b) testing the production of the altered protein.

28. The method of claim 27, wherein said protein and said altered protein are produced by genetic engineering techniques.

29. A fusion protein comprising an immunoglobulin moiety fused to a non-immunoglobulin moiety, wherein the immunoglobulin moiety comprises an antibody binding site and a heavy chain comprising an IgG1 hinge and an IgG2 CH2 domain.

18. The fusion protein of claim 29 wherein the antibody binding site of the immunoglobulin moiety comprises a variable domain directed against a tumor-associated antigen.

19. The protein of claim 18, comprising an inactive Fc receptor I binding site.

25. A nucleic acid encoding the fusion protein of claim 29.

30. The fusion protein of claim 29, wherein the non-immunoglobulin moiety is a cytokine.

31. The fusion protein of claim 30, wherein the cytokine is interleukin-2.

32. The fusion protein of claim 29, wherein the non-immunoglobulin moiety is erythropoietin.

33. The fusion protein of claim 29, wherein the IgG1 hinge comprises three or fewer cysteines.

34. The fusion protein of claim 29, wherein the non-immunoglobulin moiety is fused to the C-terminus of the immunoglobulin moiety.

35. The fusion protein of claim 29, wherein the non-immunoglobulin moiety is an antibody.