Readily isolated bispecific antibodies with native immunoglobulin format

• US 8,586,713 B2
• Filed: 06/25/2010
• Issued: 11/19/2013
• Est. Priority Date: 06/26/2009
• Status: Active Grant

• Current Assignee: Regeneron Pharmaceuticals Incorporated
• Original Assignee: Regeneron Pharmaceuticals Incorporated
• Inventors: Davis, Samuel, Smith, Eric, MacDonald, Douglas, Olson, Kara Louise
• Primary Examiner(s): Goddard, Laura B.
• Assistant Examiner(s): Natarajan, Meera
• Application Number:
  US12/823,838
• Publication Number:
  US 20100331527A1
• Time in Patent Office: 1,243 Days
• Field of Search: None
• US Class Current: 530/387.3
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: C07K 1/22 Affinity chromatography or related techniques based upon selective absorption processes
• CPC Class Codes: C07K 16/247 IL-4
• CPC Class Codes: C07K 16/248 IL-6
• CPC Class Codes: C07K 16/2809 against the T-cell receptor (TcR)-CD3 complex
• CPC Class Codes: C07K 16/2866 against receptors for cytokines, lymphokines, interferons
• CPC Class Codes: C07K 16/2887 against CD20
• CPC Class Codes: C07K 16/468 Immunoglobulins having two or more different antigen binding sites, e.g. multifunctional antibodies
• CPC Class Codes: C07K 2317/21 from primates, e.g. man
• CPC Class Codes: C07K 2317/31 multispecific
• CPC Class Codes: C07K 2317/52 Constant or Fc region; Isotype
• 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 2317/76 Antagonist effect on antigen, e.g. neutralization or inhibition of binding

Abstract

A bispecific antibody format providing ease of isolation is provided, comprising immunoglobulin heavy chain variable domains that are differentially modified in the CH3 domain, wherein the differential modifications are non-immunogenic or substantially non-immunogenic with respect to the CH3 modifications, and at least one of the modifications results in a differential affinity for the bispecific antibody for an affinity reagent such as Protein A, and the bispecific antibody is isolable from a disrupted cell, from medium, or from a mixture of antibodies based on its affinity for Protein A.

Claims

1. A bispecific antigen-binding protein that is heterodimeric with respect to Protein A binding, comprising:

a. a first polypeptide comprising, from N-terminal to C-terminal, a first epitope-binding region that selectively binds a first epitope, an immunoglobulin constant region that comprises a first CH3 region of a human IgG selected from IgG1, IgG2, and IgG4, wherein the first CH3 region binds to Protein A; and

b. a second polypeptide comprising, from N-terminal to C-terminal, a second epitope-binding region that selectively binds a second epitope, an immunoglobulin constant region that comprises a second CH3 region of a human IgG selected from IgG1, IgG2, and IgG4, wherein the second CH3 region comprises a modification that reduces or eliminates binding of the second CH3 region to Protein A.

2. The bispecific protein of claim 1, wherein the first polypeptide and the second polypeptide are human IgG heavy chains.

3. The bispecific protein of claim 1, further comprising an immunoglobulin light chain.

4. The bispecific protein of claim 3, wherein the immunoglobulin light chain is a human immunoglobulin light chain.

5. The bispecific protein of claim 1, wherein the first and the second polypeptides are each human IgG1 heavy chains.

6. The bispecific protein of claim 1, wherein the modification is selected from the group consisting of (a) 95R, and (b) 95R and 96F in the IMGT exon numbering system, or (a′

) 435R, and (b′

) 435R and 436F in the EU numbering system.

7. The bispecific protein of claim 6, further comprising one to five modifications selected from the group consisting of 16E, 18M, 44S, 52N, 57M, and 82I in the IMGT exon numbering system, or 356E, 358M, 384S, 392N, 397M, and 422I In the EU numbering system.

8. The bispecific protein of claim 6, wherein the CH3 region of the bispecific antibody is non-immunogenic or substantially non-immunogenic in a human.