Identification and engineering of antibodies with variant Fc regions and methods of using same

• US 20070036799A1
• Filed: 08/10/2006
• Published: 02/15/2007
• Est. Priority Date: 08/10/2005
• Status: Active Grant

• Current Assignee: MacroGenics, Inc.
• Original Assignee: MacroGenics, Inc.
• Inventors: Rankin, Christopher, Gorlatov, Sergey, Stavenhagen, Jeffrey, Tuaillon, Nadine
• Granted Patent: US 8,217,147 B2
• Time in Patent Office: Days
• US Class Current: 424/155.100
• CPC Class Codes: A61K 2039/505 comprising antibodies
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C07K 16/283 against Fc-receptors, e.g. CD16, CD32, CD64 CD23 C07K16/2851
• CPC Class Codes: C07K 16/2887 against CD20
• CPC Class Codes: C07K 16/32 against translation products of oncogenes
• CPC Class Codes: C07K 16/44 against material not provided for elsewhere , e.g. haptens, metals, DNA, RNA, amino acids
• CPC Class Codes: C07K 2317/24 containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
• CPC Class Codes: C07K 2317/52 Constant or Fc region; Isotype
• CPC Class Codes: C07K 2317/72 Increased effector function due to an Fc-modification
• CPC Class Codes: C07K 2317/732 Antibody-dependent cellular cytotoxicity [ADCC]
• CPC Class Codes: C07K 2317/734 Complement-dependent cytotoxicity [CDC]
• CPC Class Codes: C07K 2317/76 Antagonist effect on antigen, e.g. neutralization or inhibition of binding
• CPC Class Codes: C07K 2317/92 Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
• 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
• CPC Class Codes: G01N 33/574 for cancer

Abstract

The present invention relates to molecules, particularly polypeptides, more particularly immunoglobulins (e.g., antibodies), comprising a variant Fc region, wherein said variant Fc region comprises at least one amino acid modification relative to a wild-type Fc region, which variant Fc region binds FcγRIIIA and/or FcγRIIA with a greater affinity, relative to a comparable molecule comprising the wild-type Fc region. The molecules of the invention are particularly useful in preventing, treating, or ameliorating one or more symptoms associated with a disease, disorder, or infection. The molecules of the invention are particularly useful for the treatment or prevention of a disease or disorder where an enhanced efficacy of effector cell function (e.g., ADCC) mediated by FcγR is desired, e.g., cancer, infectious disease, and in enhancing the therapeutic efficacy of therapeutic antibodies the effect of which is mediated by ADCC.

Claims

1. A polypeptide comprising a variant Fc region, wherein said variant Fc region comprises at least one amino acid modification relative to a wild-type Fc region, such that said polypeptide binds an Fcγ

R with an altered affinity relative to a polypeptide comprising a wild-type Fc region and wherein said at least one amino acid modification comprises a set of substitutions selected from the group consisting of a substitution at position 243 with leucine, at position 292 with proline, at position 300 with leucine, at position 305 with isoleucine, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, at position 300 with leucine, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 300 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 305 with isoleucine;

a substitution at position 243 with leucine and at position 292 with proline;

a substitution at position 243 with leucine; and

a substitution at position 273 with phenylalanine.

4. The polypeptide of claim 1, wherein the Fc region is a human IgG Fc region.

5. The polypeptide of claim 4, wherein the human IgG Fc region is a human IgG1, IgG2, IgG3, or IgG4 Fc region.

6. The polypeptide of claim 1, wherein said polypeptide is an antibody.

7. The antibody of claim 6, wherein said antibody is a monoclonal antibody, a humanized antibody, or a human antibody.

8. The antibody of claim 6, wherein said antibody is a humanized antibody.

9. The antibody of claim 6, wherein said antibody comprises a variable domain which binds to CD16A or CD32B.

10. The antibody of claim 6, wherein said antibody is 2B6.

13. A nucleic acid comprising a nucleotide sequence encoding the polypeptide of claim 1.

14. A vector comprising the nucleic acid of claim 13.

15. The vector of claim 14 which is an expression vector.

16. A host cell comprising the nucleic acid of claim 15.

17. A method for recombinantly producing the polypeptide of claim 1, said method comprising:

(i) culturing in a medium a host cell comprising a nucleic acid encoding said polypeptide under conditions suitable for the expression of said polypeptide; and

(ii) recovery of said polypeptide from said medium.

18. The polypeptide of claim 1 which is a therapeutic antibody that specifically binds a cancer antigen, said therapeutic antibody comprising a variant Fc region, wherein said variant Fc region comprises at least one amino acid modification relative to a wild-type Fc region, such that said therapeutic antibody specifically binds Fcγ

RIIIA with a greater affinity than the therapeutic antibody comprising the wild-type Fc region binds Fcγ

RIIIA and wherein said at least one amino acid modification comprises a set of substitutions selected from the group consisting of a substitution at position 243 with leucine, at position 292 with proline, at position 300 with leucine, at position 305 with isoleucine, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, at position 300 with leucine, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 300 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 305 with isoleucine;

a substitution at position 243 with leucine and at position 292 with proline;

a substitution at position 243 with leucine; and

a substitution at position 273 with phenylalanine.

19. The polypeptide of claim 1 which is a therapeutic antibody that specifically binds a cancer antigen, said therapeutic antibody comprising a variant Fc region, wherein said variant Fc region comprises at least one amino acid modification relative to a wild-type Fc region, such that said therapeutic antibody specifically binds Fcγ

RIIIA with a greater affinity than a therapeutic antibody comprising the wild-type Fc region binds Fcγ

RIIIA, and said therapeutic antibody further specifically binds Fcγ

RIIB with a lower affinity than a therapeutic antibody comprising the wild-type Fc region binds Fcγ

RIIB, and wherein said at least one amino acid modification comprises a set of substitutions selected from the group consisting of a substitution at position 243 with leucine, at position 292 with proline, and at position 300 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 305 with isoleucine;

a substitution at position 243 with leucine and at position 292 with proline;

a substitution at position 243 with leucine; and

a substitution at position 273 with phenylalanine.

23. A method of treating cancer in a patient having a cancer characterized by a cancer antigen, said method comprising administering to said patient a therapeutically effective amount of a therapeutic antibody of claim 18 that binds said cancer antigen.

29. A pharmaceutical composition comprising a therapeutically effective amount of the polypeptide of claim 1, and a pharmaceutically acceptable carrier.

39. A method for recombinantly producing the antibody of claim 6, said method comprising:

(i) culturing in a medium a host cell comprising

(a) a nucleic acid comprising the nucleotide sequence encoding the light chain of said antibody; and

(b) a nucleic acid comprising the nucleotide sequence encoding the heavy chain of said antibody,

under conditions suitable for the expression of said antibody; and

(ii) recovery of said antibody from said medium.

40. A method of treating or managing cancer in a patient having a cancer characterized by a cancer antigen, said method comprising administering to said patient a therapeutically effective amount of the antibody of claim 6 that binds said cancer antigen.

42. The antibody of claim 6, wherein said antibody has an enhanced ADCC activity relative to a comparable antibody comprising the wild type Fc region.

43. The polypeptide of claim 1, wherein said variant Fc region further comprises one or more of the amino acid modifications listed in table 5, 6, 7, or 8.

2. A polypeptide comprising a variant Fc region, wherein said variant Fc region comprises at least one amino acid modification relative to a wild-type Fc region, such that said polypeptide specifically binds Fcγ

RIIIA with a greater affinity than a comparable polypeptide comprising the wild-type Fc region binds Fcγ

RIIIA and wherein said at least one amino acid modification comprises a set of substitutions selected from the group consisting of a substitution at position 243 with leucine, at position 292 with proline, at position 300 with leucine, at position 305 with isoleucine, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, at position 300 with leucine, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 300 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 305 with isoleucine;

a substitution at position 243 with leucine and at position 292 with proline;

a substitution at position 243 with leucine; and

a substitution at position 273 with phenylalanine.

3. A polypeptide comprising a variant Fc region, wherein said variant Fc region comprises at least one amino acid modification relative to a wild-type Fc region, such that said polypeptide specifically binds Fcγ

RIIIA with a greater affinity than a comparable polypeptide comprising the wild-type Fc region binds Fcγ

RIIIA, and said polypeptide further specifically binds Fcγ

RIIB with a lower affinity than a comparable polypeptide comprising the wild-type Fc regions binds Fcγ

RIIB, and wherein said at least one amino acid modification comprises a set of substitutions selected from the group consisting of a substitution at position 243 with leucine, at position 292 with proline, and at position 300 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 396 with leucine;

a substitution at position 243 with leucine, at position 292 with proline, and at position 305 with isoleucine;

a substitution at position 243 with leucine and at position 292 with proline;

a substitution at position 243 with leucine; and

a substitution at position 273 with phenylalanine.

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