ANTIBODY-BASED THERAPEUTICS WITH ENHANCED ADCC ACTIVITY

• US 20070092521A1
• Filed: 10/20/2006
• Published: 04/26/2007
• Est. Priority Date: 10/21/2005
• Status: Active Grant

• Current Assignee: Genzyme Corporation (Sanofi )
• Original Assignee: Genzyme Corporation (Sanofi )
• Inventors: Zhou, Qun, Edmunds, Tim, McPherson, John
• Granted Patent: US 7,700,321 B2
• Time in Patent Office: Days
• US Class Current: 424/155.1
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 1/04 for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 13/12 of the kidneys
• CPC Class Codes: A61P 15/00 Drugs for genital or sexual disorders for disorders of sex hormones A61P5/24; Contraceptives
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/06 Antipsoriatics
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 21/04 for myasthenia gravis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• 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 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 5/14 of the thyroid hormones, e.g. T3, T4
• CPC Class Codes: A61P 7/06 Antianaemics
• CPC Class Codes: A61P 9/00 : Drugs for disorders of the cardiovascular system
• CPC Class Codes: A61P 9/04 : Inotropic agents, i.e. stimulants of cardiac contraction; Drugs for heart failure
• 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 16/30 : from tumour cells
• CPC Class Codes: C07K 16/3023 : Lung
• CPC Class Codes: C07K 2317/41 : Glycosylation, sialylation, or fucosylation
• 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 2317/92 : Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value

Abstract

Methods for producing antibody-based therapeutics with enhanced ADCC activity are disclosed. The enhanced ADCC activity is attributed to oligomannose-type N-glycans on the antibodies and Fc fusion proteins of the invention. Also disclosed are methods of using such antibody-based therapeutics for targeted killing of cells in a mammal, including therapeutic methods of treating cancers, autoimmune diseases and other diseases.

Claims

1. A pharmaceutical composition comprising a monoclonal antibody or Fc fusion protein comprising an oligomannose-type N-glycan.

2. The composition of claim 1, wherein the monoclonal antibody or Fc fusion protein comprises an Fc domain derived from a human antibody.

3. The composition of claim 1, wherein the monoclonal antibody is human or humanized.

4. The composition of claim 1, wherein the monoclonal antibody is directed against a human antigen.

5. The composition of claim 2, wherein the human antibody from which the Fc domain is derived is of a class selected from the group consisting of IgG, IgA, IgD, IgE, and IgM.

6. The composition of claim 2, wherein the human antibody from which the Fc domain is derived is of an isotype selected from the group consisting of IgG1, IgG2, IgG3, and IgG4.

7. The composition of claim 6, wherein the Fc domain comprises an amino acid sequence which is at least 80% identical to SEQ ID NO:

n over the entire length of SEQ ID NO;

n, wherein n=1, 2, 3, or 4.

8. The composition of claim 7, wherein the Fc domain comprises an amino acid sequence which is at least 90% identical to SEQ ID NO:

1 over the entire length of SEQ ID NO;

1.

9. The composition of claim 1, wherein the oligomannose-type N-glycans comprise an oligosaccharide selected from the group consisting of Man₉(GIcNAc)₂, Man₈(GIcNAc)₂, Man₇(GIcNAc)₂, Man₆(GIcNAc)₂, and Man₅(GIcNAc)₂.

10. The composition of claim 1, wherein at least 20% of all N-glycans in the composition are Man_5-9(GIcNAc)₂.

11. The composition of claim 1, wherein the antibody or Fc fusion protein has higher ADCC activity as compared to the same antibody or Fc fusion protein with complex-type N-glycans.

12. The composition of claim 1, wherein the antibody or Fc fusion protein has higher binding affinity for Fcγ

RIIIA as compared the same antibody or Fc fusion protein with complex-type N-glycans.

13. A method of making the pharmaceutical composition of claim 1, comprising:

(a) providing a cell engineered to express the antibody or Fc fusion protein;

(b) culturing the cell under conditions resulting in secretion of the antibody or Fc fusion protein comprising oligomannose-type N-glycans; and

(c) recovering the secreted antibody or Fc fusion protein.

14. The method of claim 13, wherein the engineered cell is deficient in a glycosidase selected from the group consisting of α

-glucosidase I, α

-glucosidase II, and α

-mannosidase I.

15. The method of claim 13, wherein the engineered cell is contacted with an inhibitor of a glycosidase selected from the group consisting of α

-glucosidase I, α

-glucosidase II, and α

-mannosidase I.

16. The method of claim 15, wherein the inhibitor is a glucosidase I inhibitor selected from the group consisting of castanospermine, DNJ, N-alkyl-DNJ, DMDP, and australine.

17. The method of claim 15, wherein the inhibitor is a glucosidase I inhibitor selected from the group consisting of DNJ, N-alkyl-DNJ, N-alkenyl-DNJ, and MDL 25637.

18. The method of claim 15, wherein the inhibitor is a specific inhibitor of α

-mannosidase I.

19. The method of claim 15, wherein the specific inhibitor of α

-mannosidase I is kifunensine.

20. The method of claim 15, wherein the inhibitor is a α

-mannosidase I inhibitor selected from the group consisting of DMJ, N-methyl-DMJ, DIM, and kifunensine.

23. The method of claim 20, wherein the cell is incubated in the presence of 0.01 to 20 μ

g/ml kifunensine.

24. A method of killing a target cell in a mammal comprising administering to the mammal a pharmaceutical composition of claim 1, thereby killing the target cell.

25. The method of claim 24, wherein the target cell is a cancerous cell, an infected cell, or a cell of the immune system.

26. The method of claim 24, wherein the mammal is human.

27. The method of claim 24, wherein the mammal has a disease selected from the group consisting of cancer, infectious disease, inflammatory disease, and immune-mediated disease.

21. A method of making an antibody or Fc fusion protein, comprising:

(a) providing a cell engineered to express the antibody or Fc fusion protein;

(b) culturing the cell in the presence of kifunensine; and

(c) recovering the secreted antibody or Fc fusion protein,

wherein the antibody or Fc fusion protein comprises an Fc domain derived from IgG1.

22. The method of claim 21, wherein the mammalian cell is a hybridoma, a CHO cell or an NS0 cell.

28. Use of a monoclonal antibody or Fc fusion protein comprising an oligomannose-type N-glycan in the manufacture of a medicament for treatment of a disease in which ADCC-mediated killing of target cells is desired.