Methods and compositions for preventing oxidative degradation of proteins

• US 20050276823A1
• Filed: 07/11/2003
• Published: 12/15/2005
• Est. Priority Date: 07/12/2002
• Status: Abandoned Application

• Current Assignee: Medarex Incorporated (Bristol-Myers Squibb Company)
• Original Assignee: Medarex Incorporated (Bristol-Myers Squibb Company)
• Inventors: Cini, John K., Nagi, Athena D.
• Application Number:
  US10/618,350
• Publication Number:
  US 20050276823A1
• Time in Patent Office: Days
• US Class Current: 424/400
• CPC Class Codes: A61K 47/18 Amines; Amides; Ureas; Quaternary ammonium compounds; Amino acids; Oligopeptides having up to five amino acids
• CPC Class Codes: A61K 47/183 Amino acids, e.g. glycine, EDTA or aspartame
• CPC Class Codes: A61K 47/20 containing sulfur, e.g. dimethyl sulfoxide [DMSO], docusate, sodium lauryl sulfate or aminosulfonic acids
• CPC Class Codes: A61K 47/26 Carbohydrates, e.g. sugar alcohols, amino sugars, nucleic acids, mono-, di- or oligo-saccharides; Derivatives thereof, e.g. polysorbates, sorbitan fatty acid esters or glycyrrhizin
• 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: C08K 5/0091 Complexes with metal-heteroatom-bonds
• CPC Class Codes: C08K 5/175 containing COOH-groups; Esters or salts thereof

Abstract

Methods and compositions for preventing oxidative damage to proteins, particularly antibodies, are provided. The compositions include a combination of metal chelators, such as DTPA, EGTA, and/or DEF, and can further include one or more free radical scavengers, particularly scavengers of oxygen radicals. Methods for enhancing protein stability using the compositions of the invention are also disclosed.

Claims

1. A composition comprising a protein formulated with DTPA and another agent selected from the group consisting of DEF, mannitol, methionine, and histidine.

2. The composition of claim 1, comprising DEF.

3. The composition of claim 1, further comprising EGTA.

4. The composition of claim 1, wherein the concentration of DTPA is from about 1 μ

M to about 10 mM.

5. The composition of claim 2, wherein the concentration of DEF is from about 1 μ

M to about 5 mM.

6. The composition of claim 1, comprising mannitol at a concentration of about 0.01% to about 25%.

7. The composition of claim 1, comprising methionine at a concentration of about 10 μ

M to about 200 mM.

8. The composition of claim 1, comprising histidine at a concentration of about 100 μ

M to about 200 mM.

9. The composition of claim 1, further comprising an agent that inhibits protein aggregation.

10. The composition of claim 9, wherein the agent that inhibits protein aggregation is selected from the group consisting of polysorbate 80, polysorbate 20, glycerol, and a poloxamer polymer.

11. The composition of claim 10, wherein the agent that inhibits protein aggregation is polysorbate 80 or polysorbate 20 at a concentration of from about 0.001% to about 0.1%.

12. The composition of claim 1, further comprising a buffer that maintains the pH of the composition from about 5.0 to about 8.0.

13. The composition of claim 12, wherein the buffer is selected from the group consisting of phosphate, citrate, Tris, acetate, MES, succinic acid, PIPES, Bis-Tris, MOPS, ACES, BES, TES, HEPES, EPPS, ethylenediamine, phosphoric acid, and maleic acid.

14. The composition of claim 1, comprising mannitol, a polysorbate, Tris, and sodium chloride, wherein the protein is an antibody or fragment thereof.

15. The composition of claim 1, wherein the concentration of the protein is from about 1 μ

g/mL to about 500 mg/mL.

16. The composition of claim 1, wherein the protein is an antibody, or a fragment thereof.

17. The composition of claim 16, wherein the antibody is a monoclonal antibody, or a fragment thereof.

18. The composition of claim 16, wherein the antibody is a human antibody, or a fragment thereof.

19. The composition of claim 16, wherein the antibody is conjugated to an agent, selected from the group consisting of a toxin, a polymer, an imaging agent and a drug.

20. The composition of claim 1, wherein the protein is microencapsulated.

21. The composition of claim 1, wherein the composition is a pharmaceutical composition.

22. A composition comprising a protein formulated with EGTA and DEF.

23. A method for preparing a stabilized protein composition, comprising formulating a protein together with DTPA and another agent selected from the group consisting of DEF, mannitol, methionine, and histidine.

24. The method of claim 23, comprising DEF.

25. The method of claim 23, wherein the composition further comprises EGTA.

26. The method of claim 23, wherein the concentration of DTPA or EGTA is from about 1 μ

M to about 10 mM.

27. The method of claim 24, wherein the concentration of DEF is from about 1 μ

M to about 5 mM DEF.

28. The method of claim 23, wherein the oxidation protective compound is selected from the group consisting of about 0.01% to about 25% mannitol, about 10 μ

M to about 200 mM histidine, and about 10 μ

M to about 200 mM methionine.

29. The method of claim 23, further comprising adding an agent that inhibits protein aggregation to the composition.

30. The method of claim 23, further comprising adding a buffer that maintains the pH from about 5.0 to about 8.0 to the composition.

31. The method of claim 30, wherein the buffer is selected from the group consisting of about 5 mM to about 100 mM phosphate, citrate, Tris, acetate, MES, succinic acid, PIPES, Bis-Tris, MOPS, ACES, BES, TES, HEPES, EPPS, ethylenediamine, phosphoric acid, and maleic acid.

33. The method of claim 23, wherein the concentration of the protein is from about 1 μ

g/mL to about 500 mg/mL.

34. The method of claim 23, wherein the protein is an antibody, or a fragment thereof.

35. The method of claim 34, wherein the antibody is a human antibody, or a fragment thereof.

36. The method of claim 34, wherein the antibody is a monoclonal antibody, or a fragment thereof.

37. The method of claim 34, wherein the antibody is conjugated to an agent selected from a toxin, a polymer, an imaging agent or a drug.

38. The method of claim 23, wherein the protein is microencapsulated.

39. The method of claim 23, wherein the composition is a pharmaceutical composition.

40. The method of claim 23, wherein the protein is protected against oxidation.

32. The method of calim 23, wherein the composition comprises mannitol, a polysorbate, Tris, and sodium chloride, wherein the protein is an antibody or a fragment thereof.

41. A method for preparing a stabilized protein composition, comprising formulating a protein together with EGTA and DEF.

42. The method of claim 41, wherein the protein is protected against oxidation.