Low acidic species compositions and methods for producing and using the same

Patent metadata

Field	Value
Publication number	US-9266949-B2
Application number	US-201414575691-A
Country	US
Kind code	B2
Filing date	Dec 18, 2014
Priority date	Oct 18, 2013
Publication date	Feb 23, 2016
Grant date	Feb 23, 2016

Abstract

The instant invention relates to low acidic species (AR) compositions comprising a protein, e.g., an antibody, or antigen-binding portion thereof, and methods, e.g., cell culture and/or protein purification methods, for producing such low AR compositions. Methods for using such compositions to treat a disorder, e.g., a disorder in which TNFα is detrimental, are also provided.

First claim

The invention claimed is:

1. A method for producing a composition comprising an immunoglobulin comprising the 6 CDR domains of adalimumab, the method comprising:
culturing a mammalian cell producing an immunoglobulin comprising the 6 CDR domains of adalimumab in a cell culture media comprising 2 g/L to 11 g/L of each of one or more basic amino acids selected from the group consisting of arginine, lysine, ornithine and histidine, and combinations thereof, to produce a composition comprising an immunoglobulin comprising the 6 CDR domains of adalimumab, wherein the composition comprises less than 20% total acidic species of the immunoglobulin, and
wherein the acidic species of the immunoglobulin correspond to the peaks that elute earlier than the main peak in a WCX-10 HPLC chromatogram of the immunoglobulin, and wherein the WCX-10 HPLC chromatogram is generated using a first mobile phase of 10 mM Sodium Phosphate dibasic (pH 7.5) and a second mobile phase of 10 mM Sodium Phosphate dibasic, 500 mM Sodium Chloride (pH 5.5), and wherein the WCX-10 HPLC chromatogram is generated using detection at 280 nm.

2. The method of claim 1 , wherein the immunoglobulin is adalimumab.

3. The method of claim 2 , wherein the amino acid concentration in the cell culture media is 3 g/L to 11 g/L.

4. The method of claim 2 , wherein the amino acid concentration in the cell culture media is 3 g/L to 8 g/L.

5. The method of claim 2 , wherein the cell culture media further comprises one or more of calcium and niacinamide.

6. The method of claim 2 , wherein the composition comprises 9% to 15% total acidic species of adalimumab.

7. The method of claim 2 , wherein the composition comprises less than 10% total acidic species of adalimumab.

8. The method of claim 2 , wherein the composition comprises less than 12% total acidic species of adalimumab.

9. The method of claim 2 , wherein the pH of the cell culture media is 6.9 or lower.

10. The method of claim 2 , wherein the mammalian host cell is selected from the group consisting of a CHO cell, an NSO cell, a COS cell, and an SP2 cell.

11. The method of claim 10 , wherein the mammalian host cell is a CHO cell.

12. The method of claim 2 , wherein the one or more amino acids is arginine.

13. The method of claim 2 , wherein the one or more amino acids is lysine.

14. The method of claim 2 , wherein the one or more amino acids is histidine.

15. The method of claim 2 , wherein the one or more amino acids is ornithine.

16. The method of claim 2 , wherein the one or more amino acids are arginine, lysine, histidine, and ornithine.

17. The method of claim 2 , further comprising isolating the adalimumab.

18. A method for producing a composition comprising an immunoglobulin comprising the 6 CDR domains of adalimumab, the method comprising culturing a mammalian cell capable of producing an immunoglobulin comprising the 6 CDR domains of adalimumab in cell culture media comprising 2 g/L to 11 g/L of each one or more basic amino acids selected from the group consisting of arginine, lysine, ornithine, and histidine, and combinations thereof,
wherein the one or more amino acids is present in the cell culture media at a concentration sufficient to produce a composition that has at least 10% fewer acidic species of an immunoglobulin comprising the 6 CDR domains of adalimumab than a composition comprising an immunoglobulin comprising the 6 CDR domains of adalimumab produced by culturing the cell in a cell culture media comprising less than 2 g/L of each of the one or more basic amino acids, and
wherein the acidic species of the immunoglobulin correspond to the peaks that elute earlier than the main peak in a WCX-10 HPLC chromatogram of the immunoglobulin, and wherein the WCX-10 HPLC chromatogram is generated using a first mobile phase of 10 mM Sodium Phosphate dibasic (pH 7.5) and a second mobile phase of 10 mM Sodium Phosphate dibasic, 500 mM Sodium Chloride (pH 5.5), and wherein the WCX-10 HPLC chromatogram is generated using detection at 280 nm.

19. The method of claim 18 , wherein the cell culture media comprises 3 g/L to 11 g/L of each of the one or more amino acids.

20. The method of claim 18 , wherein said immunoglobulin is adalimumab.

21. The method of claim 20 , wherein said mammalian cell is a CHO cell.

22. The method of claim 20 , wherein the one or more amino acids is arginine.

23. The method of claim 20 , wherein the one or more amino acids is lysine.

24. The method of claim 20 , wherein the one or more amino acids is histidine.

25. The method of claim 20 , wherein the one or more amino acids is ornithine.

26. The method of claim 20 , wherein said culture media is production media.

27. The method of claim 20 , wherein said culture media is growth media.

28. The method of claim 20 , wherein said mammalian cell is cultured to a maximum viable cell density (VCD) of at least 7×10 6 cells/ml.

29. The method of claim 20 , further comprising isolating the adalimumab.

30. The method of claim 20 , wherein the composition comprises less than 10% total acidic species of adalimumab.

Assignees

• Abbvie Inc

Inventors

• Ramasubramanyan Natarajan
• Yang Lihua
• Herigstad Matthew Omon
• Yang Hong
• Subramanian Kartik
• Zeng Xiaobei
• Dong Diane D
• Lim Wen Chung
• Gifford Kathreen A
• Kaymakcalan Zehra
• Chumsae Christopher

Classifications

• A61K47/26

  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 title

• A61K47/183

  Amino acids, e.g. glycine, EDTA or aspartame · CPC title

• C07K16/00

  Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies · CPC title

• C07K2317/21

  from primates, e.g. man · CPC title

• A61K9/19

  lyophilised {, i.e. freeze-dried, solutions or dispersions (lyophilised products with subsequent particle size reduction A61K9/14; granules or pellets made by lyphilisation A61K9/1682; solid oral dosage forms made by lyophilisation A61K9/2095; lyophilisation additives A61K47/00)} · CPC title