Cell culture media and methods of antibody production

Patent metadata

Field	Value
Publication number	US-9441035-B2
Application number	US-201414211467-A
Country	US
Kind code	B2
Filing date	Mar 14, 2014
Priority date	Mar 15, 2013
Publication date	Sep 13, 2016
Grant date	Sep 13, 2016

Abstract

Cell culture media are provided herein as are methods of using the media for cell culture and antibody production from cells. Compositions comprising antibodies and fragments thereof, produced by the methods herein are also provided.

First claim

What is claimed is:

1. A method of producing bevacizumab, or a fragment thereof, comprising the step of culturing a Chinese hamster ovary (CHO) cell comprising a nucleic acid encoding bevacizumab or fragment thereof in a cell culture medium, wherein the cell culture medium comprises copper, insulin, and cystine, wherein the cystine is at a concentration of from 1.25 mM to 2.5 mM, and wherein the cell produces bevacizumab, or a fragment thereof.

2. The method of claim 1 , wherein the cell culture medium further comprises a plant peptone, a porcine peptone, or both a plant peptone and a porcine peptone.

3. The method of claim 1 , wherein the cell culture medium comprises insulin at a concentration of from about 1.0 mg/L to about 100.0 mg/L.

4. The method of claim 1 , wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 100.0 mg/L.

5. The method of claim 1 , wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 50.0 mg/L.

6. The method of claim 1 , wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 35.0 mg/L.

7. The method of claim 1 , wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 25.0 mg/L.

8. The method of claim 1 , wherein the cell culture medium comprises insulin at a concentration of about 25 mg/L.

9. The method of claim 1 , wherein the cell culture medium comprises copper at a concentration of from about 69 nM to about 1,000 nM.

10. The method of claim 1 , wherein the cell culture medium comprises copper at a concentration of from about 325 nM to about 375 nM.

11. The method of claim 1 , wherein the cell culture medium comprises copper at a concentration of from about 325 nM to about 350 nM.

12. The method of claim 1 , wherein the cell culture medium comprises copper at a concentration of about any one of 330 nM, 335 nM, 339 nM, 340 nM, 345 nM or 350 nM.

13. The method of claim 1 , wherein the cell culture medium comprises copper at a concentration of about 339 nM.

14. The method of claim 1 , wherein the cell culture medium comprises cystine at a concentration of about 1.3 mM.

15. The method of claim 1 , wherein the cell culture medium comprises a porcine peptone at a concentration of from about 6.0 g/L to about 20.0 g/L.

16. The method of claim 1 , wherein the cell culture medium comprises a porcine peptone at a concentration of from about 8.0 g/L to about 12.0 g/L.

17. The method of claim 1 , wherein the cell culture medium comprises a porcine peptone at a concentration of from about 9.0 g/L to about 11.0 g/L.

18. The method of claim 1 , wherein the cell culture medium comprises a porcine peptone at a concentration of about 13 g/L.

19. The method of claim 1 , wherein the cell culture medium comprises a plant peptone at a concentration of from about 1.0 g/L to about 10.0 g/L.

20. The method of claim 1 , wherein the cell culture medium comprises a plant peptone at a concentration of from about 2.0 g/L to about 3.0 g/L.

21. The method of claim 1 , wherein the cell culture medium comprises a plant peptone at a concentration of from about 2.25 g/L to about 2.75 g/L.

22. The method of claim 1 , wherein the cell culture medium comprises a plant peptone at a concentration of about 3.1 g/L.

23. The method of claim 1 , wherein the cell culture medium comprises insulin and the method further comprises the step of adding an additional amount of insulin to the cell culture medium.

24. The method of claim 23 , wherein the additional amount of insulin is added to the cell culture medium once during the cell culture cycle.

25. The method of claim 23 , wherein the additional amount of insulin is added to the cell culture medium at least three times during the cell culture cycle.

26. The method of claim 23 , wherein the additional amount of insulin is added to the cell culture medium at least six times during the cell culture cycle.

27. The method of claim 23 , wherein the additional amount of insulin is added in an amount to provide insulin in the cell culture medium at a concentration of from about 1.0 mg/L to about 100.0 mg/L.

28. The method of claim 23 , wherein the additional amount of insulin is added in an amount to provide insulin in the cell culture medium at a concentration of from about 10.0 mg/L to about 100.0 mg/L.

29. The method of claim 23 , wherein the additional amount of insulin is added in an amount to provide insulin in the cell culture medium at a concentration of from about 10.0 mg/L to about 50.0 mg/L.

30. The method of claim 23 , wherein the additional amount of insulin is added in an amount to provide insulin in the cell culture medium at a concentration of from about 10.0 mg/L to about 35.0 mg/L.

31. The method of claim 23 , wherein the additional amount of insulin is added in an amount to provide insulin in the cell culture medium at a concentration of from about 10.0 mg/L to about 25.0 mg/L.

32. The method of claim 23 , wherein the additional amount of insulin is added in an amount to provide insulin in the cell culture medium at a concentration of about 15 mg/L.

33. The method of claim 1 , wherein the method further comprises the step of adding cysteine to the cell culture medium.

34. The method of claim 33 , wherein cysteine is added in an amount to provide from about 0.5 to about 2.0 mM cysteine in the cell culture medium.

35. The method of claim 33 , wherein cysteine is added in an amount to provide about 0.8 mM cysteine in the cell culture medium.

36. The method of claim 1 , wherein the cell is cultured at a temperature ranging from about 28° C. to about 37° C.

37. The method of claim 36 , wherein the cell is cultured at a temperature ranging from about 31° C. to about 35° C.

38. The method of claim 1 , wherein the cell is cultured at a first temperature of about 35° C. for a first period of time, is cultured at a second temperature of about 33° C. for a second period of time, and is cultured at a third temperature of about 31° C. for a third period of time.

39. The method of claim 1 , wherein bevacizumab, or a fragment thereof, is secreted into the cell culture medium.

40. The method of claim 1 , further comprising the step of recovering the bevacizumab, or a fragment thereof, from the cell culture.

41. A method of culturing a Chinese hamster ovary (CHO) cell comprising a nucleic acid encoding bevacizumab, or a fragment thereof, the method comprising the step of contacting the CHO cell with a cell culture medium comprising copper, insulin and cystine, wherein the cystine is at a concentration of from 1.25 mM to 2.5 mM.

42. The method of claim 41 , wherein the cell culture medium further comprises a a plant peptone, a porcine peptone, or both a plant peptone and a porcine peptone.

43. The method of claim 41 , wherein the cell culture medium comprises insulin at a concentration of from about 1.0 mg/L to about 100.0 mg/L.

44. The method of claim 41 , wherein the cell culture medium comprises insulin at a concentration of from about

Assignees

• Genentech Inc

Inventors

• Carvalhal Veronica
• Vijayasankaran Natarajan
• Brown Lauren
• Dirocco Thomas
• Mcknight Nathan

Classifications

• A61P35/00

  Antineoplastic agents · CPC title

• C07K2317/14

  Specific host cells or culture conditions, e.g. components, pH or temperature · CPC title

• C12N2500/20

  Transition metals · CPC title

• C12N2501/33

  Insulin · CPC title

• C07K2317/76

  Antagonist effect on antigen, e.g. neutralization or inhibition of binding · CPC title