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Last Updated: January 28, 2022

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Claims for Patent: 9,441,035

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Summary for Patent: 9,441,035
Title:Cell culture media and methods of antibody production
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.
Inventor(s): Carvalhal; Veronica (South San Francisco, CA), Vijayasankaran; Natarajan (South San Francisco, CA), Brown; Lauren (South San Francisco, CA), DiRocco; Thomas (South San Francisco, CA), McKnight; Nathan (South San Francisco, CA)
Assignee: GENENTECH, INC. (South San Francisco, CA)
Application Number:14/211,467
Patent Litigation and PTAB cases: See patent lawsuits and PTAB cases for patent 9,441,035
Patent Claims: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.degree. C. to about 37.degree. C.

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

38. The method of claim 1, wherein the cell is cultured at a first temperature of about 35.degree. C. for a first period of time, is cultured at a second temperature of about 33.degree. C. for a second period of time, and is cultured at a third temperature of about 31.degree. 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 10.0 mg/L to about 100.0 mg/L.

45. The method of claim 41, wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 50.0 mg/L.

46. The method of claim 41, wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 35.0 mg/L.

47. The method of claim 41, wherein the cell culture medium comprises insulin at a concentration of from about 10.0 mg/L to about 25.0 mg/L.

48. The method of claim 41, wherein the cell culture medium comprises insulin at a concentration of about 25 mg/L.

49. The method of claim 41, wherein the cell culture medium comprises copper at a concentration of from about 69 nM to about 1,000 nM.

50. The method of claim 41, wherein the cell culture medium comprises copper at a concentration of from about 325 nM to about 375 nM.

51. The method of claim 41, wherein the cell culture medium comprises copper at a concentration of from about 325 nM to about 350 nM.

52. The method of claim 41, 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.

53. The method of claim 41, wherein the cell culture medium comprises copper at a concentration of about 339 nM.

54. The method of claim 41, wherein the cell culture medium comprises cystine at a concentration of about 1.3 mM.

55. The method of claim 41, 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.

56. The method of claim 41, 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.

57. The method of claim 41, 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.

58. The method of claim 41, wherein the cell culture medium comprises a porcine peptone at a concentration of about 13 g/L.

59. The method of claim 41, 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.

60. The method of claim 41, 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.

61. The method of claim 41, 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.

62. The method of claim 41, wherein the cell culture medium comprises a plant peptone at a concentration of about 3.1 g/L.

63. The method of claim 41, 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.

64. The method of claim 63, wherein the additional amount of insulin is added to the cell culture medium once during the cell culture cycle.

65. The method of claim 63, wherein the additional amount of insulin is added to the cell culture medium at least three times during the cell culture cycle.

66. The method of claim 63, wherein the additional amount of insulin is added to the cell culture medium at least six times during the cell culture cycle.

67. The method of claim 63, 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.

68. The method of claim 63, 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.

69. The method of claim 63, 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.

70. The method of claim 63, 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.

71. The method of claim 63, 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.

72. The method of claim 63, 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.

73. The method of claim 41, wherein the method further comprises the step of adding cysteine to the cell culture medium.

74. The method of claim 73, wherein cysteine is added in an amount to provide from about 0.5 to about 2.0 mM cysteine in the cell culture medium.

75. The method of claim 73, wherein cysteine is added in an amount to provide about 0.8 mM cysteine in the cell culture medium.

76. The method of claim 41, wherein the cell is cultured at a temperature ranging from about 28.degree. C. to about 37.degree. C.

77. The method of claim 76, wherein the cell is cultured at a temperature ranging from about 28.degree. C. to about 35.degree. C.

78. The method of claim 41, wherein the cell is cultured at a first temperature of about 35.degree. C. for a first period of time, is cultured at a second temperature of about 33.degree. C. for a second period of time, and is cultured at a third temperature of about 31.degree. C. for a third period of time.

79. The method of claim 41, wherein bevacizumab, or a fragment thereof, is secreted into the cell culture medium.

80. The method of claim 41, wherein the CHO cell is contacted with the cell culture medium during the cell's growth phase.

81. The method of claim 41, wherein the CHO cell is contacted with the cell culture medium during the cell's production phase.

82. A method of enhancing the amount of bevacizumab, or a fragment thereof, produced from a Chinese hamster ovary (CHO) cell comprising a nucleic acid encoding bevacizumab, or a fragment thereof, the method comprising the step of culturing the CHO cell in a cell culture medium comprising insulin, copper and cystine, wherein the cystine is at a concentration of from 1.25 mM to 2.5 mM, and wherein the amount of bevacizumab, or a fragment thereof, produced from the CHO cell is enhanced relative to culturing the CHO cell in a cell culture medium without insulin, copper and cystine.

83. A method of culturing a Chinese hamster ovary (CHO) cell comprising a nucleic acid encoding bevacizumab, or a fragment thereof, in a cell culture medium comprising insulin, copper and cystine, wherein the cystine is at a concentration of from 1.25 mM to 2.5 mM, and wherein the amount of bevacizumab, or a fragment thereof, produced from the CHO cell is enhanced relative to culturing the CHO cell in a cell culture medium without at least two of insulin, copper and cystine.

84. The method of claim 1, wherein the amount of bevacizumab, or a fragment thereof, produced from the cell is enhanced relative to culturing the cell in a cell culture medium without insulin, copper and cystine.

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