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Last Updated: March 28, 2024

Claims for Patent: 8,093,045


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Summary for Patent: 8,093,045
Title:Fed-batch cell culture methods using non-animal-based hydrolysates
Abstract: The invention describes improved methods and compositions for producing a recombinant protein, e.g., an antibody, in mammalian cell culture. In addition, the invention provides improved cell culture media, including improved production media, feed solutions, and combination feeds, which may be used to improve protein productivity in mammalian cell culture.
Inventor(s): Pla; Itzcoatl A. (Worcester, MA), Matuck; Joseph G. (Worcester, MA), Fann; John C. (Shrewsbury, MA), Schulz; Christof (Ayer, MA), Roy; Nichole A. (Worcester, MA), Bruton; David F. (Enfield, CT), McIntire; James (Castro Valley, CA), Yu-Hsiang; David Chang (Solana Beach, CA), Seewoester; Thomas (Simi Valley, CA)
Assignee: Abbott Laboratories (Abbott Park, IL)
Application Number:11/901,274
Patent Litigation and PTAB cases: See patent lawsuits and PTAB cases for patent 8,093,045
Patent Claims:1. A fed batch method of producing a protein comprising a) culturing mammalian cells comprising a nucleic acid encoding the protein in a cell culture comprising a cell culture production medium; and b) feeding the mammalian cells by adding a hydrolysate enrichment solution and a basal enrichment solution to the cell culture during a time period, wherein the hydrolysate enrichment solution comprises a plant-based hydrolysate and 75-300 g/L of a yeast-based hydrolysate, such that the protein is produced.

2. The method of claim 1, wherein the basal enrichment solution comprises PF-CHO basal medium.

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

4. The method of claim 3, wherein the antibody, or fragment thereof is an anti-IL18 antibody, or a fragment thereof, or an anti-EPO receptor (EPO-R) antibody, or a fragment thereof.

5. A fed batch method of producing an anti-TNF.alpha. antibody, or a fragment thereof, comprising a) culturing Chinese Hamster Ovary (CHO) cells comprising a nucleic acid encoding the anti-TNF.alpha. antibody, or fragment thereof, in a cell culture comprising a cell culture production medium; and b) feeding the CHO cells by adding a hydrolysate enrichment solution and a basal enrichment solution to the cell culture during a time period, wherein the basal enrichment solution comprises a plant-based hydrolysate and 75-300 g/L of a yeast-based hydrolysate such that the anti-TNF.alpha. antibody, or fragment thereof, is produced.

6. The method of claim 5, wherein the cell culture production medium comprises at least 2.0 g/L of glucose.

7. The method of claim 5, further comprising recovering the anti-TNF.alpha. antibody, or fragment thereof.

8. The method of claim 5, wherein the plant-based hydrolysate is a soy-based hydrolysate.

9. The method of claim 5, wherein the cell culture production medium comprises a) a modified basal medium which excludes the following components sodium bicarbonate, buffer, mono-basic sodium phosphate, di-basic sodium phosphate, an osmolarity regulator, a surfactant, and monosaccharide glucose; b) about 8 to 10 ml/kg or 110 to 130 mg/L ferric citrate; c) about 4 to 8 mL/kg or 10 to 14 mg/kg recombinant human insulin; d) about 5 to 9 g/kg anhydrous glucose; e) about 0.1 to 1 g/kg L-glutamine; f) about 1 to 3 g/kg sodium bicarbonate; g) about 1 to 3 g/kg HEPES; h) about 2 to 3 g/kg NaCl; i) about 0.1 to 2 g/kg Pluronic F-68; j) about 0.01 to 0.1 g/kg NaH.sub.2PO.sub.4--H.sub.2O; k) about 0.1 to 0.1 g/kg Na.sub.2HPO.sub.4-7H.sub.2O; l) about 8 to 12 g/kg yeast-based hydrolysate; and m) about 6 to 8 g/kg plant-based hydrolysate.

10. The method of claim 5, wherein the cell culture is a large scale cell culture.

11. The method of claim 5, wherein the anti-TNF.alpha. antibody, or fragment thereof, is fully human.

12. A fed batch method of producing an anti-IL-12 antibody, or a fragment thereof, comprising a) culturing CHO cells comprising a nucleic acid encoding the antibody in a cell culture comprising a cell culture production medium, b) feeding the CHO cells by adding a hydrolysate enrichment solution and a basal enrichment solution to the cell culture during a time period, wherein the basal enrichment solution comprises a basal medium, asparagine, and glucose, and wherein the hydrolysate enrichment solution comprises a plant-based hydrolysate and 75-300 g/L of a yeast-based hydrolysate such that the anti-IL-12 antibody, or fragment thereof, is produced.

13. The method of claim 12, further comprising recovering the anti-IL12 antibody, or fragment of.

14. The method of claim 12, wherein the basal medium in the basal enrichment solution is PF CHO.

15. The method of claim 12, wherein the basal enrichment solution is added to the cell culture production medium every other day beginning on day 5 of the time period.

16. The method of claim 12, wherein the hydrolysate enrichment solution is added to the cell culture production medium every day beginning on either day 5 or day 6 of the time period.

17. The method of claim 12, wherein the cell culture production medium comprises a) a modified basal medium excluding the following components sodium bicarbonate, buffer, mono-basic sodium phosphate, di-basic sodium phosphate, an osmolarity regulator, a surfactant, and monosaccharide glucose; b) about 8 to 12 ml/kg or 110 to 130 mg/L ferric citrate; c) about 5 to 8 mL/kg or 11 to 15 mg/kg recombinant human insulin; d) about 5 to 9 g/kg anhydrous glucose; e) about 0.1 to 1 g/kg L-glutamine; f) about 1 to 2 g/kg sodium bicarbonate g) about 1 to 2 g/kg HEPES; h) about 2 to 3 g/kg NaCl; i) about 0.1 to 2 g/kg Pluronic F-68; j) about 0.01 to 0.1 g/kg NaH.sub.2PO.sub.4--H.sub.2O; k) about 0.1 to 1 g/kg Na.sub.2HPO.sub.4-7H.sub.2O; l) about 6 to 12 g/kg yeast-based hydrolysate; and m) about 6 to 8 g/kg plant-based hydrolysate.

18. The method of claim 12, wherein the cell culture is a large scale cell culture.

19. The method of claim 12, wherein the anti-IL-12 antibody, or fragment thereof is fully human.

20. The method of claim 1, wherein the basal enrichment solution comprises a concentrated basal medium.

21. The method of claim 1, wherein the basal enrichment solution comprises a basal medium, asparagine, and glucose.

22. The method of claim 1, wherein the mammalian cell is a Chinese Hamster Ovary (CHO) cell.

23. The method of claim 1, wherein the plant-based hydrolysate is a soy-based hydrolysate.

24. The method of claim 5, wherein the cell culture is cultured at a temperature ranging from about 32 to 38.degree. C.

25. The method of claim 24, wherein the culturing temperature is about 35.degree. C.

26. The method of claim 5, wherein the cell culture production medium is maintained between 20 and 65% dissolved oxygen.

27. The method of claim 26, wherein the cell culture production medium is maintained at about 30% dissolved oxygen.

28. The method of claim 5, wherein the osmolarity of the cell culture production medium is maintained throughout the culturing to no more than 500 mOsm.

29. The method of claim 5, wherein the basal medium is PF CHO.

30. The method of claim 5, wherein the basal enrichment solution has a pH of about 9.0 to 10.5.

31. The method of claim 5, wherein the time period is between about 9 to 15 days.

32. The method of claim 31, wherein the time period is about 12 days.

33. The method of claim 5, wherein the basal enrichment solution is added to the cell culture production medium on at least one of the following days of the time period: Day 4, Day 6, Day 9, and Day 11.

34. The method of claim 5, wherein the hydrolysate enrichment solution is added to the cell culture production medium on Day 4, Day 7, or Day 4 and Day 7 of the time period.

35. The method of claim 5, further comprising adjusting the pH of the cell culture production medium according to a pH linear ramp, wherein the pH linear ramp comprises starting from a pH of about 7.1 to 7.2 and resulting in a final pH of about 6.9.

36. The method of claim 35, wherein the pH linear ramp is adjusted over a period of at least about 24 hours.

37. The method of claim 35, wherein the pH linear ramp is adjusted over a period of at least about 48 hours.

38. The method of claim 35, wherein the pH linear ramp is adjusted over a period of about 72 hours.

39. The method of claim 10, wherein the large scale cell culture is greater than about 10 L.

40. The method of claim 10, wherein the large scale cell culture is 13 L.

41. The method of claim 11, wherein the fully human anti-TNF.alpha. antibody, or fragment thereof, is adalimumab, or a fragment thereof.

42. The method of claim 12, wherein the hydrolysate enrichment solution further comprises glucose.

43. The method of claim 12, wherein the cell culture is cultured at a temperature ranging from about 32 to 38.degree. C.

44. The method of claim 12, wherein the culturing temperature is about 33.degree. C.

45. The method of claim 12, wherein the cell culture production medium is maintained at between 20-65% dissolved oxygen.

46. The method of claim 12, wherein the cell culture production medium is maintained at about 40% dissolved oxygen.

47. The method of claim 12, wherein the cell culture production medium has a pH of about 6.7 to 7.2.

48. The method of claim 12, wherein the plant-based hydrolysate is a soy-based hydrolysate.

49. The method of claim 12, wherein the basal enrichment solution has a pH of about 9.7 and an osmolarity of about 1400 to 1500 mOsm.

50. The method of claim 5, wherein the time period is between 14-15 days.

51. The method of claim 5, wherein the basal enrichment solution and the hydrolysate enrichment solution are added to the cell culture production medium every day beginning on day 5 of the time period.

52. The method of claim 18, wherein the large scale cell culture is greater than 10 L.

53. The method of claim 18, wherein the large scale cell culture is about 13 L.

54. The method of claim 19, wherein the fully human anti-IL-12 antibody, or fragment thereof, is ABT-874, or a fragment thereof.

55. A fed batch method of producing adalimumab, or a fragment thereof, comprising a) culturing mammalian cells expressing adalimumab, or a fragment thereof, in a cell culture, wherein the mammalian cells are cultured at a constant temperature between 32-38.degree. C.; and b) feeding the mammalian cells by adding a hydrolysate enrichment solution and a basal enrichment solution to the cell culture, wherein the hydrolysate enrichment solution comprises a plant-based hydrolysate and 75-300 g/L of a hydrolysate which is not derived from a plant or an animal, such that adalimumab, or fragment thererof, is produced.

56. The method of claim 55, wherein the basal enrichment solution comprises a basal medium, asparagine, and glucose.

57. The method of claim 56, wherein the basal medium is PF CHO.

58. The method of any one of claims 55-57, wherein the mammalian cell is a Chinese Hamster Ovary (CHO) cell.

59. The method of claim 55, wherein the hydrolysate which is not derived from a plant or an animal is a yeast-based hydrolysate.

60. The method of claim 55, wherein the plant-based hydrolysate is a soy-based hydrolysate.

61. The method of anyone of claims 55-57, wherein the temperature is 35.degree. C.

62. The method of claim 3, wherein the antibody or fragment thereof, is an anti-TNF.alpha. antibody, or a fragment thereof.

63. The method of claim 3, wherein the antibody or fragment thereof, is an anti-IL-12 antibody, or a fragment thereof.

64. The method of claim 8, wherein the soy-based hydrolysate has a concentration of 50-280 g/L.

65. The method of claim 48, wherein the soy-based hydrolysate has a concentration of 50-280 g/L.

Details for Patent 8,093,045

Applicant Tradename Biologic Ingredient Dosage Form BLA Approval Date Patent No. Expiredate
Abbvie Inc. HUMIRA adalimumab Injection 125057 12/31/2002 ⤷  Try a Trial 2026-09-13
Abbvie Inc. HUMIRA adalimumab Injection 125057 02/21/2008 ⤷  Try a Trial 2026-09-13
Abbvie Inc. HUMIRA adalimumab Injection 125057 04/24/2013 ⤷  Try a Trial 2026-09-13
Abbvie Inc. HUMIRA adalimumab Injection 125057 09/23/2014 ⤷  Try a Trial 2026-09-13
Abbvie Inc. HUMIRA adalimumab Injection 125057 11/23/2015 ⤷  Try a Trial 2026-09-13
Abbvie Inc. HUMIRA adalimumab Injection 125057 03/09/2016 ⤷  Try a Trial 2026-09-13
>Applicant >Tradename >Biologic Ingredient >Dosage Form >BLA >Approval Date >Patent No. >Expiredate

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