High Purity Lipopeptides

• US 20110207658A1
• Filed: 09/22/2010
• Published: 08/25/2011
• Est. Priority Date: 01/20/2000
• Status: Active Grant

• Current Assignee: Cubist Pharmaceuticals, LLC (Merck & Co., Inc.)
• Original Assignee: Auro Tagliani, Jan-Ji Lai, Joseph P. Decourcey, Maurizio Zenoni, Paul Lynch, Thomas Kelleher
• Inventors: Zenoni, Maurizio, Lynch, Paul D., Lai, Jan-Ji, Tagliani, Auro R., Kelleher, Thomas J., DeCourcey, Joseph P.
• Granted Patent: US 8,129,342 B2
• Time in Patent Office: Days
• US Class Current: 514/2.9
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 38/10 Peptides having 12 to 20 amino acids A61K38/043 - A61K38/046 take precedence
• CPC Class Codes: A61K 38/12 Cyclic peptides , e.g. bacitracins; Polymyxins; Gramicidins S, C; Tyrocidins A, B or C A61K38/043 - A61K38/046 take precedence
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 31/10 Antimycotics
• CPC Class Codes: C07K 1/145 by extraction or solubilisation
• CPC Class Codes: C07K 5/0821 with the first amino acid being heterocyclic, e.g. His, Pro, Trp
• CPC Class Codes: C07K 7/08 having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68
• CPC Class Codes: Y10S 435/886 Streptomyces

Abstract

The invention discloses highly purified daptomycin and to pharmaceutical compositions comprising this compound. The invention discloses a method of purifying daptomycin comprising the sequential steps of anion exchange chromatography, hydrophobic interaction chromatography and anion exchange chromatography. The invention also discloses a method of purifying daptomycin by modified buffer enhanced anion exchange chromatography. The invention also discloses an improved method for producing daptomycin by fermentation of Streptomyces roseosporus. The invention also discloses high pressure liquid chromatography methods for analysis of daptomycin purity. The invention also discloses lipopeptide micelles and methods of making the micelles. The invention also discloses methods of using lipopeptide micelles for purifying lipopeptide antibiotics, such as daptomycin. The invention also discloses using lipopeptide micelles therapeutically.

Claims

1. A composition comprising

(a) essentially pure daptomycin,

(b) daptomycin that is substantially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin,

(c) daptomycin that is essentially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin,

(d) daptomycin that is free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin,

(e) daptomycin that is substantially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12,

(f) daptomycin that is essentially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12, or

(g) substantially pure daptomycin.

2. The composition of claim 1 comprising essentially pure daptomycin.

3. The composition of claim 1 compromising daptomycin that is substantially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

4. The composition according to claim 3 that is essentially free of anhydro-daptomycin.

5. The composition according to claim 3 that is free of anhydro-daptomycin.

6. The composition of claim 1 that is substantially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

7. The composition according to claim 6 that is essentially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

8. The composition of claim 1, wherein daptomycin purity is measured by HPLC.

9. The composition of claim 1 further comprising a pharmaceutically acceptable carrier or excipient.

10. A pharmaceutical composition according to claim 9, further comprising one or more antibiotics, one or more antifungal agents, or both an antibiotic and an antifungal agent.

11. The composition according to claim 1 wherein the daptomycin is purified by a process comprising the steps of:

a) supplying a fermentation broth;

b) fermenting Streptomyces roseosporus with a feed of n-decanoic acid to produce daptomycin in the fermentation broth;

c) clarifying the fermentation broth to obtain a clarified solution;

d) subjecting the clarified solution to anion exchange chromatography to obtain an enriched daptomycin preparation;

e) subjecting the enriched daptomycin preparation to hydrophobic interaction chromatography to obtain a semi-purified daptomycin preparation; and

f) subjecting the semi-purified daptomycin preparation to anion exchange chromatography to obtain the composition of claim 1.

12. The composition according to claim 11, wherein the feed of n-decanoic acid is regulated to achieve a residual concentration of n-decanoic acid of no more than 50 parts per million (ppm) during fermentation.

13. The composition according to claim 11, wherein said clarifying comprises filtration or centrifugation and depth filtration.

14. The composition according to claim 11, wherein the anion exchange chromatography in d) is performed using a resin comprising a copolymer of 2-methacrylic acid and ethyleneglycol dimethacrylate (EDGM).

15. The composition according to claim 11, wherein the hydrophobic interaction chromatography is performed using a resin comprising a co-polymer of cross-linked divinylbenzene/stryene.

16. The composition according to claim 15, wherein the hydrophobic interaction chromatography is performed at neutral pH and a solvent concentration that is reduced compared to the solvent concentration used when performing the hydrophobic interaction chromatography at acidic pH.

17. The composition according to claim 16, wherein the resin is recycled by loading the column at an acidic pH and eluting the column at a neutral pH.

18. The composition according to claim 11, wherein the anion exchange chromatography in 0 is performed using a resin comprising a copolymer of 2-methacrylic acid and ethyleneglycol dimethacrylate (EDGM).

19. The composition according to claim 11, wherein the anion exchange chromatography is used to reduce the level of solvent in the clarified solution.

20. The composition according to claim 11, wherein the anion exchange chromatography is performed via continuous flow chromatography.

21. The composition according to claim 11, wherein the process further comprises the step of filtering daptomycin.

22. The composition according to claim 11, wherein the process further comprises the step of depyrogenating daptomycin using ultrafiltration.

23. The composition according to claim 22 wherein said depyrogenating comprises the steps of:

i) providing a daptomycin solution under conditions in which the daptomycin is in a monomeric and nonmicellar state;

ii) filtering the daptomycin solution under conditions in which the daptomycin passes through the filter but pyrogens do not pass through the filter;

iii) subjecting the daptomycin solution to conditions forming a daptomycin aggregate;

iv) filtering the daptomycin aggregate under conditions in which the daptomycin aggregate is retained on the filter; and

v) collecting the daptomycin aggregate.

24. The composition according to claim 22, wherein the process further comprises the step of lyophilizing daptomycin.

25. The composition according to claim 22, wherein the anion exchange chromatography is performed via radial flow chromatography.

26. The composition according claim 11, wherein said clarifying comprises microfiltration or centrifugation.

27. The composition according to claim 11, wherein the process further comprises the steps of filtering and concentrating daptomycin.

28. The composition according to claim 11, wherein the process further comprises the step of separating the enriched daptomycin from low molecular weight material by ultrafiltration.

29. The composition according to claim 28, wherein the process further comprises the step of depyrogenating the daptomycin.

30. The composition of claim 1 comprising substantially pure daptomycin.

31. The pharmaceutical composition of claim 9 comprising essentially pure daptomycin.

32. The pharmaceutical composition of claim 9 comprising daptomycin that is substantially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

33. The pharmaceutical composition of claim 9 comprising daptomycin that is essentially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

34. The pharmaceutical composition of claim 9 comprising daptomycin that is free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

35. The pharmaceutical composition of claim 9 comprising daptomycin that is substantially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

36. The pharmaceutical composition of claim 9 comprising daptomycin that is essentially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

37. The pharmaceutical composition of claim 9 comprising substantially pure daptomycin.

38. A method for preparing a pharmaceutical composition comprising combining the composition of claim 1 with a pharmaceutically acceptable carrier or excipient.

39. The method of claim 38 wherein the composition is essentially pure daptomycin.

40. The method of claim 38 wherein the composition is daptomycin that is substantially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

41. The method of claim 38 wherein the composition is daptomycin that is essentially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

42. The method of claim 38 wherein the composition is daptomycin that is free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

43. The method of claim 38 wherein the composition is daptomycin that is substantially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

44. The method of claim 38 wherein the composition is daptomycin that is essentially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

45. The method of claim 38 wherein the composition is substantially pure daptomycin.

46. A pharmaceutical composition prepared by the method of claim 38.

47. The pharmaceutical composition of claim 46 wherein the composition is essentially pure daptomycin.

48. The pharmaceutical composition of claim 46 wherein the composition is daptomycin that is substantially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

49. The pharmaceutical composition of claim 46 wherein the composition is daptomycin that is essentially free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

50. The pharmaceutical composition of claim 46 wherein the composition is daptomycin that is free of anhydro-daptomycin and substantially free of β

-isomer of daptomycin.

51. The pharmaceutical composition of claim 46 wherein the composition is daptomycin that is substantially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

52. The pharmaceutical composition of claim 46 wherein the composition is daptomycin that is essentially free of each of impurities 1 to 14 defined by peaks 1-14 shown in FIG. 12.

53. The pharmaceutical composition of claim 46 wherein the composition is substantially pure daptomycin.