N-acyl phosphatidylethanolamine-mediated liposomal drug delivery

• US 6,294,191 B1
• Filed: 01/19/2000
• Issued: 09/25/2001
• Est. Priority Date: 10/15/1996
• Status: Expired due to Fees

• Current Assignee: Transave Inc. (Insmed Incorporated)
• Original Assignee: The Liposome Co., Inc. (Perrigo Company PLC)
• Inventors: Meers, Paul R., Pak, Charles, Shangguan, Tong, Janoff, Andrew, Ali, Shaukat
• Primary Examiner(s): Kishore, Gollamudi S.
• Application Number:
  US09/487,053
• Time in Patent Office: 615 Days
• Field of Search: 424/450, 424/1.21, 424/9.321, 424/9.51, 424/417, 424/94.3, 424/812, 428/402.2, 436/829, 935/54
• US Class Current: 424/450
• CPC Class Codes: A61K 9/1271 Non-conventional liposomes, e.g. PEGylated liposomes, liposomes coated with polymers liposome as conjugate A61K47/6911
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: Y10S 424/812 Liposome comprising an antibody, antibody fragment, antigen, or other specific or nonspecific immunoeffector
• CPC Class Codes: Y10S 436/829 Liposomes, e.g. encapsulation
• CPC Class Codes: Y10T 428/2984 Microcapsule with fluid core [includes liposome]

Abstract

This invention provides liposomes containing one or more N-acylated phosphatidylethanolamines, such liposomes being useful for localizing the delivery of bioactive agents to cells.

Claims

1. A pharmaceutical composition comprising:

(a) a pharmaceutically acceptable carrier; and

(b) a liposome having a lipid component which comprises;

(i) an N-acyl phosphatidylethanolamine (NAPE) having the formula;

(ii) an additional lipid,

 

wherein;

each of R¹, R² and R³ is independently a group having the formula —

C(O)(CH₂)_n1(CH═

CH)_n2(CH₂)_n3(CH═

CH)_n4(CH₂)_n5(CH═

CH)_n6(CH₂)_n7(CH═

CH)_n8(CH₂)_n9CH₃;

n1 is zero or an integer equal to from 1 to 22;

n3 is zero or an integer equal to from 1 to 19;

n5 is zero or an integer equal to from 1 to 16;

n7 is zero or an integer equal to from 1 to 13;

n9 is zero or an integer equal to from 1 to 10;

for each of R¹ and R² independently the sum of n1+2n2+n3+2n4+n5+2n6+n7+2n8+n9 is an integer equal to from 12 to 22, at least one of n2, n4, n6 and n8 is equal to 1;

for R³ the sum of n1+2n2+n3+2n4+n5+2n6+n7+2n8+n9 is an integer equal to from 2 to 22;

each of n2, n4, n6 and n8 is independently equal to 0 or 1;

the lipid component comprises a destabilization-effective amount of the N-acyl phosphatidylethanolamine of at least about 25 mole %;

wherein the liposome is multilamellar or is a large unilamellar liposome wherein the destabilization-effective amount of the NAPE is that amount of NAPE which renders the liposome fusogenic when the liposome is admixed with a target, in the presence of divalent cations or at a pH of about 4.9 to about 6.0.

2. The pharmaceutical composition of claim 1, wherein for one or more of R¹ and R² at least one of n2, n4, n6 or n8 is equal to 1.

3. The pharmaceutical composition of claim 2, wherein for both of R¹ and R² at least one of n2, n4, n6 or n8 is equal to 1.

4. The pharmaceutical composition of claim 3, wherein each of R¹ and R² is —

C(O)(CH₂)₇(CH═

CH)(CH₂)₇CH₃.

5. The pharmaceutical composition of claim 1, wherein R³ is —

C(O)(CH₂)₄CH₃, —

C(O)(CH₂)₁₀CH₃ or —

C(O)(CH₂)₁₄CH₃.

6. The pharmaceutical composition of claim 5, wherein R³ is —

C(O)(CH₂)₁₀CH₃.

7. The pharmaceutical composition of claim 1, wherein the N-acyl phosphatidylethanolamine is

8. The pharmaceutical composition of claim 7, wherein the N-acyl phosphatidylethanolamine comprises from about 25 mole % to about 90 mole % of the lipid component.

9. The pharmaceutical composition of claim 1, wherein the N-acyl phosphatidylethanolamine comprises from about 25 mole % to about 80 mole % of the lipid component.

10. The pharmaceutical composition of claim 9, wherein the N-acyl phosphatidylethanolamine comprises about 70 mole % of the lipid component.

11. The pharmaceutical composition of claim 1, wherein the additional lipid is a phospholipid.

12. The pharmaceutical composition of claim 11, wherein the phospholipid is a phosphatidylcholine.

13. The pharmaceutical composition of claim 12, wherein the phosphatidylcholine is dioleoyl phosphatidylcholine.

14. The pharmaceutical composition of claim 1 wherein the lipid component comprises N-dodecanoyl phosphatidylethanolamine and dioleoyl phosphatidylcholine.

15. The pharmaceutical composition of claim 14, wherein N-dodecanoyl phosphatidylethanolamine comprises about 70 mole % of the liposome'"'"'s lipid component and dioleoyl phosphatidylcholine comprises about 30 mole % of the lipid component.

16. The pharmaceutical composition of claim 1, wherein the additional lipid is a phosphatidylethanolamine.

17. The pharmaceutical composition of claim 16, wherein the phosphatidylethanolamine is selected from the group consisting of trans-esterified phosphatidylethanolamine, dipalmitoyl phosphatidylethanolamine, palmitoyl oleoyl phosphatidylethanolamine and dioleoyl phosphatidylethanolamine.

18. The pharmaceutical composition of claim 17, wherein the phosphatidylethanolamine is a phosphatidylethanolamine conjugated to a moiety selected from the group consisting of dicarboxylic acids, polyethylene glycols, polyalkyl ethers and gangliosides.

19. The pharmaceutical composition of claim 1, further comprising a targeting moiety selected from the group consisting of antibodies, cell receptor ligands and lectins.

20. The pharmaceutical composition of claim 1, further comprising a bioactive agent selected from the group consisting of antiviral agents, antibacterial agents, antifungal agents, antineoplastic agents, antiinflammatory agents, radiolabels, radiopaque compounds, fluorescent compounds, mydriatic compounds, bronchodilators, local anesthetics, nucleic acid sequences and bioactive lipids.

23. The method of claim 1, wherein the cell is a mammalian cell, the cell is contacted with the composition in the body of the mammal and the composition comprises a pharmaceutically acceptable carrier.

24. The method of claim 23, wherein the liposome additionally comprises a targeting moiety.

25. The method of claim 23, wherein the mammal is afflicted with a cancer and wherein the liposome comprises an anticancer effective amount of an anticancer agent.

51. The pharmaceutical composition of claim 1, wherein the divalent cations are employed at physiological concentrations.

52. The pharmaceutical composition of claim 1, wherein the target is a nucleated cell, and the liposome is admixed with the nucleated cell in the absence of added divalent cations.

21. A method of delivering a bioactive agent to a target which comprises contacting the target with a composition, wherein the composition comprises:

(a) a liposome having a lipid component which comprises;

(i) an N-acyl phosphatidylethanolamine (NAPE) having the formula;

(ii) an additional lipid,

 

wherein;

each of R¹, R² and R³ is independently a group having the formula —

C(O)(CH₂)_n1(CH═

CH)_n2(CH₂)_n3(CH═

CH)_n4(CH₂)_n5(CH═

CH)_n6(CH₂)_n7(CH═

CH)_n8(CH₂)_n9CH₃;

n1 is zero or an integer equal to from 1 to 22;

n3 is zero or an integer equal to from 1 to 19;

n5 is zero or an integer equal to from 1 to 16;

n7 is zero or an integer equal to from 1 to 13;

n9 is zero or an integer equal to from 1 to 10;

for each of R¹ and R² independently the sum of n1+2n2+n3+2n4+n5+2n6+n7+2n8+n9 is an integer equal to from 12 to 22, at least one of n2, n4, n6 and n8 is equal to 1;

for R³ the sum of n1+2n2+n3+2n4+n5+2n6+n7+2n8+n9 is an integer equal to from 2 to 22;

each of n2, n4, n6 and n8 is independently equal to 0 or 1;

the lipid component comprises a destabilization-effective amount of the N-acyl phosphatidylethanolamine of at least about 25 mole %; and

wherein the liposome is multilamellar or is a large unilamellar liposome.

22. The method of claim 21, wherein the contacting is in vitro and wherein the contacting is in the presence of a physiological concentration of a cation or is at an acidic pH.

26. The method of claim 21, wherein the target is a cell.

27. The method of claim 22, wherein the acidic pH is between pH about 4.9 to about 6.0.

28. The method of claim 26, wherein the cell is a nucleated cell, and the liposome is admixed with the nucleated cell at an acidic pH.

29. The method of claim 26, wherein the cell is a nucleated cell, and the liposome is admixed with the nucleated cell in the absence of added divalent cations.

30. The method of claim 28, wherein the acidic pH is between pH about 4.9 to about 6.0.

31. The method of claim 26, wherein the targeting moiety is selected from the group consisting of antibodies, cell receptor ligands and lectins.

32. The method of claim 21, wherein for one or more of R¹ and R² of the liposome at least one of n2, n4, n6 or n8 is equal to 1.

33. The method of claim 32, wherein for both of R¹ and R² at least one of n2, n4, n6 or n8 is equal to 1.

34. The method of claim 33, wherein each of R¹ and R² is —

C(O)(CH₂)₇(CH═

CH)(CH₂)₇CH₃.

35. The method of claim 21, wherein R³ of the liposome is —

C(O)(CH₂)₄CH₃, —

C(O)(CH₂)₁₀CH₃ or —

C(O)(CH₂)₁₄CH₃.

36. The method of claim 35, wherein R³ is —

C(O)(CH₂)₁₀CH₃.

37. The method of claim 21, wherein the N-acyl phosphatidylethanolamine is

38. The method of claim 21, wherein the N-acyl phosphatidylethanolamine comprises from about 10 mole % to about 90 mole % of the lipid component.

39. The method of claim 38, wherein the N-acyl phosphatidylethanolamine comprises from about 20 mole % to about 80 mole % of the lipid component.

40. The method of claim 39, wherein the N-acyl phosphatidylethanolamine comprises about 70 mole % of the lipid component.

41. The method of claim 21, wherein the additional lipid is a phospholipid.

42. The method of claim 41, wherein the phospholipid is a phosphatidylcholine.

43. The method of claim 42, wherein the phosphatidylcholine is dioleoyl phosphatidylcholine.

44. The method of claim 21, wherein the liposome has a lipid component comprising N-dodecanoyl phosphatidylethanolamine and dioleoyl phosphatidylcholine.

45. The method of claim 43, wherein N-dodecanoyl phosphatidylethanolamine comprises about 70 mole % of the liposome'"'"'s lipid component and dioleoyl phosphatidylcholine comprises about 30 mole % of the lipid component.

46. The method of claim 21, wherein the additional lipid is a phosphatidylethanolamine.

47. The method of claim 45, wherein the phosphatidylethanolamine is selected from the group consisting of trans-esterified phosphatidylethanolamine, dipalmitoyl phosphatidylethanolamine, palmitoyl oleoyl phosphatidylethanolamine and dioleoyl phosphatidylethanolamine.

48. The method of claim 45, wherein the phosphatidylethanolamine is a phosphatidylethanolamine conjugated to a moiety selected from the group consisting of dicarboxylic acids, polyethylene glycols, polyalkyl ethers and gangliosides.

49. The method of claim 21 wherein the liposome comprises a targeting moiety selected from the group consisting of antibodies, cell receptor ligands and lectins.

50. The method of claim 21, wherein the liposome comprises a bioactive agent selected from the group consisting of antiviral agents, antibacterial agents, antifungal agents, antineoplastic agents, antiinflammatory agents, radiolabels, radiopaque compounds, fluorescent compounds, mydriatic compounds, bronchodilators, local anesthetics, nucleic acid sequences and bioactive lipids.