Peptide-lipid conjugates, liposomes and lipsomal drug delivery

• US 6,339,069 B1
• Filed: 06/29/1999
• Issued: 01/15/2002
• Est. Priority Date: 10/15/1996
• Status: Expired due to Term

• Current Assignee: Transave Inc. (Insmed Incorporated)
• Original Assignee: ELAN Pharmaceutical Technologies
• Inventors: Ahl, Patrick L., Meers, Paul R., Ali, Shaukat, Pak, Charles, Cabral-Lilly, Donna, Erukulla, Ravi K., Franklin, J. Craig, Janoff, Andrew
• Primary Examiner(s): NGUYEN, DAVE TRONG
• Application Number:
  US09/343,650
• Time in Patent Office: 931 Days
• Field of Search: 514/44, 514/2, 424/450, 435/320.1, 435/455, 435/458
• US Class Current: 514/44.00R
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: A61K 47/543 Lipids, e.g. triglycerides; Polyamines, e.g. spermine or spermidine
• CPC Class Codes: A61K 47/544 Phospholipids
• CPC Class Codes: A61K 47/6911 the form being a liposome
• CPC Class Codes: A61K 9/127 Liposomes
• 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: A61K 9/1272 with substantial amounts of non-phosphatidyl, i.e. non-acylglycerophosphate, surfactants as bilayer-forming substances, e.g. cationic lipids with cholesterol as the only non-phosphatidyl surfactant A61K9/127; lipids as modifying agent A61K47/543
• CPC Class Codes: A61K 9/1275 Lipoproteins; Chylomicrons; Artificial HDL, LDL, VLDL, protein-free species thereof; Precursors thereof
• CPC Class Codes: A61P 11/08 Bronchodilators
• CPC Class Codes: A61P 23/02 Local anaesthetics
• CPC Class Codes: A61P 27/08 Mydriatics or cycloplegics
• 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 31/04 Antibacterial agents
• CPC Class Codes: A61P 31/10 Antimycotics
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: C07K 1/1077 by covalent attachment of residues other than amino acids or peptide residues, e.g. sugars, polyols, fatty acids
• CPC Class Codes: C07K 5/06026 the side chain containing 0 or 1 carbon atom, i.e. Gly or Ala
• CPC Class Codes: C07K 5/06191 containing heteroatoms different from O, S, or N
• CPC Class Codes: C07K 5/0806 : the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
• CPC Class Codes: C07K 5/0808 : the side chain containing 2 to 4 carbon atoms, e.g. Val, Ile, Leu
• CPC Class Codes: C07K 5/0819 : with the first amino acid being acidic
• CPC Class Codes: C07K 5/0823 : and Pro-amino acid; Derivatives thereof
• CPC Class Codes: C07K 5/0827 : containing heteroatoms different from O, S, or N
• CPC Class Codes: C07K 5/1008 : the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
• CPC Class Codes: C07K 5/1013 : the side chain containing O or S as heteroatoms, e.g. Cys, Ser
• CPC Class Codes: C07K 5/1027 : containing heteroatoms different from O, S, or N
• CPC Class Codes: C07K 7/06 : having 5 to 11 amino acids
• CPC Class Codes: C07K 9/001 : the peptide sequence having less than 12 amino acids and not being part of a ring structure

Abstract

Peptide-lipid conjugates are incorporated into liposomes so as to selectively destabilize the liposomes in the vicinity of target peptidase-secreting cells, and hence, to deliver the liposomes to the vicinity of the target cells, or directly into the cells. The liposomes can thus be used to treat mammals for diseases, disorders or conditions, e.g., tumors, microbial infection and inflammations, characterized by the occurrence of peptidase-secreting cells.

Claims

1. A method of administering a bioactive agent to a mammal, which comprises administering to the mammal a composition comprising:

(i) a pharmaceutically acceptable carrier; and

,

(ii) a liposome comprising a bioactive agent and a lipid component, the lipid component comprising a peptide-lipid conjugate having the formula;

wherein;

X is a linker selected from the group consisting of a single bond or the group R³;

each of R¹ and R² is —

OC(O)(CH₂)_n1(CH═

CH)_n2(CH₂)_n3(CH═

CH)_n4(CH₂)_n5(CH═

CH)_n6(CH₂)_n7(CH═

CH)_n8(CH₂)_n9CH₃; and

R³ is —

C(O)(CH₂)_n1(CH═

CH)_n2(CH₂)_n3(CH═

CH)_n4(CH₂)_n5(CH═

CH)_n6(CH₂)_n7(CH═

CH)_n8(CH₂)_n9HN—

,

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

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

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

n7 is zero or is an integer equal to from 1 to 13, and

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

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

for R³ the sum of n1+2n2+n3+2n4+n5+2n6+n7+2n8+n9 is zero or is an integer equal to from 1 to 22;

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

Y is a peptide comprising an amino acid sequence which is the substrate of a cell-secreted or cell-associated peptidase; and

the bioactive agent is delivered to the vicinity of cells in the mammal which secrete a peptidase which recognizes the amino acid substrate.

2. The method of claim 1, wherein the liposome is a large unilamellar liposome.

3. The method of claim 1, wherein the peptide comprises the amino acid sequence Ala-Ala.

4. The method of claim 3, wherein the peptide comprises an amino acid sequence selected from the group consisting of the sequences Ala-Ala, Ala-Ala-Pro-Val (SEQ ID NO:

1), Ala-Ala-Met, Ala-Ala-Pro-Phe (SEQ ID NO;

3), Ala-Ala-Pro-Met (SEQ ID NO;

4), Ala-Ala-Arg, Ser-Ala-Ala-Arg (SEQ ID NO;

5), Ser-Ser-Ala-Ala-Arg (SEQ ID NO;

6), Ser-S carboxyl sugar-Ala-Ala-Arg- (SEQ ID NO;

7), Ala-Ala-Asp-, Ser-Ala-Ala-Asp- (SEQ ID NO;

8) and Ser-Ser-Ala-Ala-Asp-(SEQ ID NO;

9).

5. The method of claim 4, wherein the peptide comprises the amino acid sequence Ala-Ala-Pro-Val (SEQ ID NO:

1).

6. The method of claim 1, wherein the peptide comprises an amino acid sequence selected from the group consisting of Arg-Pro-Lys-Pro-Leu-Ala-Nva- (SEQ ID NO:

10), Ser-Arg-Pro-Lys-Pro-Leu-Ala-Nva- (SEQ ID NO;

11), Ser-Ser-Arg-Pro-Lys-Pro-Leu-Ala-Nva- (SEQ ID NO;

12), Pro-Cha-Gly-Nva-His-Ala-Dpa-NH₂ (SEQ ID NO;

13), Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH₂, (SEQ ID NO;

14);

Pro-Cha-Gly-Nva-;

Pro-Leu-Gly-Leu- (SEQ ID NO;

15) and Gly-Pro-Gln-Gly-Ile- (SEQ ID NO;

16).

7. The method of claim 1, wherein the peptide is modified at its amino terminus by a moiety selected from the group consisting of acetyl, methoxy, carboxy sugar, polyethylene glycol and methoxy-substituted carboxy sugar modifications.

8. The method of claim 7, wherein the peptide is preferably N-acetyl modified at its amino terminus and the peptide-lipid conjugate comprises from about 20 mole % to about 80 mole % of the lipid component.

9. The method of claim 1, wherein X is a single bond, each of R¹ and R² is —

OC(O)(CH₂)₇(CH═

CH)(CH₂)₇CH₃ and the peptide comprises the amino acid sequence Ala-Ala.

10. The method of claim 9, wherein the peptide-lipid conjugate is:

the peptide is N-methoxysuccinyl modified at its amino terminus and the peptide-lipid conjugate comprises from about 20 mole % to about 80 mole % of the lipid component.

11. The method of claim 10, wherein the lipid component further comprises an additional lipid which is a positively charged lipid selected from the group consisting of 1,2-bis(oleoyloxy)-3(trimethylammonio)propane (DOTAP);

1-N,N-dimethylamino dioleoyl propane (DODAP);

1-oleoyl-2-hydroxy-3-N,N-dimethylamino propane;

1,2-diacyl-3-N,N-dimethylamino propane;

1,2-didecanoyl-1-N,N,-dimethylamino propane, 3-beta-[N-[(N′

,N′

-dimethylamino)ethane]carbamoyl]cholesterol (DC-Chol), 1,2-dimyristooxypropyl-3-dimethylhydroxyethyl ammonium bromide (DMRIE); and

1,2-dioloeooxypropyl-3-dimethylhydroxyethyl ammonium bromide (DORI).

12. The method of claim 11, wherein the positively charged lipid is DODAP.

13. The method of claim 12, wherein the lipid component comprises 50 mole % DODAP and 50 mole % of the peptide-lipid conjugate.

14. The method of claim 12, wherein the lipid component comprises DODAP and N-Ala-Ala-Pro-Val-DOPE.

15. The method of claim 11, wherein the positively charged lipid is DOTAP.

16. The method of claim 15, wherein the lipid component comprises 50 mole % DOTAP and 50 mole % of the peptide-lipid conjugate.

17. The method of claim 15, wherein the lipid component comprises 50 mole % DOTAP and 50 mole % N-Ala-Ala-Pro-Val.

18. The method of claim 10, wherein the lipid component further comprises an additional lipid selected from the group consisting of trans-esterified phosphatidylethanolamine (tPE), dipalmitoyl phosphatidylethanolamine (DPPE), palmitoyloleoyl phosphatidylethanolamine (POPE) and dioleoyl phosphatidylethanolamine (DOPE).

19. The method of claim 1, wherein the peptidase is selected from the group consisting of elastase, plasmin, plasminogen activator, urokinase;

stromelysin, human collagenases, cathepsins, lysozyme, granzymes, dipeptidyl peptidases, peptide hormone-inactivating enzymes, kininases, bacterial peptidases and viral proteases.

20. The method of claim 19, wherein the peptidase is elastase.

21. The method of claim 19, wherein the peptidase is stromelysin, a cathepsin, plasmin or a plasminogen activator.

22. The method of claim 1, wherein the bioactive agent is 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, plasmid deoxyribonucleic acid sequences and bioactive lipids.