Use of the cathelicidin LL-37 and derivatives thereof for wound healing

• US 7,452,864 B2
• Filed: 01/28/2004
• Issued: 11/18/2008
• Est. Priority Date: 01/29/2003
• Status: Active Grant

• Current Assignee: Promore Pharma AB
• Original Assignee: Lipopeptide AB
• Inventors: Carlsson, Anders, Bogentoft, Conny, Heilborn, Johan, Stahle-Backdahl, Mona
• Primary Examiner(s): Russel; Jeffrey E
• Application Number:
  US10/543,659
• Publication Number:
  US 20070149448A1
• Time in Patent Office: 1,756 Days
• Field of Search: None
• US Class Current: 514/1.1
• CPC Class Codes: A61K 38/16 Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof enzyme inhibitors A61K38/005
• CPC Class Codes: A61K 38/1709 from mammals
• 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: A61P 1/04 for ulcers, gastritis or reflux esophagitis, e.g. antacids, inhibitors of acid secretion, mucosal protectants
• CPC Class Codes: A61P 17/00 Drugs for dermatological disorders
• CPC Class Codes: A61P 17/02 for treating wounds, ulcers, burns, scars, keloids, or the like
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 14/47 from mammals
• CPC Class Codes: C07K 14/4723 Cationic antimicrobial peptides, e.g. defensins
• CPC Class Codes: C12N 2500/32 Amino acids
• CPC Class Codes: C12N 2500/36 Lipids
• CPC Class Codes: C12N 5/0018 Culture media for cell or tissue culture media for specific animal cell type C12N5/06
• CPC Class Codes: C12N 5/0629 Keratinocytes; Whole skin

Abstract

Use of the antimicrobial cathelicidin peptide ll-37, N-terminal fragments of LL-37 or extended sequences of LL-37 having 1-3 amino acids in the C-terminal end, for stimulating proliferation of epithalial and stromal cells and thereby healing of wounds, such as chronic ulcers. The cytotoxic effect of LL-37 may be reduced by including a bilayer-forming polar lipid, especially a digalactosyldiacylglycerol, in pharmaceutical compositions and growth media comprising LL-37.

Claims

1. A method of regenerating epithelial cells, healing of wound epithelium or healing of wound stroma, which comprises administering to a subject a peptide comprising an amino acid sequence containing at least 25 amino acids of the N-terminal fragment of SEQ ID NO:

1 or a pharmaceutically acceptable salt or ester or amide thereof, wherein said peptide enhances proliferation of epithelial and/or stromal cells through a non-lytic mechanism.

2. The method according to claim 1, wherein the peptide is LL-37, SEQ ID NO:

1.

3. The method according to claim 1, wherein the peptide is selected from the group consisting of LL-36, LL-35, LL-34, LL-33, LL-32, LL-31, LL-30, LL-29, LL-28, LL-27, LL-26, and LL-25, having the sequence SEQ ID NO:

2, SEQ ID NO;

3, SEQ ID NO;

4, SEQ ID NO;

5, SEQ ID NO;

6, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, SEQ ID NO;

12, SEQ ID NO;

13, respectively.

4. The method according to claim 1, wherein the peptide is in the form of an acetate salt.

5. The method according to claim 1, wherein the peptide is LL-37 (SEQ ID NO:

1) in the form of an acetate salt.

17. The method according to claim 1, wherein the subject has a chronic ulcer due to venous insufficiency, metabolic dysfunction, or immunological dysregulation.

18. The method according to claim 1, wherein the subject has a wound due to trauma or a burn.

6. A pharmaceutical composition for topical application to epithelial cells comprising:

a) a peptide comprising an amino acid sequence containing at least 25 amino acids of the N-terminal fragment of LL-37 (SEQ ID NO;

     1) or a pharmaceutically acceptable salt or ester or amide thereof; and

b) a bilayer forming polar lipid carrier comprising a galactolipid.

7. The pharmaceutical composition according to claim 6, wherein the peptide is LL-37 (SEQ ID NO:

1).

8. The pharmaceutical composition according to claim 6, wherein the peptide is in the form of an acetate salt.

9. The pharmaceutical composition according to claim 6, wherein the the peptide is selected from the group consisting of:

LL-36, LL-35, LL-34, LL-33, LL-32, LL-31, LL-30, LL-29, LL-28, LL-27, LL-26, and LL-25, having the amino acid sequence SEQ ID NO;

2, SEQ LD NO;

3, SEQ ID NO;

4, SEQ II) NO;

5, SEQ ID NO;

6, SEQ ID NO;

7, SEQ ID NO;

8, SEQ ID NO;

9, SEQ ID NO;

10, SEQ ID NO;

11, SEQ ID NO;

12, and SEQ ID NO;

13, respectively.

10. The pharmaceutical composition according to claim 6, wherein the peptide is LL-37 in the form of an acetate salt.

11. The pharmaceutical composition according to claim 6, further comprising an aqueous solution.

12. The pharmaceutical composition according to claim 6, wherein the bilayer-forming polar lipid cater contains at least 50% (w/v) of a digalactosyldiacylglycerol.

13. The pharmaceutical composition according to claim 6, wherein the bilayer-forming polar lipid cater contains 50 to 70% (w/w) of digalactosyldiacylglycerols and 30 to 50% (w/w) of other polar lipid.

14. The pharmaceutical composition according to claim 6, wherein the ratio between the peptide as a salt and the bilayer-forming polar lipid cater is 1:

10 to 1;

50 by weight.

15. A bandage for wound healing comprising a pharmaceutical composition according to claim 6.

16. A kit for culturing of epithelial and/or stromal cells comprising:

(a) a peptide comprising an amino acid sequence containing at least 25 amino acids of the N-terminal fragment of LL-37 (SEQ ID NO;

1) or a pharmaceutically acceptable salt or ester or amide thereof; and

(b) a cytotoxicity reducing agent comprising a galactolipid.

19. A growth medium for culturing epithelial and/or stromal cells comprising:

(a) a peptide comprising an amino acid sequence containing at least 25 amino acids of the N-terminal fragment of LL-37 (SEQ ID NO;

1), or a pharmaceutically acceptable salt or ester or amide thereof;

(b) a cytotoxicity reducing agent comprising a galactolipid; and

(c) a basal medium.