Uses of sysnergistic bacteriophage lytic enzymes for prevention and treatment of bacterial infections

• US 20060159671A1
• Filed: 12/23/2003
• Published: 07/20/2006
• Est. Priority Date: 12/23/2002
• Status: Abandoned Application

• Current Assignee: Rockefeller University
• Original Assignee: Rockefeller University
• Inventors: Fischetti, Vincent A
• Application Number:
  US10/540,666
• Publication Number:
  US 20060159671A1
• Time in Patent Office: Days
• US Class Current: 424/94.610
• CPC Class Codes: A01N 63/40 Viruses, e.g. bacteriophages
• CPC Class Codes: A01N 63/50 Isolated enzymes; Isolated proteins peptides A01N37/46
• 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 38/47 acting on glycosyl compounds (3.2), e.g. cellulases, lactases
• CPC Class Codes: A61K 38/50 acting on carbon-nitrogen bonds, other than peptide bonds (3.5), e.g. asparaginase
• CPC Class Codes: A61L 2/18 Liquid substances or solutions comprising solids or dissolved gases
• CPC Class Codes: A61L 2/23 Solid substances, e.g. granules, powders, blocks, tablets
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: C12Y 302/01017 Lysozyme (3.2.1.17)
• CPC Class Codes: C12Y 305/01028 N-Acetylmuramoyl-L-alanine amidase (3.5.1.28)

Abstract

Methods for treating and preventing bacterial infections are described using at least two lytic enzymes obtained from a bacteriophage isolated from Streptococci. Two of these enzymes, Pal and Cpl-1, showed synergy when tested for cleavage of peptidoglycan in the cell walls of Streptococcus pneumoniae. Moreover, the combination of these two enzymes resulted in killing of both penicillin sensitive as well as penicillin resistant strains of S. pneumoniae. The synergy displayed by the combined use of these two enzymes may establish a means for identification of agents that mimic or enhance this activity and may lead to the identification of new antimicrobial agents and pharmaceutical compositions useful for treating and preventing a variety of bacterial infections in mammals. Further compositions are described for disinfecting or sanitizing porous and non-porous surfaces suspected of harboring infectious organisms.

Claims

1. A method of treating or preventing bacterial infections, comprising administering to a subject in need of such therapy a therapeutically effective amount of a synergistic combination of:

(a) at least two bacteriophage derived lytic enzymes; and

(b) a suitable carrier for delivery of the lytic enzymes to the site of infection.

2. The method of claim 1, wherein said at least two bacteriophage derived lytic enzymes are selected from the group consisting of an amidase, a muramidase, an endopeptidase, a glucosaminidase and combinations thereof.

3. The method of claim 1, wherein said at least two bacteriophage derived lytic enzymes are an amidase and a muramidase.

4. The method of claim 3, wherein the muramidase is lysozyme and the amidase is Pal.

5. The method of claim 4, wherein the lysozyme is Cpl-1.

6. The method of claim 1, wherein the lytic enzymes are selected from the group consisting of Pal and Cpl-1.

7. The method of claim 1, wherein the bacterial infections are caused by Streptococcus pneumoniae.

8. The method of claim 1, wherein the lytic enzymes decrease the occurrence or severity of local and systemic pneumococcal disease.

9. The method of claim 1, wherein the lytic enzymes prevent or eliminate pneumococcal colonization.

10. The method of claim 1, wherein said at least two lytic enzymes are isolated from the phage of disease causing bacteria, and wherein said disease causing bacteria are gram positive bacteria.

12. The method of claim 10, wherein the gram positive bacteria is Streptococcus.

13. The method of claim 12, wherein the Streptococcus is Streptococcus pneumoniae.

11. A method of treating or preventing bacterial infections, comprising administering to a subject in need of such therapy a therapeutically effective amount of bacteriophage derived lytic enzymes selected from the group consisting of an amidase, a muramidase, an endopeptidase, a glucosaminidase, and synergistic combinations thereof.

14. The method of claim 11, wherein the amidase is Pal and the muramidase is Cpl-1.

15. A pharmaceutical composition comprising:

(a) at least two therapeutically effective synergistic bacteriophage derived lytic enzymes; and

(b) a carrier suitable for delivery of the lytic enzymes to the site of infection.

16. The composition of claim 15, wherein said at least two bacteriophage derived lytic enzymes are selected from the group consisting of an amidase, a muramidase, an endopeptidase, a glucosaminidase and combinations thereof.

17. The pharmaceutical composition of claim 16, wherein the amidase is Pal and the muramidase is Cpl-1.

18. A screening method for identifying agents capable of enhancing the activity of Pal and Cpl-1, comprising:

(a) preparing purified Pal and Cpl-1;

(b) contacting the Pal and Cpl-1 to a bacteria having radioactively labeled peptidoglycan in the cell wall in the presence or absence of a test compound under conditions which allow binding to the peptidoglycan; and

(c) determining the amount of peptidoglycan cleavage, wherein an agent capable of enhancing Pal and Cpl-1 activity is identified when the release of radioactivity is enhanced in the presence but not the absence of the agent.

19. An anti-microbial composition for sanitizing or decontaminating porous or non-porous surfaces comprising at least two bacteriophage derived synergistic lytic enzymes.

20. The composition of claim 19, wherein said at least two bacteriophage derived synergistic lytic enzymes are selected from the group consisting of an amidase, a muramidase, an endopeptidase, a glucosaminidase and combinations thereof.

21. The composition of claim 19, wherein said at least two bacteriophage derived lytic enzymes are an amidase and a muramidase.

22. The composition of claim 21, wherein the muramidase is lysozyme and the amidase is Pal.

23. The composition of claim 22, wherein the lysozyme is Cpl-1.

24. A method for decontaminating inanimate surfaces suspected of containing infectious bacteria comprising treatment of said surfaces with a bacteriocidal or bacteriostatically effective amount of the composition of claim 19.

25. (canceled)

26. (canceled)

27. (canceled)

28. (canceled)

29. (canceled)