Identification of compounds for inhibiting complexation of C-reactive protein with fibronectin

• US 20070134813A1
• Filed: 12/14/2005
• Published: 06/14/2007
• Est. Priority Date: 12/14/2005
• Status: Active Grant

• Current Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Original Assignee: Kimberly-Clark Worldwide Incorporated (Kimberly-Clark Corporation)
• Inventors: Boga, RameshBabu, Malik, Sohail, Quirk, Stephen
• Granted Patent: US 7,645,583 B2
• Time in Patent Office: Days
• US Class Current: 436/518
• CPC Class Codes: G01N 2333/4737 C-reactive protein
• CPC Class Codes: G01N 2333/78 Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin, cold insoluble globulin [CIG]
• CPC Class Codes: G01N 2500/02 Screening involving studying the effect of compounds C on the interaction between interacting molecules A and B (e.g. A = enzyme and B = substrate for A, or A = receptor and B = ligand for the receptor)
• CPC Class Codes: G01N 33/54388 based on lateral flow
• CPC Class Codes: G01N 33/558 using diffusion or migration of antigen or antibody immunochromatographic test strips G01N33/54387
• CPC Class Codes: G01N 33/68 involving proteins, peptides or amino acids involving lipoproteins G01N33/92
• CPC Class Codes: Y10S 436/815 Test for named compound or class of compounds

Abstract

Disclosed are screening processes for identification of compounds that may inhibit complex formation between C-reactive protein and fibronectin. Compounds identified by the disclosed methods may be utilized to inhibit the target analytes that are known to occur during skin aging as well as during the course of several diseases. Accordingly, inhibitory compounds identified by the disclosed methods may be utilized to prevent and treat damaged tissue, inflammatory conditions, cardiovascular conditions, renal conditions, periodontal conditions and obesity, among other conditions. For example, the compounds identified by the disclosed methods may be utilized to treat tissue that has suffered trauma, e.g., burns or wounds, as well as tissue that is inflamed due to any of several causes. The disclosed compounds may also be utilized to improve the accuracy of assays designed to assess CRP or Fn levels in a sample, for instance in a plasma sample, as a risk assessment tool in certain medical conditions and to discover new treatments.

Claims

1. A method for identifying a compound that inhibits the complexation of C-reactive protein with fibronectin, the method comprising:

contacting a first polypeptide with a test compound, the first polypeptide comprising at least a segment of a C-reactive protein, the first polypeptide including the fibronectin binding site of the C-reactive protein;

contacting the first polypeptide with a second polypeptide, the second polypeptide comprising at least a segment of a fibronectin protein, the second polypeptide including the C-reactive protein binding site of the fibronectin; and

determining whether complexation of the first polypeptide with the second polypeptide is decreased in the presence of the test compound, wherein a decrease in said complexation is an indication that the test compound inhibits the complexation of C-reactive protein to fibronectin.

2. The method of claim 1, wherein either the first polypeptide or the second polypeptide is immobilized in the detection zone of a separation device.

3. The method of claim 2, wherein the step of determining whether complexation of the first polypeptide with the second polypeptide is decreased in the presence of the test compound includes generating a detection signal within the detection zone, the detection signal corresponding to the complexation of the first polypeptide with the second polypeptide.

4. The method of claim 2, wherein the detection zone is in fluid communication with detection probes.

5. The method of claim 4, wherein the detection probes are conjugated with the first polypeptide.

6. The method of claim 4, wherein the detection probes are conjugated with the second polypeptide.

7. The method of claim 1, wherein the first polypeptide comprises SEQ ID NO.:

20.

8. The method of claim 7, wherein the first polypeptide comprises SEQ ID NO.:

19.

9. The method of claim 1, wherein the second polypeptide comprises SEQ ID NO.:

18.

10. The method of claim 9, wherein the second polypeptide comprises SEQ ID NO.:

17.

11. A method for inhibiting the complexation of C-reactive protein with fibronectin comprising exposing C-reactive protein and fibronectin to an effective amount of an inhibitor identified according to the method of claim 1.

12. A method for treating tissue comprising:

providing a composition including an inhibitor identified according to the method of claim 1, wherein the composition comprises the inhibitor in an amount between about 0.001% and about 90% by weight of the composition ; and

applying the composition to the tissue, wherein the tissue comprises C-reactive protein and fibronectin.

13. A method for inhibiting the complexation of C-reactive protein with fibronectin comprising exposing C-reactive protein and fibronectin to an effective amount of an inhibitor of the complexation reaction, wherein the inhibitor is a polypeptide inhibitor, a giberellic acid compound or a derivative thereof, a jasmonic acid compound or a derivative thereof, or a mixture thereof.

14. The method of claim 13, wherein the inhibitor is a polypeptide inhibitor including an amino acid sequence from the linking region or the proenzyme leader region of a matrix metalloproteinase or a variant thereof.

15. The method of claim 13, wherein the inhibitor is a polypeptide inhibitor that includes the following segment:

16. The method of claim 15, wherein the inhibitor is a polypeptide inhibitor that includes the following segment:

Xaa₁₀-Xaa₁₃-Xaa₁₂-Xaa₁₃-Xaa₁₄-Xaa₁₅-Xaa₁₆-Xaa₁₇-Xaa₁₈-Xaa₁₉.

17. The method of claim 13, wherein the inhibitor is a polypeptide inhibitor that includes SEQ ID NO.:

14 or SEQ ID NO.;

15.

18. The method of claim 13, wherein the inhibitor is a polypeptide inhibitor that includes SEQ ID NO.:

1, 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, or SEQ ID NO.;

12.

19. The method of claim 13, wherein the inhibitor is a gibberellic acid compound having one of the following general backbone structures:

20. The method of claim 19, wherein the gibberellic acid compound is gibberellic acid or a derivative thereof.

21. The method of claim 13, wherein the inhibitor is a jasmonic acid compound have one of the following structures:

22. The method of claim 21, wherein the jasmonic acid compound is jasmonic acid.

23. The method of claim 13, wherein the inhibitor is a component of a composition, and the inhibitor is in the composition in an amount of between about 0.001% and about 90% by weight of the composition.

24. The method of claim 23, wherein the composition is a sterile solution.

25. The method of claim 23, wherein the composition comprises a pharmaceutically acceptable vehicle.

26. The method of claim 23, wherein the composition is capable of being topically applied to a surface.

27. The method of claim 26, further comprising applying the composition to a skin covering material, a wound dressing material, or an implantable prosthetic device.

28. The method of claim 13, wherein at least one of the C-reactive protein or the fibronectin is at a pH of less than about 6.5.

29. A method for treating tissue comprising:

providing a composition including an inhibitor of the complexation of C-reactive protein with fibronectin, wherein the inhibitor is a polypeptide inhibitor, a giberellic acid compound or a derivative thereof, a jasmonic acid compound or a derivative thereof;

or a mixture thereof, and wherein the composition comprises the inhibitor in an amount between about 0.001% and about 90% by weight of the composition; and

applying the composition to the tissue, wherein the tissue comprises C-reactive protein and fibronectin.

30. The method of claim 29, wherein the inhibitor is a polypeptide inhibitor including an amino acid sequence from the linking region or the proenzyme leader region of a matrix metalloproteinase or a variant thereof.

31. The method of claim 29, wherein the inhibitor is a polypeptide inhibitor that includes the following segment:

32. The method of claim 31, wherein the inhibitor is a polypeptide inhibitor that includes the following segment:

Xaa₁₀-Xaa₁₁-Xaa₁₂-Xaa₁₃-Xaa₁₄-Xaa₁₅-Xaa₁₆-Xaa₁₇-Xaa₁₈-Xaa₁₉.

33. The method of claim 29, wherein the inhibitor is a polypeptide inhibitor that includes SEQ ID NO.:

14 or SEQ ID NO.;

15.

34. The method of claim 29, wherein the inhibitor is a polypeptide inhibitor that includes SEQ ID NO.:

1, 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, or SEQ ID NO.;

12.

35. The method of claim 29, wherein the composition is topically applied to the tissue.

36. The method of claim 29, wherein the tissue is at a pH of less than about 6.5.

37. The method of claim 29, wherein the inhibitor is a gibberellic acid compound having one of the following general backbone structures:

38. The method of claim 29, wherein the inhibitor is a jasmonic acid compound have one of the following structures:

39. The method of claim 29, wherein the composition is applied to the tissue via a pharmaceutically acceptable route of administration selected from the group consisting of orally, intravenously, intramuscularly, and subcutaneously.

40. The method of claim 29, wherein the tissue is that of a mammal afflicted with inflammatory disease.

41. The method of claim 29, wherein the tissue is that of a mammal afflicted with cardiovascular disease, infection, obesity, transplant rejection, necrosis, tumor embolization, acute pancreatitis, a wound, a burn, diabetes, renal disease, or a fracture.