Novel isoforms of vascular endothelial cell growth inhibitor

• US 20030170242A1
• Filed: 11/12/2002
• Published: 09/11/2003
• Est. Priority Date: 11/09/2001
• Status: Active Grant

• Current Assignee: Georgetown University
• Original Assignee: Georgetown University
• Inventors: Pan, Hongguang, Li, Luyuan
• Granted Patent: US 7,498,407 B2
• Time in Patent Office: Days
• US Class Current: 424/145.1
• 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: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07K 14/70575 NGF/TNF-superfamily, e.g. CD70, CD95L, CD153, CD154 NGF C07K14/48, TNF C07K14/525
• CPC Class Codes: C07K 2319/00 Fusion polypeptide

Abstract

This invention discloses two new VEGI isoforms named VEGI-_192a and VEGI-_192b consisting of 192 amino acid residues. These isoforms show endothelial cell-specific expression and share a C-terminal 151-residues segment with the previously described VEGI-₁₇₄ and VEGI-₂₅₁. Methods of using these isoforms of VEGI in diagnosing, screening agonist and antagonist of the isoforms, and treating various angiogenesis-related diseases are also disclosed.

Claims

1. An isolated polynucleotide comprising:

(a) the sequence of SEQ ID NO;

1;

(b) at least about 15 contiguous nucleotides of SEQ ID NO;

1, wherein said contiguous nucleotides are within nucleotides 1-93 of SEQ ID NO;

1;

(c) at least 15 contiguous nucleotides of SEQ ID NO;

1, wherein said contiguous nucleotides comprise nucleotides 93 and 94 of SEQ ID NO;

1;

(d) a nucleotide sequence encoding the polypeptide of SEQ ID NO;

4;

(e) a nucleotide sequence encoding at least 10 contiguous amino acids of SEQ ID NO;

4, wherein said contiguous amino acids are within amino acids 1-26 of SEQ ID NO;

4;

(f) a nucleotide sequence encoding at least 10 contiguous amino acids of SEQ ID NO;

4, wherein said contiguous amino acids comprise amino acids 26 and 27 of SEQ ID NO;

4;

or (g) a complement of the above thereof.

2. The polynucleotide of claim 1, wherein the polynucleotide comprises the sequence of SEQ ID NO:

1.

3. The polynucleotide of claim 1, wherein said polynucleotide of (b) or (c) comprises at least about 20 contiguous nucleotides.

4. The polynucleotide of claim 1, wherein said polynucleotide further comprising a detectable label.

5. The polynucleotide of claim 1, wherein said polynucleotide is immobilized on a surface.

6. The polynucleotide of claim 1, wherein said polynucleotide encodes a polypeptide having the biological activity of anti-angiogenesis.

7. The polynucleotide of claim 1, wherein said polynucleotide encodes a fusion protein comprising the polypeptide of SEQ ID NO:

4.

8. A vector comprising a polynucleotide according to claim 1.

9. The vector of claim 8, wherein the vector is an expression vector.

10. A host cell comprising a vector according to claim 8.

11. A pharmaceutical composition comprising a polynucleotide according to claim 6 and a pharmaceutically acceptable excipient.

54. A method for detecting VEGI-_192a polynucleotide comprising contacting a sample from an individual with an oligonucleotide which selectively binds to the polynucleotide of claim 1;

and detecting the presence or absence of a duplex formed between the oligonucleotide and a polynucleotide in the sample.

12. An isolated polynucleotide comprising:

(a) the sequence of SEQ ID NO;

2;

(b) at least about 15 contiguous nucleotides of SEQ ID NO;

2, wherein said contiguous nucleotides are within nucleotides 1-386 of SEQ ID NO;

2;

(c) at least 15 contiguous nucleotides of SEQ ID NO;

2, wherein said contiguous nucleotides comprise nucleotides 386 and 387 of SEQ ID NO;

2;

(d) a nucleotide sequence encoding a polypeptide of SEQ ID NO;

5;

(e) a nucleotide sequence encoding at least 10 contiguous amino acids of SEQ ID NO;

5, wherein said contiguous amino acids are within amino acids 1-26 of SEQ ID NO;

5;

(f) a nucleotide sequence encoding at least 10 contiguous amino acids of SEQ ID NO;

5, wherein said contiguous amino acids comprise amino acids 26 and 27 of SEQ ID NO;

5;

or (g) a complement of the above thereof.

13. The polynucleotide of claim 12, wherein the polynucleotide comprises the sequence of SEQ ID NO:

2.

14. The polynucleotide of claim 12, wherein said polynucleotide of (b) or (c) comprises at least about 20 contiguous nucleotides.

15. The polynucleotide of claim 12, wherein said polynucleotide further comprising a detectable label.

16. The polynucleotide of claim 12, wherein said polynucleotide is immobilized on a surface.

17. The polynucleotide of claim 12, wherein said polynucleotide encodes a polypeptide having the biological activity of anti-angiogenesis.

18. The polynucleotide of claim 12, wherein said polynucleotide encodes a fusion protein comprising the polypeptide of SEQ ID NO:

5.

19. A vector comprising the polynucleotide according to claim 12.

20. The vector of claim 19, wherein the vector is an expression vector.

21. A host cell comprising the vector of claim 19.

22. A pharmaceutical composition comprising a polynucleotide according to claim 17 and a pharmaceutically acceptable excipient.

56. A method for detecting VEGI-_192b polynucleotide comprising contacting a sample from an individual with an oligonucleotide which selectively binds to the polynucleotide of claim 12;

and detecting the presence or absence of a duplex formed between the oligonucleotide and a polynucleotide in the sample.

23. An isolated polypeptide comprising:

(a) the sequence of SEQ ID NO;

4;

(b) at least 10 contiguous amino acids of SEQ ID NO;

4, wherein said contiguous amino acids are within amino acids 1-26 of SEQ ID NO;

4;

or (c) at least 10 contiguous amino acids of SEQ ID NO;

4, wherein said contiguous comprise amino acids 26 and 27 of SEQ ID NO;

4.

24. The polypeptide of claim 23, wherein the polypeptide comprises the sequence of SEQ ID NO:

4.

25. The polypeptide of claim 23, wherein the polypeptide inhibits vascular endothelial cell growth.

26. The polypeptide of claim 23, wherein the polypeptide has angiogenesis inhibiting activity.

27. A fusion protein comprising a polypeptide according to claim 23.

28. The fusion protein of claim 27, wherein the fusion protein comprises the polypeptide of SEQ ID NO:

4.

29. A pharmaceutical composition comprising the polypeptide of claim 26 and a pharmaceutically acceptable excipient.

37. An antibody that selectively binds to a polypeptide according to claim 23.

38. The antibody of claim 37, wherein the polypeptide comprises SEQ ID NO:

4.

47. A therapeutic method for inhibiting angiogenesis, comprising administering to an individual the composition of claim 29 in a dosage sufficient to inhibit angiogenesis.

48. A therapeutic method according to claim 47, wherein the composition comprises the polypeptide of SEQ ID NO:

4.

49. A therapeutic method for suppressing tumor growth comprising administering to an individual the composition of claim 29 in a dosage sufficient to suppress tumor growth.

50. A therapeutic method according to claim 49, wherein the composition comprises the polypeptide of SEQ ID NO:

4.

51. A method for testing an agent or a drug for angiogenic inhibitory activity, said method comprising measuring ability of said agent or drug to increase the anti-angiogenic activity of the polypeptide of claim 23.

52. A method for testing an agent or a drug for promoting angiogenesis, said method comprising measuring ability of said agent or drug to reduce or eliminate anti-angiogenic activity of the polypeptide of claim 23.

53. A method for detecting VEGI-_192a comprising contacting a sample from an individual with an antibody which selectively binds to the polypeptide of claim 23;

and detecting the presence or absence of a complex formed between a polypeptide in the sample and the antibody.

30. An isolated polypeptide comprising:

(a) the sequence of SEQ ID NO;

5;

(b) at least 10 contiguous amino acids of SEQ ID NO;

5, wherein said contiguous amino acids are within amino acids 1-26 of SEQ ID NO;

5;

or (c) at least 10 contiguous amino acids of SEQ ID NO;

5, wherein said contiguous comprise amino acids 26 and 27 of SEQ ID NO;

5.

31. The polypeptide of claim 30, wherein the polypeptide comprises the sequence of SEQ ID NO:

5.

32. The polypeptide of claim 30, wherein the polypeptide inhibits vascular endothelial cell growth.

33. The polypeptide of claim 30, wherein the polypeptide has angiogenesis inhibiting activity.

34. A fusion protein comprising a polypeptide according to claim 30.

35. The fusion protein of claim 34, wherein the fusion protein comprises the polypeptide of SEQ ID NO:

5.

36. A pharmaceutical composition comprising the polypeptide of claim 33 and a pharmaceutically acceptable excipient.

39. An antibody that selectively binds to a polypeptide according to claim 30.

40. The antibody of claim 39, wherein the polypeptide comprises SEQ ID NO:

5.

55. A method for detecting VEGI-_192b comprising contacting a sample from an individual with an antibody which selectively binds to the polypeptide of claim 30;

and detecting the presence or absence of a complex formed between a polypeptide in the sample and the antibodies.

41. An antibody that selectively binds to the polypeptide of SEQ ID NO:

4 and the polypeptide of SEQ ID NO;

5.

42. A method of inhibiting angiogenesis in a tissue or cell comprising causing an effective amount of a polypeptide encoded by the polynucleotide of SEQ ID NO:

3 to come into contact with, or be expressed in the proximity of, the tissue or the cell.

43. A therapeutic method for inhibiting angiogenesis, comprising administering to an individual a composition comprising a polynucleotide encoding the polypeptide of SEQ ID NO:

6 in a dosage sufficient to inhibit angiogenesis.

44. A therapeutic method according to claim 43, wherein the polynucleotide is operably associated with a regulatory sequence that controls gene expression.

45. A therapeutic method for suppressing tumor growth comprising administering to an individual a composition comprising a polynucleotide encoding the polypeptide of SEQ ID NO:

6 in a pharmaceutically acceptable carrier in a dosage sufficient to suppress tumor growth.

46. A therapeutic method according to claim 45, wherein the polynucleotide is operably associated with a regulatory sequence that controls gene expression.