Polypeptide compounds for inhibiting angiogenesis and tumor growth

• US 20060204512A1
• Filed: 09/23/2005
• Published: 09/14/2006
• Est. Priority Date: 09/23/2004
• Status: Abandoned Application

• Current Assignee: Vasgene Therapeutics, Inc.
• Original Assignee: Vasgene Therapeutics, Inc.
• Inventors: Kertesz, Nathalie, Reddy, Ramachandra, Krasnoperov, Valery, Gill, Parkash, Zozulya, Sergey
• Application Number:
  US11/234,482
• Publication Number:
  US 20060204512A1
• Time in Patent Office: Days
• US Class Current: 424/185.100
• 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 38/385 Serum albumin
• CPC Class Codes: A61K 47/60 the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: A61P 35/04 specific for metastasis
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 14/715 for cytokines; for lymphokines; for interferons

Abstract

In certain embodiments, this present invention provides polypeptide compositions, including compositions containing a modified polypeptide, and methods for inhibiting Ephrin B2 or EphB4 activity. In other embodiments, the present invention provides methods and compositions for treating cancer or for treating angiogenesis-associated diseases.

Claims

1. An isolated soluble polypeptide comprising an amino acid sequence of an extracellular domain of an EphB4 protein, wherein the polypeptide is a monomer and binds specifically to an Ephrin B2 polypeptide.

2. The polypeptide of claim 1, comprising a globular domain of an EphB4 protein or a sequence that is at least 90% identical to a globular domain of EphB4.

3. The polypeptide of claim 1, comprising a sequence at least 90% identical to residues 29-197 of the amino acid sequence defined by FIG. 65 (SEQ ID NO:

10).

4. The polypeptide of claim 1, further comprising a modification that increases serum half-life.

5. The polypeptide of claim 4, wherein said modification comprises a polyethylene glycol group.

6. The polypeptide of claim 5, wherein said modification is a single polyethylene glycol group covalently bonded to the polypeptide.

7. The polypeptide of claim 5, wherein said polypeptide is covalently bonded to two polyethylene glycol groups.

8. The polypeptide of claim 5, wherein said polypeptide is covalently bonded to multiple polyethylene glycol groups.

9. The polypeptide of claim 5, wherein said polyethylene glycol group has a molecular weight of from about 10 to about 40 kDa.

10. The polypeptide of claim 5, wherein the polyethylene glycol group has a molecular weight of from about 30 to about 40 kDa.

11. The polypeptide of claim 5, wherein said polyethylene glycol group is selected from the group of linear PEG chains and branched PEG chains.

12. The polypeptide of claim 5, wherein said polyethylene glycol group is attached to a group selected from the lysine side chains and the N-terminal amino group of the EphB4 polypeptide.

13. The polypeptide of claim 4, wherein said polypeptide has a serum half-life in vivo at least 50% greater than that of an unmodified EphB4 polypeptide.

14. The polypeptide of claim 4, wherein said polypeptide has a serum half-life in vivo at least 100% greater than that of an unmodified EphB4 polypeptide.

15. The polypeptide of claim 4, wherein the polypeptide is a fusion protein.

16. The polypeptide of claim 15, wherein the polypeptide comprises an albumin protein or fragments thereof.

17. The polypeptide of claim 16, wherein said albumin protein is selected from the group consisting of a human serum albumin (HSA) and bovine serum albumin (BSA).

18. The polypeptide of claim 16, wherein the albumin is a naturally occurring variant.

19. The polypeptide of claim 1, wherein the polypeptide has one or more activities selected from the group consisting of:

(a) inhibition of EphrinB2 activity;

(b) inhibition of EphrinB2 kinase activity;

(c) inhibition of the interaction between EphB4 and EphrinB2;

(d) inhibition of EphB4 kinase activity;

(e) inhibition of clustering of Ephrin B2; and

(f) inhibition of clustering of EphB4.

20. The polypeptide of claim 4, wherein the polypeptide has enhanced in vivo stability relative to the unmodified wildtype polypeptide.

21. A pharmaceutical composition comprising a polypeptide of claim 1, and a pharmaceutically acceptable carrier.

22. A method of inhibiting signaling through Ephrin B2/EphB4 pathway in a cell, comprising contacting the cell with an effective amount of a polypeptide of claim 1.

23. A method of reducing the growth rate of a tumor, comprising administering an amount of a polypeptide of claim 1, sufficient to reduce the growth rate of the tumor.

24. A method for treating a patient suffering from a cancer, comprising administering to the patient a polypeptide of claim 1.

25. A method of inhibiting angiogenesis, comprising contacting a cell with a polypeptide of claim 1.

26. A method for treating a patient suffering from an angiogenesis-associated disease, comprising administering to the patient a polypeptide of claim 1.

27. The polypeptide of claim 1, wherein the polypeptide comprises one or more modified amino acid residues.

28. A cosmetic composition comprising the polypeptide of claim 1, and a pharmaceutically acceptable carrier.

29. A method of reducing the growth rate of a tumor, comprising administering an amount of a polypeptide agent sufficient to reduce the growth rate of the tumor, wherein the polypeptide agent is selected from the group consisting of:

(a) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an EphB4 protein, wherein the EphB4 polypeptide is a monomer and binds specifically to an Ephrin B2 polypeptide;

(b) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an Ephrin B2 protein, wherein the soluble Ephrin B2 polypeptide is a monomer and binds with high affinity to an EphB4 polypeptide.

30. The method of claim 29, wherein the tumor comprises cells expressing a higher level of EphB4 and/or EphrinB2 than noncancerous cells of a comparable tissue.

31. A method for treating a patient suffering from a cancer, comprising administering to the patient a polypeptide agent selected from the group consisting of:

(a) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an EphB4 protein, wherein the EphB4 polypeptide is a monomer and binds specifically to an Ephrin B2 polypeptide;

(b) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an Ephrin B2 protein, wherein the soluble Ephrin B2 polypeptide is a monomer and binds with high affinity to an EphB4 polypeptide.

32. The method of claim 31, wherein the cancer comprises cancer cells expressing EphrinB2 and/or EphB4 at a higher level than noncancerous cells of a comparable tissue.

33. The method of claim 31, wherein the cancer is metastatic cancer.

34. The method of claim 31, wherein the tumor is selected from the group consisting of colon carcinoma, breast tumor, mesothelioma, prostate tumor, squamous cell carcinoma, Kaposi sarcoma, and leukemia.

35. The method of claim 31, wherein the cancer is an angiogenesis-dependent cancer.

36. The method of claim 31, wherein the cancer is an angiogenesis-independent cancer.

37. The method of claim 31, wherein the polypeptide agent is a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an Ephrin B2 protein, wherein the soluble Ephrin B2 polypeptide is a monomer and binds with high affinity to an EphB4 polypeptide and further comprises a modification that increases serum half-life.

38. The method of claim 31, further including administering at least one additional anti-cancer chemotherapeutic agent that inhibits cancer cells in an additive or synergistic manner with the polypeptide agent.

39. A method of inhibiting angiogenesis, comprising contacting a cell an amount of a polypeptide agent sufficient to inhibit angiogenesis, wherein the polypeptide agent is selected from the group consisting of:

(a) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an EphB4 protein, wherein the EphB4 polypeptide is a monomer and binds specifically to an Ephrin B2 polypeptide;

(b) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an Ephrin B2 protein, wherein the soluble Ephrin B2 polypeptide is a monomer and binds with high affinity to an EphB4 polypeptide.

40. A method for treating a patient suffering from an angiogenesis-associated disease, comprising administering to the patient a polypeptide agent selected from the group consisting of:

(a) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an EphB4 protein, wherein the EphB4 polypeptide is a monomer and binds specifically to an Ephrin B2 polypeptide;

(b) a soluble polypeptide comprising an amino acid sequence of an extracellular domain of an Ephrin B2 protein, wherein the soluble Ephrin B2 polypeptide is a monomer and binds with high affinity to an EphB4 polypeptide.

41. An isolated soluble polypeptide comprising an amino acid sequence of a fibronectin type 3 domain of an EphB4 protein, wherein the polypeptide inhibits tumor growth in a mouse xenograft model of cancer.

42. The polypeptide of claim 41, wherein the polypeptide does not bind to EphrinB2.

43. The polypeptide of claim 41, wherein the polypeptide does not include a substantial portion of the globular domain of an EphB4 protein.

44. The polypeptide of claim 41, wherein the polypeptide comprises an amino acid sequence of amino acids 324-526 of the sequence of FIG. 65 (SEQ ID NO:

10).

45. The polypeptide of claim 41, wherein the polypeptide is a monomer.

46. The polypeptide of claim 41, wherein the polypeptide further comprises a modification that increases serum half-life.

47. A polypeptide dimer or multimers comprising two or more polypeptides of claim 41.