Modified peptides as therapeutic agents

• US 20040053845A1
• Filed: 07/31/2003
• Published: 03/18/2004
• Est. Priority Date: 10/23/1998
• Status: Active Grant

• Current Assignee: Amgen Inc.
• Original Assignee: Amgen Inc.
• Inventors: Liu, Chuan-Fa, Feige, Ulrich, Cheetham, Janet C., Boone, Thomas Charles
• Granted Patent: US 7,189,827 B2
• Time in Patent Office: Days
• US Class Current: 514/12
• 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 11/06 Antiasthmatics
• CPC Class Codes: A61P 19/00 Drugs for skeletal disorders
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 3/04 Anorexiants; Antiobesity agents
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 31/18 for HIV
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 35/02 specific for leukemia
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: A61P 7/00 Drugs for disorders of the blood or the extracellular fluid
• CPC Class Codes: A61P 7/02 Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
• CPC Class Codes: A61P 7/04 Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
• CPC Class Codes: A61P 7/06 Antianaemics
• CPC Class Codes: A61P 7/08 Plasma substitutes; Perfusion solutions; Dialytics or haemodialytics; Drugs for electrolytic or acid-base disorders, e.g. hypovolemic shock artificial tears A61P27/04
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07K 14/505 Erythropoietin [EPO]
• CPC Class Codes: C07K 14/52 : Cytokines; Lymphokines; Interferons
• CPC Class Codes: C07K 14/524 : Thrombopoietin, i.e. C-MPL ligand
• CPC Class Codes: C07K 14/525 : Tumour necrosis factor [TNF]
• CPC Class Codes: C07K 14/545 : IL-1
• CPC Class Codes: C07K 14/8146 : Metalloprotease (E.C. 3.4.24) inhibitors, e.g. tissue inhibitor of metallo proteinase, TIMP
• CPC Class Codes: C07K 2319/00 : Fusion polypeptide
• CPC Class Codes: C07K 2319/30 : Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
• CPC Class Codes: C12N 9/6491 : Matrix metalloproteases [MMP's], e.g. interstitial collagenase (3.4.24.7); Stromelysins (3.4.24.17; 3.2.1.22); Matrilysin (3.4.24.23)

Abstract

The present invention concerns fusion of Fc domains with biologically active peptides and a process for preparing pharmaceutical agents using biologically active peptides. In this invention, pharmacologically active compounds are prepared by a process comprising:

a) selecting at least one peptide that modulates the activity of a protein of interest; and

b) preparing a pharmacologic agent comprising an Fc domain covalently linked to at least one amino acid of the selected peptide.

Linkage to the vehicle increases the half-life of the peptide, which otherwise would be quickly degraded in vivo. The preferred vehicle is an Fc domain. The peptide is preferably selected by phage display, E. coli display, ribosome display, RNA-peptide screening, or chemical-peptide screening.

Claims

1. A composition of matter of the formula

(X¹)_a—

F¹—

(X²)_b

and multimers thereof, wherein;

F¹ is an Fc domain;

X¹ and X² are each independently selected from -(L¹)_c-P¹, -(L¹)_c-P¹-(L¹)_c-P¹-(L²)_d-P²-(L³)_e-P³-(L⁴)_f-P⁴

P¹, P², P³, and P⁴ are each independently sequences of pharmacologically active peptides;

L¹, L², L³, and L⁴ are each independently linkers; and

a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a and b is 1.

2. The composition of matter of claim 1 of the formulae

X¹— F¹

or

F¹—

X².

3. The composition of matter of claim 1 of the formula

F¹-(L¹)_c-P¹.

4. The composition of matter of claim 1 of the formula

F¹-(L¹)_c-P¹-(L²)_d-P².

5. The composition of matter of claim 1 wherein F¹ is an IgG Fc domain.

6. The composition of matter of claim 1 wherein F¹ is an IgG1 Fc domain.

7. The composition of matter of claim 1 wherein F¹ comprises the sequence of SEQ ID NO:

2.

8. The composition of matter of claim 1 wherein X¹ and X² comprise an IL-1 antagonist peptide sequence.

9. The composition of matter of claim 8 wherein the IL-1 antagonist peptide sequence is selected from SEQ ID NOS:

212, 907, 908, 909, 910, 917, and 979.

10. The composition of matter of claim 8 wherein the IL-1 antagonist peptide sequence is selected from SEQ ID NOS:

213 to 271, 671 to 906, 911 to 916, and 918 to 1023.

11. The composition of matter of claim 8 wherein F¹ comprises the sequence of SEQ ID NO:

2.

12. The composition of matter of claim 1 wherein X¹ and X² comprise an EPO-mimetic peptide sequence.

13. The composition of matter of claim 12 wherein the EPO-mimetic peptide sequence is selected from Table 5.

14. The composition of matter of claim 12 wherein F¹ comprises the sequence of SEQ ID NO:

2.

15. The composition of matter of claim 12 comprising a sequence selected from SEQ ID NOS:

83, 84, 85, 124, 419, 420, 421, and 461.

16. The composition of matter of claim 12 comprising a sequence selected from SEQ ID NOS:

339 and 340.

17. The composition of matter of claim 12 comprising a sequence selected from SEQ ID NOS:

20 and 22.

18. The composition of matter of claim 3 wherein P¹ is a TPO-mimetic peptide sequence.

19. The composition of matter of claim 18 wherein P¹ is a TPO-mimetic peptide sequence selected from Table 6.

20. The composition of matter of claim 18 wherein F¹ comprises the sequence of SEQ ID NO:

2.

21. The composition of matter of claim 18 having a sequence selected from SEQ ID NOS:

6 and 12.

22. A DNA encoding a composition of matter of any of claims 1 to 21.

23. An expression vector comprising the DNA of claim 22.

24. A host cell comprising the expression vector of claim 23.

25. The cell of claim 24, wherein the cell is an E. coli cell.

26. A process for preparing a pharmacologically active compound, which comprises

a) selecting at least one randomized peptide that modulates the activity of a protein of interest;

and

b) preparing a pharmacologic agent comprising at least one Fc domain covalently linked to at least one amino acid sequence of the selected peptide or peptides.

27. The process of claim 26, wherein the peptide is selected in a process comprising screening of a phage display library, an E. coli display library, a ribosomal library, or a chemical peptide library.

28. The process of claim 26, wherein the preparation of the pharmacologic agent is carried out by:

a) preparing a gene construct comprising a nucleic acid sequence encoding the selected peptide and a nucleic acid sequence encoding an Fc domain; and

b) expressing the gene construct.

29. The process of claim 26, wherein the gene construct is expressed in an E. coli cell.

30. The process of claim 26, wherein the protein of interest is a cell surface receptor.

31. The process of claim 26, wherein the protein of interest has a linear epitope.

32. The process of claim 26, wherein the protein of interest is a cytokine receptor.

33. The process of claim 26, wherein the peptide is an EPO-mimetic peptide.

34. The process of claim 26, wherein the peptide is a TPO-mimetic peptide.

35. The process of claim 26, wherein the peptide is an IL-1 antagonist peptide.

36. The process of claim 26, wherein the peptide is an MMP inhibitor peptide or a VEGF antagonist peptide.

37. The process of claim 26, wherein the peptide is a TNF-antagonist peptide.

38. The process of claim 26, wherein the peptide is a CTLA4-mimetic peptide.

39. The process of claim 26, wherein the peptide is selected from Tables 4 to 20.

40. The process of claim 26, wherein the selection of the peptide is carried out by a process comprising:

a) preparing a gene construct comprising a nucleic acid sequence encoding a first selected peptide and a nucleic acid sequence encoding an Fc domain;

b) conducting a polymerase chain reaction using the gene construct and mutagenic primers, wherein

i) a first mutagenic primer comprises a nucleic acid sequence complementary to a sequence at or near the 5′

end of a coding strand of the gene construct, and

ii) a second mutagenic primer comprises a nucleic acid sequence complementary to the 3′

end of the noncoding strand of the gene construct.

41. The process of claim 26, wherein the compound is derivatized.

42. The process of claim 26, wherein the derivatized compound comprises a cyclic portion, a cross-linking site, a non-peptidyl linkage, an N-terminal replacement, a C-terminal replacement, or a modified amino acid moiety.

43. The process of claim 26 wherein the Fc domain is an IgG Fc domain.

44. The process of claim 26, wherein the vehicle is an IgG1 Fc domain.

45. The process of claim 26, wherein the vehicle comprises the sequence of SEQ ID NO:

2.

46. The process of claim 26, wherein the compound prepared is of the formula

(X¹)_a—

F¹—

(X²)_b

and multimers thereof,wherein;

F¹ is an Fc domain;

X¹ and X² are each independently selected from -(L¹)_c-P¹, -(L¹)_c-P¹-(L²)_d-P², -(L¹)_c-P¹-(L²)_d-P²-(L³)_e-P³, and -(L¹)_c-P¹-(L²)_d-P²-(L³)_e-P³-(L⁴)_f-P⁴

P¹, P², P³, and P⁴ are each independently sequences of pharmacologically active peptides;

L¹, L², L³, and L⁴ are each independently linkers; and

a, b, c, d, e, and f are each independently 0 or 1, provided that at least one of a and b is 1.

47. The process of claim 46, wherein the compound prepared is of the formulae

X¹—

F¹

or

F¹—

X².

48. The process of claim 46, wherein the compound prepared is of the formulae

F¹-(L¹)_cP¹

or

F¹-(L¹)_c-P¹-(L²)_d-P².

49. The process of claim 46, wherein F¹ is an IgG Fc domain.

50. The process of claim 46, wherein F¹ is an IgG1 Fc domain.

51. The process of claim 46, wherein F¹ comprises the sequence of SEQ ID NO:

2.