Fibrinogen binding moieties

• US 7,041,790 B2
• Filed: 03/25/2003
• Issued: 05/09/2006
• Est. Priority Date: 03/26/2002
• Status: Active Grant

• Current Assignee: Takeda Pharmaceutical Company Limited
• Original Assignee: Dyax Corporation (Shire plc)
• Inventors: Sato, Aaron K., Wescott, Charles R.
• Primary Examiner(s): Weber, Jon
• Assistant Examiner(s): NOAKES, SUZANNE MARIE
• Application Number:
  US10/396,073
• Publication Number:
  US 20030207330A1
• Time in Patent Office: 1,141 Days
• Field of Search: 530324-328, 530/382, 514/2
• US Class Current: 530/328
• CPC Class Codes: C07K 14/00 Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
• CPC Class Codes: C07K 7/06 having 5 to 11 amino acids
• CPC Class Codes: C07K 7/08 having 12 to 20 amino acids gastrins C07K14/595; somatostatins C07K14/655; melanotropins C07K14/68
• CPC Class Codes: G01N 2333/75 Fibrin; Fibrinogen
• CPC Class Codes: G01N 33/86 involving blood coagulating time or factors, or their receptors

Abstract

Compositions comprising non-naturally occurring fibrinogen binding moieties are described, together with methods of use thereof, e.g., for detecting or isolating fibrinogen molecules in a solution, for blood circulation imaging, and for linking therapeutics or other molecules to fibrinogen. Preferred binding peptides having a high affinity for fibrinogen are particularly disclosed.

Claims

1. A fibrinogen binding moiety comprising a polypeptide comprising an amino acid sequence of the formula:

2. The binding moiety according to claim 1, wherein said polypeptide comprises an amino acid sequence selected from the group consisting of:

3. The binding moiety according to claim 1, wherein said polypeptide comprises an amino acid sequence of the formula:

Ac-AEGTGSLYLCNSYPMHPYCNPGK-NH₂ (SEQ ID NO;

48)

where Ac—

denotes acetylation and —

NH₂ denotes amidation.

4. A fibrinogen binding moiety according to any one of claims 1 or 3, wherein said polypeptide is expressed by a recombinant bacteriophage.

5. A composition of matter comprising a binding moiety according to any one of claims 1 or 3 linked to a solid support material selected from the group consisting of cellulose, plastic, metal, rubber, wood, nylon, glass, acrylamide, agarose, and combinations thereof.

6. The composition according to claim 4, wherein said binding moiety is linked to a chromatographic matrix material.

7. A composition, of matter comprising a binding moiety according to any one of claims 1 or 3 linked to a detectable label.

8. The composition according to claim 7, wherein said detectable label is selected from the group consisting of radionuclides, detectable proteins, epitope tags, biotin, streptavidin, enzymes, antibodies, and fluorescent labels.

9. The composition according to claim 8, wherein the detectable label is a technetium-containing compound.

10. A composition of matter comprising a binding moiety according to any one of claims 1 or 3 linked to a drug, biopharmaceutical, or polypeptide of interest.

11. A method for detecting fibrinogen in a solution comprising:

(a) contacting said solution with a binding moiety according to any one of claims 1 or 3 under conditions wherein said binding moiety will form a complex with said fibrinogen, and (b) detecting said complex.

12. The method according to claim 11, wherein the solution is blood and the binding moiety is linked to a detectable label.

13. A method for isolating fibrinogen from a solution containing it comprising:

(a) immobilizing a composition according to any one of claims 1 or 3 on a solid support;

(b) contacting a solution containing fibrinogen with said solid support of (a) under conditions wherein said binding moiety will form a complex with said fibrinogen; and

, thereafter, (c) separating the solid support from the unbound components of said solution.

14. The method according to claim 13, wherein said solid support is selected from the group consisting of chromatographic matrix materials, filters, magnetic beads, and the surface of a plastic or glass container.

15. The method according to claim 13, further comprising the step:

(d) eluting and recovering the fibrinogen from said support.

16. The method according to claim 15, wherein the eluant used in step (d) is a citrate buffer or a glycine buffer.

17. The method according to claim 13, wherein the solution is selected from the group consisting of a recombinant eukaryotic or prokaryotic cell extract, and whole blood.

18. A method for isolating fibrinogen from a solution containing it comprising:

(a) contacting a solution containing fibrinogen with a binding moiety according to any one of claims 1 or 3, in which said binding moiety is conjugated to an affinity ligand, under conditions suitable for formation of a binding complex between said binding moiety and said fibrinogen;

(b) contacting the solution of step (a) with an immobilized binding partner for said affinity ligand under conditions suitable for formation of a binding complex between the affinity ligand and the binding partner; and

(c) removing unbound materials in the solution from the complex formed in step (b).

19. The method according to claim 18, wherein said affinity ligand is a polyhistidine tag.

20. The method according to claim 18, wherein said affinity ligand is biotin and said immobilized binding partner for said affinity ligand is streptavidin.

21. A method for assessing blood circulation in a subject comprising:

(a) introducing into the circulatory system of the subject a composition comprising a detectable binding moiety according to claim 7, and (b) detecting any circulation of complexes formed between said detectable binding moiety and fibrinogen in the circulatory system of said subject.

22. The method according to claim 21, wherein said detectable compound consists essentially of a complex of said detectably labeled binding moiety and fibrinogen.

23. The method according to claim 21, wherein the binding moiety is detectably labeled with a magnetic resonance imaging agent.