COMPOSITION, METHOD, AND KIT FOR PREPARING PLASMIN

• US 20110201077A1
• Filed: 06/03/2009
• Published: 08/18/2011
• Est. Priority Date: 06/04/2008
• Status: Active Grant

• Current Assignee: Grifols Therapeutics LLC (Grifols SA)
• Original Assignee: Talecris Biotherapeutics Incorporated (Grifols SA)
• Inventors: Koepf, Edward, Zimmerman, Thomas
• Granted Patent: US 8,617,863 B2
• Time in Patent Office: Days
• US Class Current: 435/174
• CPC Class Codes: C07K 14/3153 Streptokinase
• CPC Class Codes: C12N 11/10 the carrier being a carbohydrate
• CPC Class Codes: C12N 9/6435 Plasmin (3.4.21.7), i.e. fibrinolysin
• CPC Class Codes: C12Y 304/21007 Plasmin (3.4.21.7), i.e. fibrinolysin

Abstract

A streptokinase immobilized on a surface, in particular an immobilized plasmin-resistant streptokinase, and compositions, methods and kits of utilizing same for preparing plasmin are provided.

Claims

1. A composition comprising a streptokinase immobilized on a matrix, wherein the streptokinase is a streptokinase mutant characterized as capable of activating plasminogen to plasmin, yet resistant to plasmin degradation relative to its corresponding wild-type streptokinase.

10. A kit for preparing plasmin, the kit comprising:

a) the composition of claim 1; and

b) a plasmin-binding matrix having a molecule disposed thereon having affinity for the plasmin.

2. An article of manufacture comprising a matrix having a streptokinase immobilized thereon, wherein the streptokinase is a streptokinase mutant characterized as capable of activating plasminogen to plasmin, yet resistant to plasmin degradation relative to its corresponding wild-type streptokinase.

3. A method for preparing plasmin, the method comprising:

a) contacting a composition comprising a plasminogen with a streptokinase immobilized on a matrix thereby converting the plasminogen to a plasmin; and

b) purifying the plasmin.

4. The method of claim 3, wherein purifying comprises contacting the composition with a plasmin-binding matrix so that the plasmin is retained by the plasmin-binding matrix, the plasmin-binding matrix having a molecule disposed thereon having affinity for the plasmin.

5. The method of claim 3, wherein the streptokinase is a streptokinase mutant characterized as capable of activating plasminogen to plasmin, yet resistant to plasmin degradation relative to its corresponding wild-type streptokinase

6. The method of claim 3, wherein the streptokinase comprises an amino acid sequence having an amino acid residue other than lysine at a position corresponding to position 85, 412, or both as shown in SEQ ID NO:

1.

7. The method of claim 3, wherein the streptokinase comprises the amino acid sequence shown as amino acid residues 27 through 440 of SEQ ID NO:

2.

8. The method of claim 6, wherein the streptokinase, optionally, further comprises polar or charged residues at one or more positions corresponding to positions 406-410 in SEQ ID NO:

1.

9. The method of claim 6, wherein the amino acid is glutamine or asparagine.

11. The method of claim 7, wherein the streptokinase, optionally, further comprises polar or charged residues at one or more positions corresponding to positions 406-410 in SEQ ID NO:

1.