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Stable protease composition

A composition, serine protease technology, applied in the field of enzyme composition, can solve the problem of loss of enzyme initial activity and so on

Inactive Publication Date: 2008-09-24
TROBIO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, even in this case, a considerable part of the initial activity of the enzyme is still lost, since higher concentrations of inhibitors are required for good stabilization

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0131] Example 1 - Stabilization of human thrombin

[0132] To determine the procoagulant activity of the thrombin solution, the coagulation of the fibrinogen solution (1.3 mg / ml) was determined after addition of different dilutions of a specific human thrombin (from plasma, 3300 units / mg, Biovitrum AB, Sweden) solution time. Clotting times were measured with an Amelungen Kc 1 hemagglutination meter (Amelungen, Germany). In order to study the stability of thrombin solutions with different additives, the samples were incubated in a constant temperature chamber maintained at 37°C. Aliquots were taken at various time intervals and the thrombin activity retained in these aliquots was determined. Activity decay curves can be constructed from the values ​​obtained.

[0133] The activity decay curve for a 1 mg / ml solution of thrombin in 10 mM HEPES, 0.13 M NaCl buffer, pH 7.4 showed a T 70% value of approximately 1.6 days at 37°C. Corresponding experiments at room temperature (ab...

Embodiment 2

[0138] Example 2 - Stabilization of bovine thrombin

[0139] The stabilization of bovine thrombin was studied. The experimental procedure was the same as in Example 1, but the concentration of bovine thrombin (Baxter) used was 0.4 mg / ml. The thrombin solution in HEPES buffer showed a T 70% value of 1.3 days when stored at 37°C. A solution of thrombin in HEPES buffer plus 3.0 mM N-(2'-phenoxy)-4-aminopyridine and 0.20 M MOPS showed a T70% value of 54 days.

[0140] The results obtained show that bovine thrombin is somewhat less stable than the human thrombin preparations studied, but that a very good stabilization is obtained by the combination according to the invention.

Embodiment 3

[0141] Example 3 - Low concentration of thrombin

[0142] The stabilization of thrombin in compositions comprising low concentrations of thrombin was investigated. The stabilizing effect of the combination according to the invention has also been shown to be usable at relatively low concentrations of thrombin.

[0143] A solution of 15.0 activity units / ml of human thrombin (from plasma, 3300 units / mg, Biovitrum AB, Sweden) in HEPES buffer, pH 7.4 showed a T 70% value of 23 days. The corresponding solution in HEPES buffer, pH 7.4 plus 2.0 mM N-(2'-phenoxy)-4-aminopyridine and 0.20 M MOPS showed a T 70% value of 92 days.

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Abstract

A composition is provided, which comprises a serine protease; a reversible inhibitor of said serine protease; and a stabilizing agent M having the formula (I): Also provided are uses of the composition as a medicament, and other uses and methods employing its various properties.

Description

field of invention [0001] The present invention relates to an enzyme composition wherein the enzyme is stabilized by certain additives in the composition of the invention. More specifically, the present invention relates to a serine protease composition comprising a reversible inhibitor of a serine protease and an additional stabilizer M as defined below. Background technique [0002] Serine proteases are a class of proteolytic enzymes characterized by having serine and histidine residues in their active sites. Many well-known enzymes belong to this category, such as trypsin, kallikrein, thrombin and plasmin. Practical applications of some of them have already been found. Trypsin is used in the leather industry. Thrombin is used as a hemostat to stop bleeding from wounds. Two other serine proteases, urokinase and tissue plasminogen activator, are clinically used as thrombolytic agents for the treatment of acute myocardial infarction. Many such enzymes are already widely...

Claims

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Application Information

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IPC IPC(8): A61K31/5377A61K45/06
Inventor L-O·安德松H·阿格兰
Owner TROBIO
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