Enzymatic preparation of increased anticoagulant bovine sourced heparin

Patent metadata

Field	Value
Publication number	US-10864227-B2
Application number	US-201816496149-A
Country	US
Kind code	B2
Filing date	Mar 20, 2018
Priority date	Mar 20, 2017
Publication date	Dec 15, 2020
Grant date	Dec 15, 2020

Abstract

A heparin structure with increased anticoagulant activity and method of making the same are disclosed. A heparin sample is provided and treated with a heparan sulfate sulfotransferase in an enzymatic reaction to add sulfuryl groups from a sulfuryl group source to the heparin sample, resulting in a heparin structure having above about 8% more 3-O-sulfo groups relative to wild-type bovine intestinal heparin. The added sulfuryl groups modify the heparin structure and increase the sample's binding to antithrombin III and its anticoagulant activity to be more similar and a viable alternative to porcine intestinal heparin. The modified heparin exhibits an anti-FXa activity and an anti-FIIa activity greater than about 180 U/mg, and a ratio of the anti-FXa activity to the anti-FIIa activity of about 0.9 to about 1.1, consistent with U.S. Pharmacopeia (USP) heparin activity specifications.

First claim

What is claimed is:

1. A heparin structure exhibiting increased anticoagulant activity, the heparin comprising the structure:

wherein:
a+d is about 14 to about 18;
b is 2;
c is about 2 to about 8;
X and X′ are SO 3 − , H, or combinations thereof;
Y and Y′ are SO 3 − , COCH3, H, or combinations thereof; and
Z is SO 3 − , H, or combinations thereof.

2. The heparin structure according to claim 1 , wherein X and X′ in b and c is about 4H or about 4 SO 3 − .

3. The heparin structure according to claim 1 , wherein Y and Y′ in b and c is about 6 SO 3 − , about 1 COCH 3 , and about 1H.

4. The heparin structure according to claim 1 , wherein Z is about 0.8 SO 3 − and about 0.2H.

5. The heparin structure according to claim 1 , wherein the heparin structure includes above about 8% more 3-O-sulfo groups relative to wild-type bovine intestinal heparin.

6. The heparin structure according to claim 5 , wherein the heparin structure includes about 15% to about 25% more 6-O-sulfo groups and about 8% to about 20% more 3-O-sulfo groups relative to wild-type bovine intestinal heparin.

7. The heparin structure according to claim 1 , wherein the heparin structure includes an anti-FXa activity and an anti-FIIa activity greater than about 180 U/mg, and the ratio of the anti-FXa activity to the anti-FIIa activity is about 0.9 to about 1.1.

Assignees

• Rensselaer Polytech Inst

Inventors

• Fu Li
• Li Kevin
• Mori Daisuke
• Hirakane Makoto
• Dordick Jonathan S
• Linhardt Robert J

Classifications

• C12N9/13

  transferring sulfur containing groups (2.8) · CPC title

• C12Y208/02

  Sulfotransferases (2.8.2) · CPC title

• C12P19/26

  Preparation of nitrogen-containing carbohydrates · CPC title

• A61K31/727 Primary

  Heparin; Heparan · CPC title