Polymer engineered regenerating bioscavengers

• US 10,400,232 B2
• Filed: 10/03/2014
• Issued: 09/03/2019
• Est. Priority Date: 10/03/2013
• Status: Active Grant

• Current Assignee: Carnegie Mellon University
• Original Assignee: Carnegie Mellon University
• Inventors: Russell, Alan J., Murata, Hironobu, Cummings, Chad, Koepsel, Richard R.
• Primary Examiner(s): Underdahl, Thane
• Application Number:
  US15/026,093
• Publication Number:
  US 20160244741A1
• Time in Patent Office: 1,796 Days
• Field of Search: None
• CPC Class Codes: A61K 38/465 acting on ester bonds (3.1), e.g. lipases, ribonucleases
• CPC Class Codes: A61K 38/4826 Trypsin (3.4.21.4) Chymotrypsin (3.4.21.1)
• CPC Class Codes: A61K 47/58 obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. poly[meth]acrylate, polyacrylamide, polystyrene, polyvinylpyrrolidone, polyvinylalcohol or polystyrene sulfonic acid resin
• CPC Class Codes: C12N 11/08 the carrier being a synthetic polymer
• CPC Class Codes: C12N 9/16 acting on ester bonds (3.1)
• CPC Class Codes: C12N 9/6427 Chymotrypsins (3.4.21.1; 3.4.21.2); Trypsin (3.4.21.4)
• CPC Class Codes: C12N 9/96 Stabilising an enzyme by forming an adduct or a composition; Forming enzyme conjugates
• CPC Class Codes: C12Y 301/01007 Acetylcholinesterase (3.1.1.7)
• CPC Class Codes: C12Y 304/21001 Chymotrypsin (3.4.21.1)

Abstract

Embodiments of the invention provide at least one polymer covalently conjugated to an esterase. The at least one polymer includes a plurality of oxime functional groups.

Claims

1. A composition comprising:

at least one polymer covalently conjugated to an enzyme, the at least one polymer comprising a reactive functional group, wherein the reactive functional group reacts with an inhibitor attached to an active site of the enzyme.

2. The composition of claim 1, wherein the enzyme comprises an esterase, and the reactive functional group comprises a plurality of oxime functional groups.

3. The composition of claim 2, wherein the esterase is selected from the group consisting of acetylcholinesterase and butyrylcholinesterase.

4. The composition of claim 2, wherein the esterase comprises chymotrypsin.

5. The composition of claim 2, wherein the plurality of oxime functional groups comprises alkyne derivatives of 2-pyridine aldoxime.

6. The composition of claim 2, wherein the plurality of oxime functional groups comprises an aldoxime.

7. The composition of claim 2, wherein the plurality of oxime functional groups comprises a ketoxime.

8. The composition of claim 1, wherein the reactive functional group reacts with the inhibitor attached to the active site of the enzyme, thus releasing the inhibitor from the active site and restoring activity to the active site of the enzyme.

9. The composition of claim 1, wherein the at least one polymer comprises at least one environmentally responsive monomer.

10. The composition of claim 1, wherein the at least one polymer is a copolymer comprising at least two different monomers, wherein at least one monomer is selected from the group consisting of aldoximes, ketoximes, muco-adhesion monomers, polyethylene glycol, bis-pyridinium oximes, N,N-dimethylacrylamide, N-isopropylacrylamide, (meth)acrylate, N,N-dimethylaminoethyl methacrylate, carboxyl acrylamide, 2-hydroxylethylmethacrylate, N-(2-hydroxypropyl)methacrylamide, quaternary ammonium monomers, sulfobetain methacrylate, oligo(ethylene glycol) methyl ether methacrylate, 2-PAM monomers, 4-PAM monomers, Clickable azide monomers, and combinations thereof.

11. A composition comprising:

a bioconjugate comprising;

an enzyme; and

at least one polymer covalently conjugated to the enzyme, wherein the at least one polymer comprises a plurality of reactive functional groups, and at least one reactive functional group of the plurality of reactive functional groups is positioned to react, in use, with a functional group on an inhibitor attached to an active site of the enzyme.

12. The composition of claim 11, wherein the enzyme comprises an esterase, and the at least one reactive functional group comprises a plurality of oxime functional groups.

13. The composition of claim 12, wherein the at least one reactive functional group comprises an oxime functional group positioned to exert a nucleophilic attack on a phosphoryl functional group of the inhibitor when the phosphoryl functional group is attached to the active site of the esterase, said nucleophilic attack resulting in removal of the phosphoryl functional group from the active site and restoration of activity to the esterase.

14. The composition of claim 12, wherein the at least one polymer of the bioconjugate comprises a flexibility sufficient to react, in use, with a phosphoryl functional group of the inhibitor attached to the active site of the esterase.

15. The composition of claim 12, wherein the at least one polymer covalently conjugated to the esterase forms a long lived covalent conjugate that is maintained in the body for a time period ranging from more than about one day to more than about one week.

16. The composition of claim 12, wherein the at least one polymer is a co-polymer comprising at least two different monomers, wherein at least one monomer comprises a member selected from the group consisting of aldoximes, ketoximes, muco-adhesion monomers, polyethylene glycol, bis-pyridinium oximes, N,N-dimethylacrylamide, N-isopropylacrylamide, (meth)acrylate, N,N-dimethylaminoethyl methacrylate, carboxyl acrylamide, 2-hydroxylethylmethacrylate, N-(2-hydroxypropyl)methacrylamide, quaternary ammonium monomers, sulfobetain methacrylate, oligo(ethylene glycol) methyl ether methacrylate, 2-PAM monomers, 4-PAM monomers, Clickable azide monomers, and combinations thereof.