# US 11,304,995 B2 · Prohemostatic proteins for the treatment of bleeding

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- Number: US 11,304,995 B2
- Document ID: US11304995B2
- Status: Granted
- Kind: B2
- Application number: 16/719,875
- Filed: 2019-12-18
- Published: 2020-05-07
- Granted: 2022-04-19
- Earliest claimed: 2014-05-26
- Assignees: Academisch Ziekenhuis Leiden
- Applicants: Academisch Ziekenhuis Leiden
- Inventors: Daniël Verhoef; Pieter H. Reitsma; Mettine H. A. Bos
- Web page: https://patentgrep.com/patent/US11304995B2

## CPC

- A61K 38/4846: {Factor VII (3.4.21.21); Factor IX (3.4.21.22); Factor Xa (3.4.21.6); Factor XI (3.4.21.27); Factor XII (3.4.21.38)}
- A61P 7/02: Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
- A61P 7/04: Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
- A61P 43/00: Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
- C07K 14/46: from vertebrates
- C12N 9/6432: {Coagulation factor Xa (3.4.21.6)}
- C12Y 304/21006: Coagulation factor Xa (3.4.21.6)
- C07K 2319/00: Fusion polypeptide
- A61K: PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES (devices or methods specially adapted for bringing pharmaceutical products into particular physical or administering forms A61J3/00; chemical aspects of, or use of materials for deodorisation of air, for disinfection or sterilisation, or for bandages, dressings, absorbent pads or surgical articles A61L; soap compositions C11D)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C12Y: ENZYMES

## Abstract

This disclosure relates to recombinant FXa polypeptides that can be used as antidotes to completely or partially reverse an anti-coagulant effect of a coagulation inhibitor in a subject, preferably a direct factor Xa inhibitor. Disclosed herein are recombinant factor Xa proteins and a method of completely or partially reversing an anti-coagulant effect of a coagulation inhibitor in a subject.

## Claims

### Claim 1

A recombinant protein comprising a coagulation factor Xa polypeptide, said coagulation factor Xa polypeptide having an alteration or deletion of an amino acid residue corresponding to amino acid residue Phe-396 as indicated in SEQ ID NO: 1.

### Claim 2 (depends on 1)

The protein according to claim 1, wherein the alteration or deletion of an amino acid residue corresponding to amino acid residue Phe-396 is combined with an insertion of 1-50 amino acid residue(s) in a region corresponding to the region of amino acid residues between Gly-289 and Asp-320.

### Claim 3 (depends on 1)

The protein according to claim 1, wherein the alteration or deletion of an amino acid residue corresponding to amino acid residue Phe-396 is combined with an insertion of 1-50 amino acid residue(s) in a region corresponding to the region of amino acid residues between His-311 and Asp-320 of SEQ ID NO: 1.

### Claim 4 (depends on 3)

The protein according to claim 3, wherein the region of amino acid residues corresponding to amino acid residues between His-311 and Asp-320 of SEQ ID NO: 1 has the amino acid sequence of SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 9, SEQ ID NO: 10, or SEQ ID NO: 11.

### Claim 5 (depends on 1)

A pharmaceutical composition comprising the protein of claim 1, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

### Claim 6 (depends on 1)

A method of completely or partially reversing an anti-coagulant effect of a direct factor Xa inhibitor in a subject in need thereof, the method comprising:
administering to the subject a therapeutically effective amount of the protein of claim 1 so as to completely or partially reverse the anti-coagulant effect of the direct factor Xa inhibitor in the subject.

### Claim 7 (depends on 6)

The method according to claim 6, wherein the direct factor Xa inhibitor is rivaroxaban (5-chloro-N-[[(5S)-2-oxo-3-[4-(3-oxo-4-morpholinyl)phenyl]-5-oxazolidinyl]methyl]-2-thiophenecarboxamide), apixaban (1-(4methoxyphenyl)-7-oxo-6-[4-(2-oxopiperidin-1-yl)phenyl]-4,5,6,7-tetrahydro-1H-pyrazolo[3,4-c]pyridine-3-carboxamide), edoxaban (N′-(5-chloropyridin-2-yl)-N-[(1S,2R,4S)-4-(dimethylcarbamoyl)-2-[(5-methyl-6,7-dihydro-4H-[1,3]thiazolo[5,4-c]pyridine-2-carbonyl)amino]cyclohexyl]oxamide; 4-methylbenzenesulfonic acid), or betrixaban (N-(5-chloropyridin-2-yl)-2-[[4-(N,N-dimethylcarbamimidoyl)benzoyl]amino]-5-methoxybenzamide).

### Claim 8 (depends on 1)

A method of making the protein of claim 1, the method comprising:
expressing a nucleic acid molecule comprising a DNA sequence that encodes the protein in a host cell.

### Claim 9 (depends on 8)

The method according to claim 8, wherein the nucleic acid molecule is comprised within an expression vector.