# US 10,501,729 B2 · Engineered heme-binding compositions and uses thereof

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- Number: US 10,501,729 B2
- Document ID: US10501729B2
- Status: Granted
- Kind: B2
- Application number: 15/968,116
- Filed: 2018-05-01
- Published: 2018-11-08
- Granted: 2019-12-10
- Earliest claimed: 2013-05-21
- Assignees: President and Fellows of Harvard College
- Applicants: President and Fellows of Harvard College
- Inventors: Michael Super; Alexander L. Watters; Philip T. Snell; Donald E. Ingber
- Web page: https://patentgrep.com/patent/US10501729B2

## CPC

- C12N 9/2474: {Hyaluronoglucosaminidase (3.2.1.35), i.e. hyaluronidase}
- A61K 38/00: Medicinal preparations containing peptides (peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00)
- A61P 7/00: Drugs for disorders of the blood or the extracellular fluid
- A61P 7/08: Plasma substitutes; Perfusion solutions; Dialytics or haemodialytics; Drugs for electrolytic or acid-base disorders, e.g. hypovolemic shock (artificial tears A61P27/04)
- A61P 21/00: Drugs for disorders of the muscular or neuromuscular system
- A61P 31/00: Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P 31/04: Antibacterial agents
- C07K 14/805: Haemoglobins; Myoglobins
- C12N 9/96: Stabilising an enzyme by forming an adduct or a composition; Forming enzyme conjugates
- C12Y 302/01035: Hyaluronoglucosaminidase (3.2.1.35), i.e. hyaluronidase
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- 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)
- C12Y: ENZYMES

## Abstract

Described herein are heme-binding compositions and methods relating to their use, for example methods of treatment of sepsis and rhabdomyolysis.

## Claims

### Claim 1

A composition comprising:
an engineered heme-binding fusion protein comprising:
a hemopexin domain comprising a polypeptide having a sequence corresponding to residues 27-213 of SEQ ID NO: 1 or SEQ ID NO: 2; and
a Fc domain comprising the polypeptide sequence of SEQ ID NO: 8, SEQ ID NO: 7, or SEQ ID NO: 17.

### Claim 2 (depends on 1)

The composition of claim 1, wherein the hemopexin domain binds to heme.

### Claim 3 (depends on 1)

The composition of claim 1, wherein the Fc domain is linked to the N-terminus or C-terminus of the hemopexin domain.

### Claim 4 (depends on 1)

The composition of claim 1, wherein the hemopexin domain is a polypeptide comprising the sequence of SEQ ID NO: 1.

### Claim 5 (depends on 1)

The composition of claim 1, wherein the hemopexin domain comprises a polypeptide having a sequence corresponding to:
residues 24-462 of SEQ ID NO: 1;
residues 24-256 of SEQ ID NO: 1;
residues 27-233 of SEQ ID NO: 1;
residues 1-233 of SEQ ID NO: 1;
residues 27-220 of SEQ ID NO: 1;
residues 1-220 of SEQ ID NO: 1;
residues 27-213 of SEQ ID NO: 1;
residues 1-213 of SEQ ID NO: 1;
residues 27-256 of SEQ ID NO: 1;
or residues 1-256 of SEQ ID NO: 1.

### Claim 6 (depends on 5)

The composition of claim 5, wherein the hemopexin domain further comprises a mutation wherein the residues corresponding to residues 220-226 of SEQ ID NO: 1 have been replaced with a polypeptide linker of about 1-10 amino acids in length.

### Claim 7 (depends on 5)

The composition of claim 5, wherein the hemopexin domain further comprises a mutation wherein the residues corresponding to residues 220-226 of SEQ ID NO: 1 have been replaced with the sequence GSGS (SEQ ID NO: 18).

### Claim 8 (depends on 1)

The composition of claim 1, wherein the hemopexin domain is a polypeptide comprising the sequence of SEQ ID NO: 2.

### Claim 9 (depends on 1)

The composition of claim 1, wherein the hemopexin domain comprises a polypeptide having a sequence corresponding to:
residues 27-233 of SEQ ID NO: 2;
residues 1-233 of SEQ ID NO: 2;
residues 27-220 of SEQ ID NO: 2;
residues 1-220 of SEQ ID NO: 2;
residues 27-213 of SEQ ID NO: 2;
residues 1-213 of SEQ ID NO: 2;
residues 27-256 of SEQ ID NO: 2;
or residues 1-256 of SEQ ID NO: 2.

### Claim 10 (depends on 9)

The composition of claim 9, wherein the hemopexin domain further comprises a mutation wherein the residues corresponding to residues 220-226 of SEQ ID NO: 2 have been replaced with a polypeptide linker of about 1-10 amino acids in length.

### Claim 11 (depends on 9)

The composition of claim 9, wherein the hemopexin domain further comprises a mutation wherein the residues corresponding to residues 220-226 of SEQ ID NO: 2 have been replaced with the sequence GSGS (SEQ ID NO: 18).

### Claim 12 (depends on 1)

The composition of claim 1, wherein the engineered heme-binding fusion protein comprises the sequence of SEQ ID NO: 4 or SEQ ID NO: 5.

### Claim 13 (depends on 1)

The composition of claim 1, wherein the second polypeptide domain multimerizes to form a multimeric complex of the engineered heme-binding fusion protein.

### Claim 14 (depends on 13)

The composition of claim 13, wherein the multimeric complex is a dimeric complex.

### Claim 15 (depends on 1)

The composition of claim 1, wherein the second polypeptide domain is linked to the N-terminus or C-terminus of the hemopexin domain via a linker.

### Claim 16 (depends on 1)

The composition of claim 1, wherein the second polypeptide domain is linked to the N-terminus of the hemopexin domain.

### Claim 17 (depends on 1)

The composition of claim 1, wherein the second polypeptide domain is linked to the C-terminus of the hemopexin domain.

### Claim 18 (depends on 1)

A pharmaceutical composition comprising the composition of claim 1 and a pharmaceutically acceptable carrier.