# US 11,987,601 B2 · Norovirus fusion proteins and VLPs comprising norovirus fusion proteins

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- Number: US 11,987,601 B2
- Document ID: US11987601B2
- Status: Granted
- Kind: B2
- Application number: 17/735,706
- Filed: 2022-05-03
- Published: 2023-01-26
- Granted: 2024-05-21
- Earliest claimed: 2017-03-23
- Assignees: Aramis Biotechnologies Inc.
- Applicants: Aramis Biotechnologies Inc.
- Inventors: Pierre-Olivier Lavoie; Marc-Andre D'Aoust
- Web page: https://patentgrep.com/patent/US11987601B2

## CPC

- C12N 15/8257: {for the production of primary gene products, e.g. pharmaceutical products, interferon}
- A61K 39/12: Viral antigens
- A61P 1/00: Drugs for disorders of the alimentary tract or the digestive system
- A61P 31/14: for RNA viruses
- C07K 14/005: from viruses
- C07K 16/10: RNA viruses
- C12N 7/00: Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00)
- C12N 15/8258: {for the production of oral vaccines (antigens) or immunoglobulins}
- A61K 9/0019: {Injectable compositions; Intramuscular, intravenous, arterial, subcutaneous administration; Compositions to be administered through the skin in an invasive manner (non-active ingredients are additionally classified in A61K47/00)}
- A61K 2039/5258: {Virus-like particles}
- C07K 2319/00: Fusion polypeptide
- C12N 2770/16022: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2770/16023: Virus like particles [VLP]
- C12N 2770/16034: Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
- C12N 2770/16051: Methods of production or purification of viral material
- C12N 2770/16071: Demonstrated in vivo effect
- 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)

## Abstract

Nucleic acids encoding norovirus VP1 fusion proteins and VLPs comprising the norovirus VP1 fusion proteins are provided. Methods for norovirus VP1 fusion protein and norovirus VLP production in plants are also described. The VP1 fusion protein comprises, a first sequence encoding an S domain derived from a first norovirus strain, and a second sequence encoding a P domain derived from a second norovirus strain.

## Claims

### Claim 1

A method of producing a norovirus VP1 fusion protein in a plant, a portion of the plant, or a plant cell comprising,
introducing into the plant, the portion of the plant, or the plant cell a nucleic acid comprising a first sequence encoding an S domain derived from a first norovirus strain and a second sequence encoding a P domain derived from a second norovirus strain, the first norovirus strain selected from norovirus genotypes GI.1, GI.5, GII.1, GII.12, GII.14, and GII.21, and the second norovirus strain selected from norovirus genogroups GI, and GIV or selected from norovirus genotypes GII.1, GII.2, GII.3, GII.4, GII.5, GII.7, GII.12, GII.13, GII.14, GII.17 and GII.21, and
incubating the plant, the portion of the plant, or the plant cell under conditions that permit the expression and production of the norovirus VP1 fusion protein.

### Claim 2 (depends on 1)

The method of claim 1, wherein the second norovirus strain is selected from norovirus genotypes GI.1, GI.2, GI.3, GI.4, GI.5, GII.1, GII.2, GII.3, GII.4, GII.5, GII.7, GII.12, GII.13, GII.14, GII.17 and GII.21.

### Claim 3 (depends on 1)

The method of claim 1, wherein the first norovirus strain and the second norovirus strain are selected from norovirus subtypes GI.1/US/Norwalk/1968; GI.2/Leuven/2003/Bel; GI.3/S29/2008/Lilla Edet/Sweden; GII.5/AlbertaEI390/2013/CA; GII.1/Ascension208/2010/USA; GII.12/H5206/2010/USA; GII.13/VA173/2010/USA; GII.14/8610/Saga/2008/JPN; and GII.21/Salisbury150/2011/USA.

### Claim 4 (depends on 1)

The method of claim 1, wherein the nucleic acid further comprises a third sequence encoding a Cowpea Mosaic Virus (CPMV) enhancer, the CPMV enhancer operatively linked with the first and second sequences.

### Claim 5 (depends on 1)

The method of claim 1, wherein the first, the second, or both the first and second, sequence is optimized for human codon usage, increased GC content, or a combination thereof.

### Claim 6 (depends on 1)

The method of claim 1, wherein the method further comprises a step of harvesting the plant, the portion of the plant, or the plant cell.

### Claim 7 (depends on 6)

The method of claim 6, wherein the method further comprises a step of extracting, purifying, or both extracting and purifying, the norovirus VP1 fusion protein from the plant, the portion of the plant, or the plant cell.

### Claim 8 (depends on 6)

The method of claim 6, wherein the method further comprises a step of extracting, purifying, or both extracting and purifying a virus like particle (VLP) from the plant, the portion of the plant, or the plant cell, wherein the VLP comprises the norovirus VP1 fusion protein and has a diameter of about 15 nm to 50 nm.

### Claim 9 (depends on 8)

The method of claim 8, wherein the VLP has a diameter of either about 23 nm or about 38 nm.

### Claim 10 (depends on 1)

The method of claim 1, wherein in the step of introducing, a second nucleic acid sequence encoding a norovirus VP2 protein is introduced in the plant, the portion of the plant, or the plant cell, and in the step of incubating, the conditions permit co-expression and co-production, of both the norovirus VP1 fusion protein and the norovirus VP2 protein in the plant, the portion of the plant, or the plant cell.

### Claim 11 (depends on 10)

The method of claim 10, further comprising a step of harvesting the plant, the portion of the plant, or the plant cell.

### Claim 12 (depends on 11)

The method of claim 11, further comprising a step of extracting,
purifying, or both extracting and purifying, the norovirus VP1 fusion protein and norovirus VP2 protein.

### Claim 13 (depends on 11)

The method of claim 11, further comprising a step of extracting, purifying, or both extracting and purifying a virus like particle (VLP) from the plant, the portion of the plant, or the plant cell, wherein the VLP comprises the norovirus VP1 fusion protein and the norovirus VP2 protein, and has a diameter of about 15 nm to 50 nm.

### Claim 14 (depends on 13)

The method of claim 13, wherein the VLP has a diameter of either about 23 nm or about 38 nm.

### Claim 15 (depends on 1)

A plant, a portion of the plant or plant cell comprising the norovirus VP1 fusion protein produced by the method of claim 1.

### Claim 16 (depends on 1)

A plant, a portion of the plant or plant cell comprising a VLP comprising the VP1 fusion protein produced by the method of claim 1.

### Claim 17 (depends on 10)

A plant, a portion of the plant or plant cell comprising a VLP comprising the VP1 fusion protein and the VP2 protein produced by the method of claim 10.

### Claim 18 (depends on 1)

A plant extract comprising the norovirus VP1 fusion protein produced by the method of claim 1.

### Claim 19 (depends on 1)

A plant extract comprising a VLP comprising the VP1 fusion protein produced by the method of claim 1.

### Claim 20 (depends on 10)

A plant extract comprising a VLP comprising the VP1 fusion protein and the VP2 protein produced by the method of claim 10.