# US 11,028,131 B2 · Mutant of adeno-associated virus (AAV) capsid protein

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- Number: US 11,028,131 B2
- Document ID: US11028131B2
- Status: Granted
- Kind: B2
- Application number: 16/481,159
- Filed: 2018-01-29
- Published: 2020-01-02
- Granted: 2021-06-08
- Earliest claimed: 2017-01-30
- Assignees: Nippon Medical School Foundation; National Center of Neurology and Psychiatry; Takara Bio Inc.
- Applicants: Nippon Medical School Foundation; National Center of Neurology and Psychiatry; Takara Bio Inc.
- Inventors: Takashi Okada; Hironori Okada; Hiromi Kinoh; Tatsuji Enoki; Toshikazu Nishie; Junichi Mineno
- Web page: https://patentgrep.com/patent/US11028131B2

## CPC

- C07K 14/005: from 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 15/85: for animal cells
- C12N 2750/14121: Viruses as such, e.g. new isolates, mutants or their genomic sequences
- C12N 2750/14122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2750/14123: Virus like particles [VLP]
- C12N 2750/14143: viral genome or elements thereof as genetic vector
- C12N 2750/14151: Methods of production or purification of viral material
- 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)

## Abstract

The present invention provides: a mutant of adeno-associated virus (AAV) capsid protein, which contains at least one amino acid substitution in PLA2 domain when compared with the amino acid sequence for wild-type AAV capsid protein; a nucleic acid encoding the mutant; a cell containing the nucleic acid; a method for producing a recombinant AAV particle, comprising a step of culturing the cell to produce the recombinant AAV particle; a recombinant AAV particle containing the mutant; a composition containing the recombinant AAV particle; and a method for transferring a gene into a target cell, comprising a step of bringing the recombinant AAV particle into contact with the target cell.

## Claims

### Claim 1

A mutant of an adeno-associated virus (AAV) capsid protein, comprising one or more amino acid replacements in a PLA2 domain as compared with the amino acid sequence of a wild-type AAV capsid protein,
wherein the one or more amino acid replacements comprise an amino acid replacement of alanine at position 68 in the amino acid sequence of AAV2 VP1 capsid protein, and said position is based on the amino acid sequence of wild-type AAV2 VP1 capsid protein shown by SEQ ID NO: 2, and
wherein the mutant is a mutant of AAV2 capsid protein.

### Claim 2 (depends on 1)

The mutant of an AAV capsid protein according to claim 1, wherein the one or more amino acid replacements comprise a replacement of alanine at position 68 by valine (A68V), in the amino acid sequence of AAV2 VP1 capsid protein, and said position is based on the amino acid sequence of wild-type AAV2 VP1 capsid protein shown by SEQ ID NO: 2.

### Claim 3 (depends on 1)

The mutant of an AAV capsid protein according to claim 1, wherein the one or more amino acid replacements comprise:
(a) a replacement of alanine at position 3 by threonine (A3T),
(b) a replacement of tyrosine at position 6 by histidine (Y6H), and
(c) a replacement of alanine at position 68 by valine (A68V)
in the amino acid sequence of AAV2 VP1 capsid protein, and the positions indicated in (a) to (c) are based on the amino acid sequence of wild-type AAV2 VP1 capsid protein shown by SEQ ID NO: 2.

### Claim 4 (depends on 1)

A recombinant AAV particle containing the mutant of an AAV capsid protein according to claim 1.

### Claim 5 (depends on 4)

A composition containing the recombinant AAV particle according to claim 4.

### Claim 6 (depends on 4)

A method of introducing a gene into a target cell, the method comprising a step of bringing the recombinant AAV particle according to claim 4 into contact with a target cell.