# US 10,370,432 B2 · Heterologous targeting peptide grafted AAVS

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- Number: US 10,370,432 B2
- Document ID: US10370432B2
- Status: Granted
- Kind: B2
- Application number: 15/516,582
- Filed: 2015-10-02
- Published: 2018-09-20
- Granted: 2019-08-06
- Earliest claimed: 2014-10-03
- Assignees: University of Massachusetts
- Applicants: University of Massachusetts
- Inventors: Miguel Sena Esteves; Sourav Roy Choudhury
- Web page: https://patentgrep.com/patent/US10370432B2

## CPC

- C12N 15/8645: {Adeno-associated virus}
- A61K 48/0025: {wherein the non-active part clearly interacts with the delivered nucleic acid}
- A61K 48/0058: {Nucleic acids adapted for tissue specific expression, e.g. having tissue specific promoters as part of a contruct}
- C07K 7/00: Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K 7/04: Linear peptides containing only normal peptide links
- C07K 14/005: from viruses
- C07K 14/775: Apolipopeptides
- C07K 16/00: Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies
- C12N 15/79: Vectors or expression systems specially adapted for eukaryotic hosts
- C12N 15/85: for animal cells
- C12N 15/86: Viral vectors
- C07K 14/00: Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K 14/075: Adenoviridae
- C07K 19/00: Hybrid peptides {, i.e. peptides covalently bound to nucleic acids, or non-covalently bound protein-protein complexes}
- C07K 2319/01: containing a localisation/targetting motif
- C07K 2319/30: Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto
- C07K 2319/33: fusions for targeting to specific cell types, e.g. tissue specific targeting, targeting of a bacterial subspecies
- C12N 2310/141: MicroRNAs, miRNAs
- C12N 2750/14145: Special targeting system for viral vectors
- C12N 2810/85: mammalian
- 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)
- 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

The disclosure in some aspects relates to recombinant adeno-associated viruses having distinct tissue targeting capabilities. In some aspects, the disclosure relates to gene transfer methods using the recombinant adeno-associated viruses. In some aspects, the disclosure relates to isolated AAV capsid proteins and isolated nucleic acids encoding the same.

## Claims

### Claim 1

An adeno associated virus capsid protein comprising an N-terminally grafted heterologous targeting peptide, wherein:
(i) the capsid protein is a VP2 capsid protein that is not of an AAV2 serotype; and
(ii) the targeting peptide is a CNS-targeting polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 5, that is inserted between the first and second amino acid residues of the VP2 capsid protein.

### Claim 2 (depends on 1)

The capsid protein of claim 1, further comprising a linker conjugated to the C-terminus of the N-terminally grafted heterologous targeting peptide.

### Claim 3 (depends on 2)

The capsid protein of claim 2, wherein the linker comprises at least one polypeptide repeat, each repeat comprising at least two glycine residues.

### Claim 4 (depends on 3)

The capsid protein of claim 3, wherein the linker is of the formula [G]nS, wherein n is an integer in a range of 2 to 10.

### Claim 5 (depends on 3)

The capsid protein of claim 3, wherein the linker comprises the formula [GGGGS]n , wherein n is an integer in a range of 1 to 4.

### Claim 6 (depends on 5)

The capsid protein of claim 5, wherein the linker comprises SEQ ID NO: 28.

### Claim 7

An adeno associated virus capsid protein comprising an N-terminally grafted heterologous targeting peptide, wherein:
(i) the capsid protein is a VP2 capsid protein that is not of an AAV2 serotype; and
(ii) the targeting peptide is a CNS-targeting polypeptide consisting of the amino acid sequence set forth in SEQ ID NO: 7, that is inserted between the first and second amino acid residues of the VP2 capsid protein.

### Claim 8 (depends on 7)

The capsid protein of claim 7, further comprising a linker conjugated to the C-terminus of the N-terminally grafted heterologous targeting peptide.

### Claim 9 (depends on 8)

The capsid protein of claim 8, wherein the linker comprises at least one polypeptide repeat, each repeat comprising at least two glycine residues.

### Claim 10 (depends on 9)

The capsid protein of claim 9, wherein the linker is of the formula [G]nS, wherein n is an integer in a range of 2 to 10.

### Claim 11 (depends on 9)

The capsid protein of claim 9, wherein the linker comprises the formula [GGGGS]n, wherein n is an integer in a range of 1 to 4.

### Claim 12 (depends on 11)

The capsid protein of claim 11, wherein the linker comprises SEQ ID NO:28.

### Claim 13

An adeno associated virus capsid protein comprising an N-terminally grafted heterologous targeting peptide, wherein:
(i) the capsid protein is a VP2 capsid protein that is not of an AAV2 serotype; and
(ii) the targeting peptide is a CNS-targeting polypeptide encoded by a nucleic acid consisting of the nucleotide sequence set forth in SEQ ID NO: 29, that is inserted between the first and second amino acid residues of the VP2 capsid protein.

### Claim 14 (depends on 13)

The capsid protein of claim 13, further comprising a linker conjugated to the C-terminus of the N-terminally grafted heterologous targeting peptide.

### Claim 15 (depends on 14)

The capsid protein of claim 14, wherein the linker comprises at least one polypeptide repeat, each repeat comprising at least two glycine residues.

### Claim 16 (depends on 15)

The capsid protein of claim 15, wherein the linker is of the formula [G]nS, wherein n is an integer in a range of 2 to 10.

### Claim 17 (depends on 15)

The capsid protein of claim 15, wherein the linker comprises the formula [GGGGS]n, wherein n is an integer in a range of 1 to 4.

### Claim 18 (depends on 17)

The capsid protein of claim 17, wherein the linker comprises SEQ ID NO:28.