# US 10,370,662 B2 · Replication capable rAAV vectors encoding inhibitory siRNA and methods of their use

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- Number: US 10,370,662 B2
- Document ID: US10370662B2
- Status: Granted
- Kind: B2
- Application number: 15/728,447
- Filed: 2017-10-09
- Published: 2018-02-01
- Granted: 2019-08-06
- Earliest claimed: 2013-02-28
- Assignees: City of Hope
- Applicants: City of Hope
- Inventors: Kamehameha K. Wong, Jr.; Cam Mroske; Saswati Chatterjee
- Web page: https://patentgrep.com/patent/US10370662B2

## CPC

- C12N 15/1131: {against viruses}
- C12N 2330/51: Specially adapted vectors
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

In some embodiments, an antiviral vector is provided. The antiviral vector includes a replication competent adeno-associated virus (AAV) and an inhibitory expression cassette that includes a nucleotide sequence that encodes an RNAi molecule that inhibits expression of a targeted helper virus (THV) gene. The THV gene may be part of an Adenovirus (Ad) genome, a Human Papillomavirus (HPV) genome, a Human Herpes Virus (HHV) genome, or a Vaccinia virus (VV) genome.

## Claims

### Claim 1

A pharmaceutical composition comprising:
(i) one or more antiviral vectors, each comprising
an inhibitory expression cassette; and
a replication competent adeno-associated virus (AAV); and
(ii) a pharmaceutically acceptable carrier,
wherein the inhibitory expression cassette comprises a nucleotide sequence that encodes an RNAi molecule that inhibits expression of a targeted helper virus (THV) gene, and
wherein the RNAi molecule is an shRNA comprising a sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:57, SEQ ID NO:58, and SEQ ID NO:59.

### Claim 2 (depends on 1)

The pharmaceutical composition of claim 1, wherein the inhibitory expression cassette further comprises an H6 or U6 promoter.

### Claim 3 (depends on 1)

The pharmaceutical composition of claim 1, wherein the THV gene is part of an Adenovirus (Ad) genome, a Human Papillomavirus (HPV) genome, a Human Herpes Virus (HHV) genome, or a Vaccinia virus (VV) genome.

### Claim 4 (depends on 3)

The pharmaceutical composition of claim 3, wherein the THV gene is HPV E6 or HPV E7.

### Claim 5 (depends on 3)

The pharmaceutical composition of claim 3, wherein the THV gene is Ad E1, Ad IVa, or Ad Hexon.

### Claim 6 (depends on 3)

The pharmaceutical composition of claim 3, wherein the RNAi molecule targets Ad Tripartite Leader Sequence (TriP).

### Claim 7 (depends on 1)

The pharmaceutical composition of claim 1, wherein the replication competent AAV virus is a wild type serotype-2 AAV.

### Claim 8

A method of producing an antiviral vector comprising:
culturing the antiviral vector with a population of cells infected with a first THV, such that the antiviral vector co-infects the population of cells; and
isolating the antiviral vector after at least one full infectious cycle;
wherein the antiviral vector comprises a replication competent AAV virus and a nucleotide sequence that encodes an RNAi molecule that inhibits expression of a targeted helper virus (THV) gene, and
wherein the RNAi molecule is an shRNA comprising a sequence selected from the group consisting of SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, SEQ ID NO:8, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, SEQ ID NO:19, SEQ ID NO:26, SEQ ID NO:27, SEQ ID NO:28, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:31, SEQ ID NO:32, SEQ ID NO:33, SEQ ID NO:57, SEQ ID NO:58, and SEQ ID NO:59.

### Claim 9 (depends on 8)

The method of claim 8, wherein the THV gene is part of an Adenovirus (Ad) genome, a Human Papillomavirus (HPV) genome, a Human Herpes Virus (HHV) genome, or a Vaccinia virus (VV) genome.

### Claim 10 (depends on 9)

The method of claim 9, wherein the THV gene is HPV E6 or HPV E7.

### Claim 11 (depends on 9)

The method of claim 9, wherein the THV gene is Ad E1, Ad IVa, or Ad Hexon.

### Claim 12 (depends on 9)

The method of claim 9, wherein the RNAi molecule targets Ad Tripartite Leader Sequence (TriP).

### Claim 13 (depends on 1)

The pharmaceutical composition of claim 1, comprising two or more antiviral vectors, wherein the inhibitory expression cassettes of the antiviral vectors comprise nucleotide sequences that encode RNAi molecules that inhibit expression of different THV genes.