# US 11,993,783 B1 · Nucleic acid molecule comprising asymmetrically modified ITR for improving expression rate of inserted gene, and use thereof

> Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt

- Number: US 11,993,783 B1
- Document ID: US11993783B1
- Status: Granted
- Kind: B1
- Application number: 18/190,590
- Filed: 2023-03-27
- Granted: 2024-05-28
- Earliest claimed: 2023-03-27
- Assignees: Genecraft Inc.
- Applicants: Genecraft Inc.
- Inventors: Suk Chul Bae; You Soub Lee; Xinzi Chi; Seo Yeong Yoo; Woo-Jin Kim
- Web page: https://patentgrep.com/patent/US11993783B1

## CPC

- C12N 15/86: Viral vectors
- C12N 15/90: Stable introduction of foreign DNA into chromosome
- C12N 2750/14122: New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
- C12N 2750/14143: viral genome or elements thereof as genetic vector
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)

## Abstract

Described herein is a nucleic acid molecule including an asymmetrically modified inverted terminal repeat (ITR). An AAV vector including the nucleic acid molecule has advantages of increased productivity and expression efficiency of a transgene, and decreased genotoxicity, by having an asymmetric ITR in which any one of two ITRs is modified. Also, described herein is are compositions and vectors.

## Claims

### Claim 1

A nucleic acid molecule comprising:
a gene expression cassette between a first inverted terminal repeat (ITR) and a second ITR,
wherein the gene expression cassette comprises a heterologous polynucleotide sequence,
wherein:
the first ITR is an adeno-associated virus (AAV) wild-type TTR, and
the second ITR consists essentially of a nucleotide sequence or a complementary sequence thereof, the nucleotide sequence of the second ITR having at least about 90% sequence identity with a nucleotide sequence of any one of SEQ ID NOs: 1 to 9,
wherein the second ITR comprises a terminal resolution site (trs) sequence and a rep-binding element (RBE) sequence,
wherein in the second ITR, all of B, B′, C, C′, and RBE′ regions are deleted, and either A or A′ region, whichever is closer to 3′-end of the (+) strand of the transgene or 5′-end of the (−) strand of the transgene, is deleted,
wherein the second ITR exists as an open-end without forming any of the stem region and the loop region of the stem-loop structure, and
wherein the second ITR retains functional activities of the trs and the RBE.

### Claim 2 (depends on 1)

The nucleic acid molecule of claim 1, wherein the first ITR sequence and the second ITR sequence are each independently based on an ITR sequence of an AAV serotype selected from AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, and AAV12.

### Claim 3 (depends on 1)

The nucleic acid molecule of claim 1, wherein
the first ITR is an AAV wild-type ITR, and
the second ITR consists essentially of a nucleotide sequence or a complementary sequence thereof, the nucleotide sequence of the second ITR having 100% sequence identity with a nucleotide sequence of any one of SEQ ID NOs: 1 to 9 or a functional derivative thereof.

### Claim 4 (depends on 1)

The nucleic acid molecule of claim 1, wherein
the first ITR is an AAV wild-type ITR, and
the second ITR consists essentially of any one sequence selected from nucleotide sequences of SEQ ID NOs: 1 to 9.

### Claim 5 (depends on 1)

The nucleic acid molecule of claim 1, wherein
the first ITR is an AAV wild-type ITR, and
the second ITR consists essentially of a nucleotide sequence of SEQ ID NO: 1.

### Claim 6 (depends on 1)

The nucleic acid molecule of claim 1, wherein the gene expression cassette further comprises at least one of a promoter, and a polyadenylation sequence.

### Claim 7 (depends on 1)

The nucleic acid molecule of claim 1, wherein the heterologous polynucleotide sequence encodes a therapeutic gene.

### Claim 8 (depends on 1)

A vector comprising the nucleic acid molecule of claim 1.

### Claim 9 (depends on 8)

The vector of claim 8, wherein the vector is an AAV vector.

### Claim 10 (depends on 8)

A composition comprising: the vector of claim 8; and a pharmaceutically acceptable carrier.

### Claim 11 (depends on 10)

The composition of claim 10, wherein the composition is for delivering a therapeutic gene for gene therapy.