# US 9,518,259 B2 · Compounds and methods for modulating interaction between proteins and target nucleic acids

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- Number: US 9,518,259 B2
- Document ID: US9518259B2
- Status: Granted
- Kind: B2
- Application number: 13/703,322
- Filed: 2011-06-15
- Published: 2013-10-31
- Granted: 2016-12-13
- Earliest claimed: 2010-06-15
- Assignees: Ionis Pharmaceuticals, Inc.
- Applicants: Frank Rigo; C. Frank Bennett; Adrian R. Krainer; Yimin Hua
- Inventors: Frank Rigo; C. Frank Bennett; Adrian R. Krainer; Yimin Hua
- Web page: https://patentgrep.com/patent/US9518259B2

## CPC

- C12N 15/113: Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; {Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing (when used in plants C12N15/8218)}
- A61P 21/00: Drugs for disorders of the muscular or neuromuscular system
- A61P 35/00: Antineoplastic agents
- C12N 15/111: {General methods applicable to biologically active non-coding nucleic acids}
- C12N 2310/11: Antisense
- C12N 2310/321: 2'-O-R Modification
- C12N 2310/322: 2'-R Modification
- C12N 2310/3231: having an additional ring, e.g. LNA, ENA
- C12N 2310/3341: 5-Methylcytosine
- C12N 2310/341: Gapmers, i.e. of the type ===---===
- C12N 2310/3525: MOE, methoxyethoxy
- C12N 2310/3533: Halogen
- C12N 2320/33: Alteration of splicing
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- A61P: SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS

## Abstract

Provided herein are antisense compounds and methods for recruiting one or more non-cleaving protein to a target nucleic acid in a cell. In certain instances such recruitment of a non-cleaving protein alters the function or activity of the target nucleic acid. In certain such instances, the target nucleic acid a pre-mRNA and the recruitment of the non-cleaving protein results in a change in splicing of the pre-mRNA.

## Claims

### Claim 1

A method of inducing exon skipping in a target pre-mRNA in a cell comprising contacting the cell with an antisense compound comprising a chemically modified oligonucleotide consisting of 18 linked nucleosides; and thereby inducing exon skipping in the pre-mRNA in the cell; wherein the antisense compound comprises:
a 5′-wing region consisting of 2 linked duplex stabilizing nucleosides;
a 3′-wing region consisting of 2 linked duplex stabilizing nucleosides; and
a central gap region located between the 5′-wing region and the 3′-wing region
and consisting of 14 contiguous nucleosides comprising 2′-F modifications;
wherein each duplex stabilizing nucleoside is a 2′-methoxyethyl (2′-MOE) nucleoside, and
wherein the antisense compound is complementary to an intron of the target pre-mRNA.

### Claim 2 (depends on 1)

The method of claim 1, wherein the antisense compound binds to the target pre-mRNA and recruits at least one non-cleaving nucleic acid binding protein to the antisense compound/target pre-mRNA duplex.

### Claim 3 (depends on 2)

The method of claim 2, wherein at least one of the at least one non-cleaving nucleic acid binding proteins recruited to the compound/target pre-mRNA duplex is Interleukin Enhancer Binding Factor 2 or Interleukin Enhancer Binding Factor 3.

### Claim 4 (depends on 1)

The method of claim 1, wherein the target pre-mRNA is associated with a disease or disorder.

### Claim 5 (depends on 4)

The method of claim 4, wherein the target pre-mRNA is Bcl-x.

### Claim 6 (depends on 4)

The method of claim 4, wherein the disease or disorder is cancer.

### Claim 7 (depends on 4)

The method of claim 4, wherein the disease or disorder is Duchenne muscular dystrophy.

### Claim 8 (depends on 1)

The method of claim 1, wherein the cell is in vitro.

### Claim 9 (depends on 1)

The method of claim 1, wherein the cell is in an animal.

### Claim 10 (depends on 1)

The method of claim 1 comprising performing an assay to determine whether the exon has been skipped.

### Claim 11 (depends on 1)

The method of claim 1 comprising performing an assay to determine whether a protein has been recruited to the antisense compound/target pre-mRNA duplex.

### Claim 12 (depends on 11)

The method of claim 11 comprising performing an assay to determine the identity of one or more protein recruited to the antisense compound/target pre-mRNA duplex.

### Claim 13 (depends on 6)

The method of claim 6, wherein the cell is in an animal.

### Claim 14 (depends on 7)

The method of claim 7, wherein the cell is in an animal.