# US 10,370,660 B2 · Antisense oligonucleotides to treat Dystrophic Epidermolysis Bullosa

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- Number: US 10,370,660 B2
- Document ID: US10370660B2
- Status: Granted
- Kind: B2
- Application number: 15/575,559
- Filed: 2016-05-20
- Published: 2018-08-30
- Granted: 2019-08-06
- Earliest claimed: 2015-05-21
- Assignees: Wings Therapeutics, Inc.
- Applicants: ProQR Therapeutics II B.V.
- Inventors: Elisabeth Marlene Haisma; Marko Potman; Gerardus Johannes Platenburg
- Web page: https://patentgrep.com/patent/US10370660B2

## 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 17/00: Drugs for dermatological disorders
- C12N 2310/11: Antisense
- C12N 2310/321: 2'-O-R Modification
- C12N 2320/32: Special delivery means, e.g. tissue-specific
- 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

An antisense oligonucleotide capable of preventing or reducing exon 80 inclusion into a human COL7A1 mRNA, and methods for preventing or reducing exon 80 inclusion into a human COL7A1 mRNA.

## Claims

### Claim 1

An antisense oligonucleotide (AON) capable of preventing or reducing exon 80 inclusion into a human COL7A1 mRNA when the mRNA is produced by splicing from a pre-mRNA in a cell, wherein the AON comprises a nucleotide sequence selected from SEQ ID NOs: 8, 25, 26, 31 and 32.

### Claim 2 (depends on 1)

The AON of claim 1, wherein the AON comprises a region of complementarity with exon 80 wherein the region of complementarity is at most 20 nucleotides in length.

### Claim 3 (depends on 1)

The AON of claim 1, wherein the AON is less than 24 nucleotides in length.

### Claim 4 (depends on 1)

The AON of claim 1, wherein the AON is an oligoribonucleotide.

### Claim 5 (depends on 1)

The AON of claim 1, wherein the internucleosidic linkages are chemically modified.

### Claim 6 (depends on 1)

The AON of claim 1, wherein the AON comprises one or more lower 2′-O-alkyl substituted sugar moieties.

### Claim 7 (depends on 1)

A method for preventing or reducing exon 80 inclusion into a human COL7A1 mRNA when the mRNA is produced by splicing from an RNA transcript in a human cell, the method comprising providing to (i) the cell, in vitro or ex vivo, (ii) a tissue comprising the cell, in vitro or ex vivo, or (iii) a living human being comprising the cell, the oligonucleotide of claim 1, under conditions conducive to uptake of the oligonucleotide by the cell, and allowing splicing to take place.

### Claim 8 (depends on 2)

The AON of claim 2, wherein the AON comprises a region of complementarity with exon 80 that is 11, 12, 13, 14, 15, 16 or 17 nucleotides in length.

### Claim 9 (depends on 3)

The AON of claim 3, wherein the AON is 20, 21, 22, or 23 nucleotides in length.

### Claim 10 (depends on 5)

The AON of claim 5, wherein the internucleosidic linkages are phosphorothioate-linkages.

### Claim 11 (depends on 6)

The AON of claim 6, wherein the lower 2′-O-alkyl substituted sugar moieties are 2′-O-methyl substituted sugar moieties.

### Claim 12 (depends on 1)

A composition comprising the oligonucleotide of claim 1, and one or more of a carrier, excipient, stabilizer, transfection agent, diluent, gelling agent or buffer.

### Claim 13 (depends on 1)

The AON of claim 1, wherein the AON consists of a nucleotide sequence selected from SEQ ID NOs: 8, 25, 26, 31 and 32.

### Claim 14 (depends on 1)

The AON of claim 1, wherein the AON comprises one or more sugar moieties that comprise a substitution at one or two of the 2′, 3′ and 5′ positions.

### Claim 15 (depends on 14)

The AON of claim 14, wherein the substitution is selected from: -OH; -F; substituted or unsubstituted, linear or branched lower (C1-C10) alkyl, alkenyl, alkynyl, alkaryl, allyl, or aralkyl, that may be interrupted by one or more heteroatoms; O-, S-, or N-alkyl; O-, S-, or N-alkenyl; O-, S- or N-alkynyl; O-, S-, or N-allyl; O-alkyl-O-alkyl, -methoxy, -aminopropoxy; -methoxyethoxy; -dimethylaminooxyethoxy; and -dimethylaminoethoxyethoxy.

### Claim 16 (depends on 1)

The AON of claim 1, wherein the AON comprises the nucleotide sequence of SEQ ID NO: 8.

### Claim 17 (depends on 1)

The AON of claim 1, wherein the AON comprises the nucleotide sequence of SEQ ID NO: 25.

### Claim 18 (depends on 1)

The AON of claim 1, wherein the AON comprises the nucleotide sequence of SEQ ID NO: 26.

### Claim 19 (depends on 1)

The AON of claim 1, wherein the AON comprises the nucleotide sequence of SEQ ID NO: 31.

### Claim 20 (depends on 1)

The AON of claim 1, wherein the AON comprises the nucleotide sequence of SEQ ID NO: 32.