Double-stranded antisense nucleic acid with exon-skipping effect

Patent metadata

Field	Value
Publication number	US-2016130583-A1
Application number	US-201414898630-A
Country	US
Kind code	A1
Filing date	Jun 16, 2014
Priority date	Jun 16, 2013
Publication date	May 12, 2016
Grant date	—

Abstract

Disclosed are double-stranded antisense nucleic acid complexes that can efficiently alter the processing of RNA in a cell via an antisense effect, and methods for using the same. One method comprises contacting with the cell a double-stranded nucleic acid complex comprising: a first nucleic acid strand annealed to a second nucleic acid strand, wherein: the first nucleic acid strand comprises (i) nucleotides independently selected from natural DNA nucleotides, modified DNA nucleotides, and nucleotide analogs, (ii) no regions that have 4 or more consecutive natural DNA nucleotides, (iii) the total number of natural DNA nucleotides, modified DNA nucleotides, and nucleotide analogs in the first nucleic acid strand is from 8 to 100, and (iv) the first nucleic acid strand is capable of hybridizing to RNA inside of the cell; and the second nucleic acid strand comprises nucleotides independently selected from natural RNA nucleotides, modified RNA nucleotides, and nucleotide analogs.

First claim

1 . A pharmaceutical composition for changing the function of a coding or non-coding RNA in a cell comprising:
a double-stranded nucleic acid complex comprising: a first nucleic acid strand annealed to a second nucleic acid strand, wherein: the first nucleic acid strand comprises (i) nucleotides independently selected from natural nucleotides, modified nucleotides, and nucleotide analogs, (ii) the total number of natural nucleotides, modified nucleotides, and nucleotide analogs in the first nucleic acid strand is from 8 to 100, and (iii) the first nucleic acid strand is capable of hybridizing to RNA inside of the cell; and the second nucleic acid strand comprises nucleotides independently selected from natural RNA nucleotides, modified RNA nucleotides, and nucleotide analogs.

2 . The pharmaceutical composition of claim 1 , wherein the function is a modulation in the process of an RNA and the first nucleic acid strand comprises no regions that have 4 or more consecutive natural DNA nucleotides.

3 . The pharmaceutical composition of claim 2 , wherein the function of a coding or non-coding RNA is changed by inducing exon skipping.

4 . The pharmaceutical composition of claim 3 , wherein the first nucleic acid strand comprises at least one region consisting of 2 or 3 consecutive natural DNA nucleotides.

5 . The pharmaceutical composition of claim 4 , wherein the first nucleic acid strand comprises a bridged nucleotide/DNA mixmer oligonucleotide.

6 . The pharmaceutical composition of claim 1 , wherein the function is a modulation in the process of an RNA and the first nucleic acid strand comprises nucleotides independently selected from a morpholino oligonucleotide. a 2′-O-methyl modified oligonucleotide, a 2′-O-(2-methoxyethyl)modified oligonucleotide, or a bridged nucleotide oligonucleotide.

7 . The pharmaceutical composition of claim 6 , wherein the function of a coding or non-coding RNA is changed by inducing exon skipping.

8 . The pharmaceutical composition of claim 1 , wherein the first nucleic acid strand comprises 4 or more consecutive natural DNA nucleotide.

9 . The pharmaceutical composition of claim 8 , wherein the function is reducing the level of a transcription product and the first nucleic acid strand is capable of hybridizing to a non-coding region of a precursor mRNA inside of the cell.

10 . The pharmaceutical composition of claim 9 , wherein the first nucleic acid strand comprises a bridged nucleotide/DNA gapmer oligonucleotide.

11 . The pharmaceutical composition of claim 1 , wherein the total number of natural DNA nucleotides, modified DNA nucleotides, and nucleotide analogs in the first nucleic acid strand is from 10 to 35.

12 . The pharmaceutical composition of claim 5 , wherein the bridged nucleotides are independently selected from LNA, cEt-BNA, amideBNA (AmNA), and cMOE-BNA.

13 . The pharmaceutical composition of claim 5 , wherein the first nucleic acid strand comprises bridged nucleotides independently selected from a nucleotide in which the carbon atom at the 2′-position and the carbon atom at the 4′-position are bridged by 4′-(CH 2 ) p -O-2′, 4′-(CH 2 ) p -CH 2 -2′, 4′-(CH 2 ) p -S-2′,4′-(CH 2 ) p -OCO-2′,4′-(CH 2 ) n -N(R 3 )-O-(CH 2 ) m -2′, where p, m and n represent an integer from 1 to 4, an integer from 0 to 2, and an integer from 1 to 3, respectively, and R 3 represents a hydrogen atom, an alkyl group, an alkenyl group, a cycloalkyl group, an aryl group, an aralkyl group, an acyl group, a sulfonyl group, a fluorescent or chemiluminescent label, a functional group with nucleic acid cleavage activity, or an intracellular or intranuclear localization signal peptide.

14 . The pharmaceutical composition of claim 1 , wherein at least one of the modified nucleotides or one of the nucleotide analogs in the first nucleic acid strand is phosphorothioated.

15 . The pharmaceutical composition of claim 1 , wherein the first nucleic acid strand and/or the second nucleic acid strand further comprises a functional moiety having a function selected from a labeling function, a purification function, and a targeted delivery function.

16 . The pharmaceutical composition according to claim 15 , wherein said functional moiety is a molecule selected from a lipid, a sugar, a peptide, and a protein.

17 . The pharmaceutical composition according to claim 16 , wherein the functional moiety is joined to the 3′-terminal nucleotide and/or the 5′-terminal nucleotide of the first, second. or third nucleic acid strand.

18 . The pharmaceutical composition according to claim 17 , wherein the functional molecule is a peptide or protein selected from a receptor ligand and an antibody.

19 . The pharmaceutical composition according to claim 18 , wherein the functional molecule is independently selected from P007 and B peptide.

Assignees

• Nat Univ Corp Tokyo Med & Dent
• Univ Osaka

Inventors

• Yokota Takanori
• Nishina Kazutaka
• Yoshioka Kotaro
• Obika Satoshi
• Shimo Takenori

Classifications

• A61P21/00

  Drugs for disorders of the muscular or neuromuscular system · CPC title

• A61K31/713

  Double-stranded nucleic acids or oligonucleotides · CPC title

• C12N2310/341

  Gapmers, i.e. of the type ===---=== · CPC title

• C12N2310/315

  Phosphorothioates · CPC title

• C12N2310/3513

  Protein; Peptide · CPC title