Modifications for antisense compounds

• US 9,506,059 B2
• Filed: 09/26/2014
• Issued: 11/29/2016
• Est. Priority Date: 03/26/2010
• Status: Active Grant

• Current Assignee: Integrated DNA Technologies (Danaher Corporation)
• Original Assignee: Integrated DNA Technologies (Danaher Corporation)
• Inventors: Behlke, Mark Aaron, Owczarzy, Richard, You, Yong, Walder, Joseph Alan, Lennox, Kim
• Primary Examiner(s): ZARA, JANE J
• Application Number:
  US14/281,646
• Publication Number:
  US 20150011744A1
• Time in Patent Office: 795 Days
• Field of Search: 435/6.1, 435/6.12, 435/91.1, 435/91.31, 436/501, 536/23.1, 536/24.5, 536/24.3, 536/26.6
• US Class Current: 1/1
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: 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
• CPC Class Codes: C12N 2310/11 Antisense
• CPC Class Codes: C12N 2310/141 MicroRNAs, miRNAs
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/344 Position-specific modifications, e.g. on every purine, at the 3'-end
• CPC Class Codes: C12N 2310/352 linked to the nucleic acid via a carbon atom
• CPC Class Codes: C12N 2310/3529 Aromatic substituent
• CPC Class Codes: C12N 2320/51 modulating the chemical stability, e.g. nuclease-resistance
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes

Abstract

The invention pertains to modifications for antisense oligonucleotides, wherein the modifications are used to improve stability and provide protection from nuclease degradation. The modifications could also be incorporated into double-stranded nucleic acids, such as synthetic siRNAs and miRNAs.

Claims

1. An antisense oligonucleotide comprising at least one modification that is incorporated between two bases of the antisense oligonucleotide, wherein the modification increases binding affinity and nuclease resistance of the antisense oligonucleotide, and wherein the modification has the structure:

2. The antisense oligonucleotide of claim 1, wherein the modification is located within three bases of a terminal nucleotide.

3. The antisense oligonucleotide of claim 1, wherein the modification is located between a terminal base and a penultimate base of either the 3′

or the 5′

end of the antisense oligonucleotide.

4. The antisense oligonucleotide of claim 1, wherein the modification is located between the terminal base and the penultimate base at both the 3′

and the 5′

ends of the antisense oligonucleotide.

5. The antisense oligonucleotide of claim 1, wherein the modification is a napthylene-azo compound.

6. The antisense oligonucleotide of claim 1, wherein the modification has the structure:

7. The antisense oligonucleotide of claim 6, wherein R₈ is an electron withdrawing group, which is NO₂.

8. The antisense oligonucleotide of claim 1, wherein the modification has the structure:

9. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide further comprises at least one 2′

-O-methyl RNA.

10. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide further comprises one or more phosphorothioate linkages.

11. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide comprises a region of bases linked through phosphodiester bonds, wherein the region is flanked at one or both ends by regions containing phosphorothioate linkages.

12. An antisense oligonucleotide having the structure:

5′

-X₁—

Z—

X₂—

X₃—

X₄—

Z—

X₅-3′

wherein;

X₁ and X₅ are independently 1-3 nucleotides;

the internucleotide linkages are optionally phosphorothioate;

Z is a napthylene-azo compound;

X₂ and X₄ are independently 1-5 nucleotides wherein the internucleotide linkages are optionally phosphorothioate; and

X₃ is 10-25 nucleotides.