TRICYCLO-PHOSPHOROTHIOATE DNA

• US 20140296323A1
• Filed: 10/12/2012
• Published: 10/02/2014
• Est. Priority Date: 10/13/2011
• Status: Active Grant

• Current Assignee: Institut National De La Sante Et De La Recherche Medicale (Inserm) (Allianz Global Investors GmbH), Centre National De La Recherche Scientifique, Institut de Myologie, Universitã¤T Bern, University Pierre et Marie Curie
• Original Assignee: Institut National De La Sante Et De La Recherche Medicale (Inserm) (Allianz Global Investors GmbH), Centre National De La Recherche Scientifique, Institut de Myologie, Universitã¤T Bern, University Pierre et Marie Curie
• Inventors: Leumann, Christian, Voit, Thomas, Garcia, Luis
• Granted Patent: US 9,738,891 B2
• Time in Patent Office: Days
• US Class Current: 514/44.A
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• CPC Class Codes: A61P 21/02 Muscle relaxants, e.g. for tetanus or cramps
• CPC Class Codes: A61P 21/04 for myasthenia gravis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 9/00 Drugs for disorders of the cardiovascular system
• 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 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/3231 having an additional ring, e.g. LNA, ENA
• CPC Class Codes: C12N 2320/33 Alteration of splicing

Abstract

The present invention relates to a nucleic acid molecule containing a sequence of tricyclo nucleosides joined by internucleoside phosphorothioate linkage. The invention also relates to synthetic antisense oligonucleotides and to methods employing the same.

Claims

1. A nucleic acid molecule comprising tricyclo-nucleosides joined by internucleoside phosphorothioate linkages (3′

-OPS-O-5′

linkages).

2. The nucleic acid molecule according to claim 1, comprising between 3 and 50 nucleotides.

3. The nucleic acid molecule according to claim 1, which is complementary to a target sequence.

4. The nucleic acid molecule according to claim 3, the nucleic acid molecule being an antisense oligonucleotide complementary to a portion of a RNA encoded by a gene.

5. The nucleic acid molecule of claim 1, comprising or consisting of the sequence selected in the group consisting of SEQ ID NOs:

1 to 11.

7. A pharmaceutical composition comprising a nucleic acid molecule according to claim 1, in a pharmaceutically acceptable carrier.

8. The pharmaceutical composition according to claim 7, said composition being an injectable composition, in particular a composition for intravenous injection.

15. A method for treating a heart disease, comprising administering to a subject in need thereof a nucleic acid molecule according to claim 1.

16. A method for treating a neuromuscular or musculoskeletal disease, comprising administering to a subject in need thereof a nucleic acid molecule according to claim 1.

17. A method for treating a central nervous system disease, comprising administering to a subject in need thereof a nucleic acid molecule according to claim 1.

18. The method according to claim 15, wherein the heart disease results from an alteration of a gene, wherein said alteration is

an in-frame mutation of an exon,

a mutation disrupting the translational reading frame of the gene, and the tc-DNA facilitates skipping of an exon so as to restore the reading frame;

a deleterious mutation that can be compensated by the inclusion of an atypical exon in the mRNA coded by said gene, and the tc-DNA is complementary to an ISS or TSL present in a pre-mRNA coded by said gene and facilitates inclusion of an atypical exon, or

a mutation resulting in the presence of deleterious 3′

CUG amplification(s) in a mRNA coded by said gene.

19. The method according to claim 16, wherein the neuromuscular or musculoskeletal disease results from an alteration of a gene, wherein said alteration is

an in-frame mutation of an exon,

a mutation disrupting the translational reading frame of the gene, and the tc-DNA facilitates skipping of an exon so as to restore the reading frame;

a deleterious mutation that can be compensated by the inclusion of an atypical exon in the mRNA coded by said gene, and the tc-DNA is complementary to an ISS or TSL present in a pre-mRNA coded by said gene and facilitates inclusion of an atypical exon, or

a mutation resulting in the presence of deleterious 3′

CUG amplification(s) in a mRNA coded by said gene.

20. The method according to claim 17, wherein the central nervous system disease results from an alteration of a gene, wherein said alteration is

an in-frame mutation of an exon,

a mutation disrupting the translational reading frame of the gene, and the tc-DNA facilitates skipping of an exon so as to restore the reading frame;

a deleterious mutation that can be compensated by the inclusion of an atypical exon in the mRNA coded by said gene, and the tc-DNA is complementary to an ISS or TSL present in a pre-mRNA coded by said gene and facilitates inclusion of an atypical exon, or

a mutation resulting in the presence of deleterious 3′

CUG amplification(s) in a mRNA coded by said gene.

21. The method according to claim 16, wherein the neuromuscular or musculoskeletal disease is selected in the group consisting of Duchenne Muscular Dystrophy, Spinal Muscular Atrophy, and Steinert'"'"'s Myotonic Dystrophy.

6. A method for synthesizing a tricyclo-phosphorothioate DNA molecule, the method comprising:

a) providing a first tricyclo-nucleoside bound to a solid phase support, said first nucleotide having a protected 5′

-OH group;

b) deprotecting the 5′

group to form a free 5′

-OH group;

c) reacting the free 5′

-OH group with a 5′

-protected tricyclonucleoside-3′

-O-cyanoethyl-N,N-diisopropylaminophosphoramidite monomer to form an internucleoside phosphoramidite linkage between the first and a second tricyclo-nucleosides; and

d) sulfurizing the internucleoside phosphoramidite group to form a phosphorothioate internucleoside linkage between the first and second tricyclo-nucleosides.

9. (canceled)

10. (canceled)

11. (canceled)

12. (canceled)

13. (canceled)

14. (canceled)