Tricyclo-DNA antisense oligonucleotides, compositions, and methods for the treatment of disease

• US 10,465,191 B2
• Filed: 02/09/2017
• Issued: 11/05/2019
• Est. Priority Date: 04/10/2009
• 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, Sorbonne Universite
• Original Assignee: Institut de Myologie
• Inventors: Schumperli, Daniel, Leumann, Christian, Furling, Denis, Garcia, Luis, Voit, Thomas
• Primary Examiner(s): Zara, Jane J
• Application Number:
  US15/428,385
• Publication Number:
  US 20170260524A1
• Time in Patent Office: 999 Days
• Field of Search: 435 61, 435 611, 435 911, 435 9131, 435455, 435375, 514 44, 536 231, 536 245
• CPC Class Codes: A61K 31/711 Natural deoxyribonucleic acids, i.e. containing only 2'-deoxyriboses attached to adenine, guanine, cytosine or thymine and having 3'-5' phosphodiester links
• CPC Class Codes: A61K 31/712 Nucleic acids or oligonucleotides having modified sugars, i.e. other than ribose or 2'-deoxyribose
• CPC Class Codes: A61P 19/08 for bone diseases, e.g. rachitism, Paget's disease
• CPC Class Codes: A61P 21/00 Drugs for disorders of the muscular or neuromuscular system
• 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 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• 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 2310/3231 having an additional ring, e.g. LNA, ENA
• CPC Class Codes: C12N 2320/33 Alteration of splicing

Abstract

Provided are tricyclo-DNA (tc-DNA) AON and methods employing tc-DNA AON for modifying splicing events that occur during pre-mRNA processing. Tricyclo-DNA (tc-DNA) AON are described that may be used to facilitate exon skipping or to mask intronic silencer sequences and/or terminal stein-loop sequences during pre-mRNA processing and to target RNase-mediated destruction of processed mRNA. Tc-DNA AON described herein may be used in methods for the treatment of Duchenne Muscular Dystrophy by skipping a mutated exon 23 or exon 51 within a dystrophin gene to restore functionality of a dystrophin protein; in methods for the treatment of Spinal Muscular Atrophy by masking an intronic silencing sequence and/or a terminal stem-loop sequence within an SMN2 gene to yield modified functional SMN2 protein, including an amino acid sequence encoded by exon 7, which is capable of at least partially complementing a non-functional SMN1 protein; and in methods for the treatment of Steinert'"'"'s Myotonic Dystrophy by targeting the destruction of a mutated DM1 mRNA comprising 3′-terminal CUG repeats.

Claims

1. A method for the delivery of an antisense oligonucleotide (AON) into a human patient comprising administering to the patient a tricyclo-DNA (tc-DNA) antisense oligonucleotide (AON),

wherein the tc-DNA AON has a length of 10 to 18 nucleotides, wherein all the nucleotides of the tc-DNA AON are tc-DNA nucleotides;

wherein the tc-DNA AON comprises a sequence of nucleotides that is complementary to an intron-exon junction; and

wherein the administration of the tc-DNA AON to the patient results in increased exon skipping at the intron-exon junction in the patient.

2. The method of claim 1, wherein the tc-DNA AON comprises a sequence of nucleotides that is complementary to an intron-exon junction within a mutated dystrophin pre-mRNA.

3. The method of claim 2, wherein the administration of the tc-DNA AON to the patient results in increased skipping of exon 23 in the patient.

4. The method of claim 2, wherein the wherein the administration of the tc-DNA AON to the patient results in increased skipping of exon 51 in the patient.

5. The method of claim 1, wherein the tc-DNA AON has a length of 12 to 16 nucleotides.

6. The method of claim 1, wherein the tc-DNA AON has a length of 13 to 15 nucleotides.

7. The method of claim 1, wherein the administration comprises intramuscular (i.m.), intravenous (i.v.), intraperitoneal (i.p.), intra-arterial, sub-cutaneous or transdermic injection of the tc-DNA AON.

8. The method of claim 1, comprising administering 1 mg/kg to 100 mg/kg body weight/day to the patient.

9. The method of claim 1, comprising injecting the tc-DNA AON directly into the muscle of the patient.

10. The method of claim 1, wherein 8-16 nucleotides of said tc-DNA AON are complementary to a dystrophin pre-mRNA intronic splice donor site, wherein 2-8 nucleotides of said tc-DNA AON are complementary to a dystrophin pre-mRNA exonic region, and wherein said exonic region is contiguous with and 3′

to said intronic splice donor site.

11. The method of claim 1, wherein the tc-DNA AON comprises the nucleotide sequence 5′

-AACCTCGGCTTACCT-3′

(SEQ ID NO;

     1).

12. The method of claim 1, wherein the tc-DNA AON is complementary to the nucleotide sequence 5′

-AGAAATGCCATCTTC-3′

(SEQ ID NO;

     2).

13. The method of claim 1, wherein the tc-DNA AON is complementary to the nucleotide sequence 5′

-AAATGCCATCTTCCT-3′

(SEQ ID NO;

     3).

14. The method of claim 1, wherein said tc-DNA AON is complementary to the nucleotide sequence 5′

-TGCCATCTTCCTTGA-3′

(SEQ ID NO;

     4).

15. The method of claim 1, wherein the patient has Duchene Muscular Dystrophy (DMD).