Antisense oligonucleotide compositions and methods for the modulation of activating protein 1

• US 6,312,900 B1
• Filed: 07/30/1999
• Issued: 11/06/2001
• Est. Priority Date: 04/14/1997
• Status: Expired due to Term

• Current Assignee: ISIS Pharmaceuticals Incorporated (Ionis Pharmaceuticals, Inc.)
• Original Assignee: ISIS Pharmaceuticals Incorporated (Ionis Pharmaceuticals, Inc.)
• Inventors: Baker, Brenda, Miraglia, Loren, McKay, Robert, Dean, Nicholas M.
• Primary Examiner(s): WANG, ANDREW J
• Application Number:
  US09/364,416
• Time in Patent Office: 830 Days
• Field of Search: 536/23.1, 536/24.5, 435/6, 435/91.1, 435/325, 435/375, 514/44
• US Class Current: 435/6.13
• CPC Class Codes: A61K 38/00 Medicinal preparations containing peptides peptides containing beta-lactam rings A61K31/00; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, A61K31/00; ergot alkaloids of the cyclic peptide type A61K31/48; containing macromolecular compounds having statistically distributed amino acid units A61K31/74; medicinal preparations containing antigens or antibodies A61K39/00; medicinal preparations characterised by the non-active ingredients, e.g. peptides as drug carriers, A61K47/00
• CPC Class Codes: C12N 15/1135 against oncogenes or tumor suppressor genes
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/3181 Peptide nucleic acid, PNA
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 2'-R Modification
• CPC Class Codes: C12N 2310/33 of the base
• CPC Class Codes: C12N 2310/334 Modified C
• CPC Class Codes: C12N 2310/335 Modified T or U
• CPC Class Codes: C12N 2310/3521 Methyl
• CPC Class Codes: C12N 2310/3525 MOE, methoxyethoxy
• CPC Class Codes: C12N 2310/3527 Other alkyl chain

Abstract

Compositions and methods for the treatment and diagnosis of diseases or disorders amenable to treatment through modulation of Activating Protein 1 (AP-1) expression are provided. In accordance with various embodiments of the present invention, oligonucleotides are provided which are specifically hybridizable with c-fos or c-jun, the genes encoding c-Fos or c-Jun, respectively. In a preferred embodiment, a method of modulating the metastasis of malignant tumors via modulation of one or more of the AP-1 subunits is provided; this method can be effected using the oligonucleotides of the invention or any other agent which modulates AP-1 or AP-1-mediated transcription.

Claims

1. An antisense oligonucleotide comprising 8-30 nucleobases connected by covalent linkages, wherein said antisense oligonucleotide comprises at least an 8 nucleobase portion of SEQ ID NO:

10, 11, 12, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 27, 120, 121, 122, 123, 131, 132, 133, 134, 135, 136, 137, or 138, wherein said antisense oligonucleotide inhibits the expression of a c-Fos protein, wherein said antisense oligonucleotide comprises at least one 2′

methoxyethoxy residue, and wherein said antisense oligonucleotide comprises at least one lipophilic moiety which enhances the cellular uptake of said antisense oligonucleotide.

2. The antisense oligonucleotide of claim 1, wherein said lipophilic moiety is selected from the group consisting of a cholesterol moiety, a cholesteryl moiety, cholic acid, a thioether, a thiocholesterol, an aliphatic chain, a phospholipid, a polyamine chain, a polyethylene glycol chain, adamantane acetic acid, a palmityl moiety, an octadecylamine moiety and a hexylamino-carbonyl-oxycholesterol moiety.

3. The antisense oligonucleotide of claim 1, wherein at least one of said linkages is selected from the group consisting of a phosphorothioate linkage, a phosphodiester linkage, a phosphotriester linkage, a methyl phosphonate linkage, a methylene(methylimino) linkage, a morpholino linkage, an amide linkage, a polyamide linkage, a short chain alkyl intersugar linkage, a cycloalkyl intersugar linkage, a short chain heteroatomic intersugar linkage and a heterocyclic intersugar linkage.

4. The antisense oligonucleotide of claim 1, wherein at least one of said nucleotides has a modified sugar moiety.

5. The antisense oligonucleotide of claim 4, wherein said modified sugar moiety is a modification at the 2′

position of any nucleotide, the 3′

position of the 3′

terminal nucleotide or the 5′

position of the 5′

terminal oligonucleotide.

6. The antisense oligonucleotide of claim 5, wherein said modification is selected from the group consisting of a substitution of an azido group for a 3′

hydroxyl group and a substitution of a hydrogen atom for a 3′

or 5′

hydroxyl group.

7. The antisense oligonucleotide of claim 5, wherein said modification is a substitution or addition at the 2′

position of a moiety selected from the group consisting of —

OH, —

SH, —

SCH₃, —

F, —

OCN, —

OCH₃OCH₃, —

OCH₃O(CH₂)_nCH₃, —

O(CH₂)_nNH₂ or —

O(CH₂)_nCH₃ where n is from 1 to about 10, a C₁ to C₁₀ lower alkyl group, an alkoxyalkoxy group, a substituted lower alkyl group, a substituted alkaryl group, a substituted aralkyl group, —

Cl, —

Br, —

CN, —

CF₃, —

OCF₃, an -O-alkyl group, an —

S-alkyl group, an N-alkyl group, an O-alkenyl group, an S-alkenyl group, an N-alkenyl group, —

SOCH₃, —

SO₂CH₃, —

ONO₂, —

NO₂, —

N₃, —

NH₂, a heterocycloalkyl group, a heterocycloalkaryl group, an aminoalkylamino group, a polyalkylamino group, a substituted silyl group, an RNA cleaving group, a reporter group, a DNA intercalating group, a group for improving the pharmacokinetic properties of an oligonucleotide, a group for improving the pharmacodynamic properties of an oligonucleotide, a methoxyethoxy group and a methoxy group.

8. The antisense oligonucleotide of claim 1, wherein at least one of said nucleotides has a modified nucleobase.

9. The antisense oligonucleotide of claim 8, wherein said modified nucleobase is selected from the group consisting of 5 hypoxanthine, 5-methylcytosine, 5-hydroxymethylcytosine, glycosyl 5-hydroxymethylcytosine, gentiobiosyl 5-hydroxymethylcytosine, 5-bromouracil, 5-hydroxymethyluracil, 6-methyladenine, N⁶-(6-aminohexyl)adenine, 8-azaguanine, 7-deazaguanine and 2,6-diaminopurine.

10. The antisense oligonucleotide of claim 1, wherein said c-Fos protein is human c-Fos.

11. A pharmaceutical composition comprising the antisense oligonucleotide of claim 1, and a pharmaceutically acceptable carrier.

23. A pharmaceutical composition comprising:

(a) a chemotherapeutic agent;

(b) the antisense oligonucleotide of claim 1, and

(c) a pharmaceutically acceptable carrier.

27. A method of inhibiting the expression of a c-Fos protein in cells or tissues comprising contacting said cells or tissues with an antisense oligonucleotide of claim 1, so that expression of said c-Fos protein is inhibited.

12. An antisense oligonucleotide comprising 8-30 nucleotides connected by covalent linkages, wherein said antisense oligonucleotide comprises at least an 8 nucleobase portion of SEQ ID NO:

1, 3, 4, 5, 6, 7, 8, 9, 36, or 37, wherein said antisense oligonucleotide inhibits the expression of a c-Jun protein, wherein said antisense oligonucleotide comprises at least one 2′

methoxyethoxy residue, and wherein said antisense oligonucleotide comprises at least one lipophilic moiety which enhances the cellular uptake of said antisense oligonucleotide.

13. The antisense oligonucleotide of claim 12, wherein said lipophilic moiety is selected from the group consisting of a cholesterol moiety, a cholesteryl moiety, cholic acid, a thioether, a thiocholesterol, an aliphatic chain, a phospholipid, a polyamine chain, a polyethylene glycol chain, adamantane acetic acid, a palmityl moiety, an octadecylamine moiety and a hexylamino-carbonyl-oxycholesterol moiety.

14. The antisense oligonucleotide of claim 12, wherein at least one of said linkages is selected from the group consisting of a phosphorothioate linkage, a phosphodiester linkage, a phosphotriester linkage, a methyl phosphonate linkage, a methylene(methylimino) linkage, a morpholino linkage, an amide linkage, a polyamide linkage, a short chain alkyl intersugar linkage, a cycloalkyl intersugar linkage, a short chain heteroatomic intersugar linkage and a heterocyclic intersugar linkage.

15. The antisense oligonucleotide of claim 12, wherein at least one of said nucleotides has a modified sugar moiety.

16. The antisense oligonucleotide of claim 15, wherein said modified sugar moiety is a modification at the 2′

position of any nucleotide, the 3′

position of the 3′

terminal nucleotide or the 5′

position of the 5′

terminal oligonucleotide.

17. The antisense oligonucleotide of claim 16, wherein said modification is selected from the group consisting of a substitution of an azido group for a 3′

hydroxyl group and a substitution of a hydrogen atom for a 3′

or 5′

hydroxyl group.

18. The antisense oligonucleotide of claim 16, wherein said modification is a substitution or addition at the 2′

position of a moiety selected from the group consisting of —

OH, —

SH, —

SCH₃, —

F, —

OCN, —

OCH₃OCH₃, —

OCH₃O (CH₂)_nCH₃, —

O(CH₂)_nNH₂ or —

O(CH₂)_nCH₃ where n is from 1 to about 10, a C₁ to C₁₀ lower alkyl group, an alkoxyalkoxy group, a substituted lower alkyl group, a substituted alkaryl group, a substituted aralkyl group, —

Cl, —

Br, —

CN, —

CF₃₁, —

OCF₃, an -O-alkyl group, an —

S-alkyl group, an N-alkyl group, an O-alkenyl group, an S-alkenyl group, an N-alkenyl group, —

SOCH₃, —

SO₂CH₃, —

ONO₂, —

NO₂, —

N₃, —

NH₂, a heterocycloalkyl group, a heterocycloalkaryl group, an aminoalkylamino group, a polyalkylamino group, a substituted silyl group, an RNA cleaving group, a reporter group, a DNA intercalating group, a group for improving the pharmacokinetic properties of an oligonucleotide, a group for improving the pharmacodynamic properties of an oligonucleotide, a methoxyethoxy group and a methoxy group.

19. The antisense oligonucleotide of claim 12, wherein at least one of said nucleotides has a modified nucleobase.

20. The antisense oligonucleotide of claim 19, wherein said modified nucleobase is selected from the group consisting of hypoxanthine, 5-methylcytosine, 5-hydroxymethylcytosine, glycosyl 5-hydroxymethylcytosine, gentiobiosyl 5-hydroxymethylcytosine, 5-bromouracil, 5-hydroxymethyluracil, 6-methyladenine, N⁶-(6-aminohexyl)adenine, 8-azaguanine, 7-5 deazaguanine and 2,6-diaminopurine.

21. The antisense oligonucleotide of claim 12, wherein said c-Jun protein is human c-Jun.

22. A pharmaceutical composition comprising the antisense oligonucleotide of claim 12, and a pharmaceutically acceptable carrier.

24. A pharmaceutical composition comprising:

(a) a chemotherapeutic agent;

(b) the antisense oligonucleotide of claim 12, and

(c) a pharmaceutically acceptable carrier.

25. A pharmaceutical composition comprising:

(a) an antisense oligonucleotide of claim 13;

(b) an antisense oligonucleotide comprising 8 to 30 nucleotides connected by covalent linkages, wherein said antisense oligonucleotide has a sequence specifically hybridizable with a nucleic acid encoding a c-Jun protein, wherein said antisense oligonucleotide inhibits the expression of said c-Jun protein and wherein said antisense oligonucleotide comprises at least one 2′

-methoxyethoxy residue; and

(c) a pharmaceutically acceptable carrier.

26. A pharmaceutical composition comprising:

(a) a chemotherapeutic agent;

(b) an antisense oligonucleotide of claim 13;

(c) an antisense oligonucleotide comprising 8 to 30 nucleotides connected by covalent linkages, wherein said antisense oligonucleotide has a sequence specifically hybridizable with a nucleic acid encoding a c-Jun protein, wherein said antisense oligonucleotide inhibits the expression of said c-Jun protein and wherein said antisense oligonucleotide comprises at least one 2′

-methoxyethoxy residue; and

(d) a pharmaceutically acceptable carrier.

28. A method of inhibiting the expression of a c-Jun protein in cells or tissues comprising contacting said cells or tissues with an antisense oligonucleotide of claim 12, so that expression of said c-Jun protein is inhibited.

29. An antisense oligonucleotide comprising 8-30 nucleotides connected by covalent linkages, wherein said antisense oligonucleotide comprises at least an 8 nucleobase portion of SEQ ID NO:

10, 11, 12, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 27, 120, 121, 122, 123, 131, 132, 133, 134, 135, 136, 137, or 138, wherein said antisense oligonucleotide inhibits the expression of a c-Fos protein, and wherein said antisense oligonucleotide comprises two or more chemically distinct regions.

30. The antisense oligonucleotide of claim 29 wherein said antisense oligonucleotide is a hemimer or a gapmer.

31. The antisense oligonucleotide of claim 29 wherein one of said chemically distinct regions comprises one or more 2′

-methoxyethoxy residues.

32. An antisense oligonucleotide comprising 8-30 nucleotides connected by covalent linkages, wherein said antisense oligonucleotide comprises at least an 8 nucleobase portion of SEQ ID NO:

1, 3, 4, 5, 6, 7, 8, 9, 36, or 37, wherein said antisense oligonucleotide inhibits the expression of a c-Jun protein, and wherein said antisense oligonucleotide comprises two or more chemically distinct regions.

33. The antisense oligonucleotide of claim 32, wherein said antisense oligonucleotide is a hemimer or a gapmer.

34. The antisense oligonucleotide of claim 32, wherein one of said chemically distinct regions comprises one or more 2′

-methoxyethoxy residues.