Immunostimulatory nucleic acids

• US 20050059619A1
• Filed: 08/19/2003
• Published: 03/17/2005
• Est. Priority Date: 08/19/2002
• Status: Active Grant

• Current Assignee: Coley Pharmaceutical GmbH (Pfizer Inc.)
• Original Assignee: Coley Pharmaceutical GmbH (Pfizer Inc.), Coley Pharmaceutical Group, Inc. (Pfizer Inc.)
• Inventors: Jurk, Marion, Krieg, Arthur M., Uhlmann, Eugen, Samulowitz, Ulrike, Vollmer, Jorg, Rankin, Robert, Lipford, Grayson
• Granted Patent: US 8,283,328 B2
• Time in Patent Office: Days
• US Class Current: 514/44.A
• CPC Class Codes: A61K 2039/55561 CpG containing adjuvants; Oligonucleotide containing adjuvants
• CPC Class Codes: A61K 31/70 Carbohydrates; Sugars; Derivatives thereof sorbitol A61K31/047
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 11/06 Antiasthmatics
• CPC Class Codes: A61P 11/08 Bronchodilators
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 31/04 Antibacterial agents
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/02 Immunomodulators
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 37/08 Antiallergic agents antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14
• CPC Class Codes: A61P 43/00 Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
• CPC Class Codes: C07H 19/00 Compounds containing a hetero ring sharing one ring hetero atom with a saccharide radical; Nucleosides; Mononucleotides; Anhydro-derivatives thereof
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07H 21/02 with ribosyl as saccharide radical
• CPC Class Codes: C07H 21/04 with deoxyribosyl as saccharide radical
• CPC Class Codes: C12N 15/117 Nucleic acids having immunomodulatory properties, e.g. containing CpG-motifs
• CPC Class Codes: C12N 2310/312 Phosphonates
• CPC Class Codes: C12N 2310/3125 Methylphosphonates
• CPC Class Codes: C12N 2310/313 : Phosphorodithioates
• CPC Class Codes: C12N 2310/315 : Phosphorothioates
• CPC Class Codes: C12N 2310/336 : Modified G
• CPC Class Codes: C12N 2310/345 : having at least two different backbone modifications

Abstract

The invention relates to a class of soft or semi-soft CpG immunostimulatory oligonucleotides that are useful for stimulating an immune response.

Claims

1. An immunostimulatory nucleic acid molecule having at least one internal pyrimidine-purine (YZ) dinucleotide and a chimeric backbone, wherein the at least one internal YZ dinucleotide has a phosphodiester or phosphodiester-like internucleotide linkage, wherein optionally each additional internal YZ dinucleotide has a phosphodiester, phosphodiester-like, or stabilized internucleotide linkage, and wherein all other internucleotide linkages are stabilized.

2. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid comprises a plurality of internal YG dinucleotides having a phosphodiester or phosphodiester-like internucleotide linkage.

3. The oligonucleotide of claim 2, wherein every internal YG dinucleotide has a phosphodiester or phosphodiester-like internucleotide linkage.

4. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid molecule is any one of SEQ ID NO:

1-54, SEQ ID NO;

55-99 and SEQ ID NO;

241, wherein * shown in the SEQ ID No'"'"'s in the specification represents phosphorothioate, _ represents phosphodiester, U represents 2′

-deoxyuracil, and 7 represents 7-deazaguanine.

5. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid molecule is selected from the group consisting of:

12. The oligonucleotide of claim 1, wherein the at least one internal YG dinucleotide having a phosphodiester or phosphodiester-like internucleotide linkage is CG.

13. The oligonucleotide of claim 1, wherein the at least one internal YG dinucleotide having a phosphodiester or phosphodiester-like internucleotide linkage is TG.

14. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid molecule is a B-Class immunostimulatory nucleic acid molecule.

15. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid molecule is a C-Class immunostimulatory nucleic acid molecule.

16. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid molecule is 4-100 nucleotides long.

17. The oligonucleotide of claim 1, wherein the immunostimulatory nucleic acid molecule is not an antisense oligonucleotide, triple-helix-forming oligonucleotide, or ribozyme.

22. The oligonucleotide of claim 1 wherein the nucleic acid has a backbone comprising deoxyribose or ribose.

23. The oligonucleotide of claim 1, wherein the oligonucleotide further comprises an adjuvant or a cytokine, or an antigen.

24. The oligonucleotide of claim 1 wherein the phosphodiester or phosphodiester-like internucleotide linkage is phosphodiester.

25. The oligonucleotide of claim 1 wherein the phosphodiester-like linkage is boranophosphonate or diastereomerically pure Rp phosphorothioate.

26. The oligonucleotide of claim 1 wherein the stabilized internucleotide linkages are selected from the group consisting of:

phosphorothioate, phosphorodithioate, methylphosphonate, methylphosphorothioate, and any combination thereof.

27. The oligonucleotide of claim 1 wherein the stabilized internucleotide linkages are phosphorothioate.

70. A method for modulating an immune response, comprising administering to a subject an oligonucleotide of claim 1, in an effective amount to modulate an immune response.

6-11. -11. (canceled)

18-21. -21. (canceled)

28. An oligonucleotide comprising:

29-31. -31. (canceled)

32. An oligonucleotide comprising:

33-35. -35. (canceled)

36. An oligonucleotide comprising:

37-38. -38. (canceled)

39. An oligonucleotide comprising:

40-43. -43. (canceled)

44. An oligonucleotide comprising:

45. (canceled)

46. An oligonucleotide comprising:

47. (canceled)

48. An oligonucleotide comprising:

49. An oligonucleotide comprising:

an octameric sequence comprising at least one YZ dinucleotide having a phosphodiester or phosphodiester-like internucleotide linkage, and at least 4 T nucleotides, wherein Y is a pyrimidine or modified pyrimidine, wherein Z is a guanosine or modified guanosine, and wherein the oligonucleotide includes at least one stabilized internucleotide linkage.

50-65. -65. (canceled)

66. An oligonucleotide comprising:

67. An oligonucleotide comprising:

5′

GNC 3′

, wherein N is a nucleic aid sequence of 4-10 nucleotides in length and is at least 50% T and does not include a CG dinucleotide, and the oligonucleotide includes at least one stabilized internucleotide linkage.

68-69. -69. (canceled)

71-87. -87. (canceled)

88. A method for treating airway remodeling, comprising:

administering to a subject an oligonucleotide comprising a CG dinucleotide, in an effective amount to treat airway remodeling in the subject.

89-93. -93. (canceled)

94. A method for stimulating an immune response, comprising

administering to a subject an oligonucleotide of at least 5 nucleotides in length in an effective amount to stimulate an immune response, wherein the oligonucleotide includes at least one immunostimulatory dinucleotide motif wherein the internucleotide linkage between the nucleotides of the dinucleotide has R chirality and wherein at least 70% of the other internucleotide linkages of the oligonucleotide have S chirality.

95. An oligonucleotide, comprising:

an immunostimulatory nucleic acid molecule comprising a chimeric backbone and at least one sequence N₁YGN₂, wherein independently for each sequence N₁YGN₂ YG is an internal pyrimidine-guanosine (YG) dinucleotide, N₁ and N₂ are each, independent of the other, any nucleotide, and wherein for the at least one sequence N₁YGN₂ and optionally for each additional sequence N₁YGN₂;

the YG dinucleotide has a phosphodiester or phosphodiester-like internucleotide linkage, and

(a) N₁ and Y are linked by a phosphodiester or phosphodiester-like internucleotide linkage when N₁ is an internal nucleotide,

(b) G and N₂ are linked by a phosphodiester or phosphodiester-like internucleotide linkage when N₂ is an internal nucleotide, or

(c) N₁ and Y are linked by a phosphodiester or phosphodiester-like internucleotide linkage when N₁ is an internal nucleotide and G and N₂ are linked by a phosphodiester or phosphodiester-like internucleotide linkage when N₂ is an internal nucleotide, wherein all other internucleotide linkages are stabilized.

96. An oligonucleotide comprising

N₁-C_G-N₂-C_G-N₃

wherein N₁ and N₃ are each independently a nucleic acid sequence 1-20 nucleotides in length, wherein _ indicates an internal phosphodiester or phosphodiester-like internucleotide linkage, wherein N₂ is independently a nucleic acid sequence 0-20 nucleotides in length, and wherein G-N₂-C includes 1 or 2 stabilized linkages.

97. An oligonucleotide comprising

N₁-C_G-N₂-C_G-N₃

wherein N₁ and N₃ are each independently a nucleic acid sequence 1-20 nucleotides in length, wherein _ indicates an internal phosphodiester or phosphodiester-like internucleotide linkage, wherein N₂ is independently a nucleic acid sequence 4-20 nucleotides in length, and wherein G-N₂-C includes at least 5 stabilized linkages.

98. An oligonucleotide comprising

N₁-C_G-N₂-C_G-N₃

wherein N₁, N₂, and N₃ are each independently a nucleic acid sequence of 0-20 nucleotides in length and wherein _ indicates an internal phosphodiester or phosphodiester-like internucleotide linkage, wherein the oligonucleotide is not an antisense oligonucleotide, triple-helix-forming oligonucleotide, or ribozyme.

99. An oligonucleotide comprising

X₁-N₁-(GTCGTT)_n-N₂-X₂

wherein N₁ and N₂ are each independently a nucleic acid sequence of 0-20 nucleotides in length, wherein n=2 or n=4-6, wherein X₁ and X₂ are each independently a nucleic acid sequence having phosphorothioate internucleotide linkages of 3-10 nucleotides, wherein N₁-(GTCGTT)_n-N₂ includes at least one phosphodiester internucleotide linkage, and wherein 3′

and 5′

nucleotides of the oligonucleotide do not include a poly-G, poly-A, poly-T, or poly-C sequence.