Method of stimulating an immune response and inhibiting expression of a gene using an oligonucleotide

• US 8,003,619 B2
• Filed: 12/09/2005
• Issued: 08/23/2011
• Est. Priority Date: 12/09/2004
• Status: Active Grant

• Current Assignee: Alnylam Pharmaceuticals Incorporated
• Original Assignee: Alnylam Pharmaceuticals Incorporated
• Inventors: Endres, Stefan, Hornung, Veit, de Fougerolles, Antonin, Hartmann, Gunther
• Primary Examiner(s): Angell; Jon E
• Application Number:
  US11/298,850
• Publication Number:
  US 20060217330A1
• Time in Patent Office: 2,083 Days
• Field of Search: 514/44.A, 514/44.R
• US Class Current: 514/44.00A
• CPC Class Codes: A61K 2039/53 DNA (RNA) vaccination
• CPC Class Codes: A61K 39/00 Medicinal preparations containing antigens or antibodies materials for immunoassay G01N33/53
• CPC Class Codes: A61K 39/39 characterised by the immunostimulating additives, e.g. chemical adjuvants
• CPC Class Codes: A61P 11/00 Drugs for disorders of the respiratory system
• CPC Class Codes: A61P 11/06 Antiasthmatics
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• 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: C12N 15/1138 against receptors or cell surface proteins
• CPC Class Codes: C12N 15/117 Nucleic acids having immunomodulatory properties, e.g. containing CpG-motifs
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/17 Immunomodulatory nucleic acids
• CPC Class Codes: C12N 2310/3231 having an additional ring, e.g. LNA, ENA

Abstract

This invention provides oligonucleotide agents that modulate an immune response by stimulating IFN production and methods of using such agents for therapeutic treatments of mammals.

Claims

1. A method of stimulating an immune response in a mammal comprising the steps of administering to said mammal an oligonucleotide agent comprising a sequence that differs by not more than 1 or not more than 2 nucleotides from SEQ ID NO:

1, and assaying the mammal for IFNα

expression prior to and after the administering step to determine that the immune response was stimulated in the mammal, wherein the oligonucleotide agent is at least 19 nucleotides in length.

2. The method of claim 1, wherein the oligonucleotide agent comprises SEQ ID NO:

1.

3. The method of claim 1, wherein said sequence comprises the sequence of any one the following:

SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

4, SEQ ID NO;

5, and SEQ ID NO;

6.

7. The method of claim 1, wherein the oligonucleotide agent is an siRNA comprising a sense strand and an antisense strand, wherein the antisense strand comprises a sequence of at least 19, 20, 21 or 23 nucleotides.

8. The method of claim 7, wherein the sense strand comprises a sequence of at least 19, 20, or 21 contiguous nucleotides of SEQ ID NO:

20.

9. The method of claim 7, wherein the sense strand consists of SEQ ID NO:

20.

10. The method of claim 7, wherein the sense strand comprises a sequence of at least 19, 20, or 21 contiguous nucleotides of SEQ ID NO:

22.

11. The method of claim 7, wherein the sense strand consists of SEQ ID NO:

22.

12. The method of claim 7, wherein the sense strand comprises a sequence that differs by not more than 1 or not more than 2 nucleotides from SEQ ID NO:

1.

13. The method of claim 1, wherein the oligonucleotide agent is single stranded.

14. The method of claim 1, wherein the oligonucleotide agent is a hybrid oligonucleotide.

15. The method of claim 1, wherein said method is used for treatment of cancer, autoimmune disorders, asthma, respiratory allergies, food allergies, bacteria infections, parasitic infections, or viral infections and wherein the mammal is in need of said treatment.

16. The method of claim 1, wherein an antigen is conjugated to the oligonucleotide agent at a position other than the accessible 5′

end.

17. The method of claim 1, wherein an antigen is conjugated to the oligonucleotide by a non-nucleotidic linker.

18. The method of claim 17, wherein the antigen is conjugated to the oligonucleotide at a position other than its 3′

end.

19. The method of claim 1, wherein immune recognition of the oligonucleotide requires toll-like receptor 7 (TLR7).

20. The method of claim 1, wherein the oligonucleotide comprises two repeats of SEQ ID NO:

1.

21. The method of claim 20, wherein the oligonucleotide comprises SEQ ID NO:

38.

22. The method of claim 7, wherein the immunostimulatory activity is maintained while abolishing the silencing activity of the siRNA.

4. A method of concomitantly inhibiting the expression of a gene and inducing an immune response in a mammal, comprising administering to said mammal an iRNA agent comprising a sequence that differs by not more than 1 or not more than 2 nucleotides of SEQ ID NO:

1 and assaying the mammal for IFNα

expression prior to and after the administering step to determine that the immune response was stimulated in the mammal, wherein the iRNA agent is at least 19 nucleotides in length.

5. The method of claim 4, wherein said sequence comprises the sequence of any one the following:

SEQ ID NO;

2, SEQ ID NO;

3, SEQ ID NO;

4, SEQ ID NO;

5, and SEQ ID NO;

6.

6. The method of claim 4, wherein the iRNA agent comprises SEQ ID NO:

1.