Ultra-small RNAs as toll-like receptor-3 antagonists

• US 8,481,508 B2
• Filed: 02/20/2009
• Issued: 07/09/2013
• Est. Priority Date: 02/21/2008
• Status: Active Grant

• Current Assignee: University Of Kentucky Research Foundation (University of Kentucky)
• Original Assignee: University Of Kentucky Research Foundation (University of Kentucky)
• Inventors: Ambati, Jayakrishna
• Primary Examiner(s): GIBBS, TERRA C
• Application Number:
  US12/918,548
• Publication Number:
  US 20110097390A1
• Time in Patent Office: 1,600 Days
• Field of Search: 514/44, 536/24.1, 536/24.5
• US Class Current: 514/44.A
• CPC Class Codes: A01K 2217/056 due to mutation of coding region of the transgene (dominant negative)
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2267/0337 Animal models for infectious diseases
• CPC Class Codes: A01K 2267/0387 Animal model for diseases of the immune system
• CPC Class Codes: A01K 67/0276 Knock-out vertebrates
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 15/06 Antiabortive agents; Labour repressants
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 27/02 Ophthalmic agents
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 33/10 Anthelmintics
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/117 Nucleic acids having immunomodulatory properties, e.g. containing CpG-motifs
• CPC Class Codes: C12N 15/8509 for producing genetically modified animals, e.g. transgenic
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/18 acting by a non-sequence specific mechanism
• CPC Class Codes: C12N 2310/3515 Lipophilic moiety, e.g. cholesterol
• CPC Class Codes: C12N 2310/53 partially self-complementary or closed
• CPC Class Codes: C12N 2320/53 reducing unwanted side-effects

Abstract

Provided are methods and compositions for the treatment or prevention of macular degeneration or other diseases or disorders associated with activation of TLR3. Administration of double stranded RNAs having a length of 22 nucleotides or less treats or prevents macular degeneration or other diseases or disorders associated with activation of TLR3 due to the ability of the RNAs to bind to but not activate TLR3. Furthermore, all double stranded RNAs (both targeted and non-targeted) of 22 nucleotides or less in length can bind to but not activate TLR3 and thereby treat or prevent such conditions. Also provided of a method for increasing the specificity of a desire siRNA target knockdown, the method comprising administering an amount of a target siRNA sufficient to knockdown a target gene and an amount of a double stranded RNA of 22 nucleotides or less which prevents the target siRNA from activating TLR3.

Claims

1. A method of treating or preventing macular degeneration comprising exposing a retinal or choroidal cell to a toll-like receptor 3 (TLR3)-antagonistic effective amount of a double-stranded RNA of 21 nucleotides or shorter length which binds to but does not activate the activity of a TLR3.

2. The method of claim 1, wherein macular degeneration is treated or prevented by inhibiting the geographic atrophy of wet or dry macular degeneration.

3. The method of claim 1, wherein the double-stranded RNA is a naked double-stranded RNA or a double-stranded RNA having an O-methyl group at one or more 2′

-positions.

4. The method of claim 1, wherein the double-stranded RNA has 5-15 nucleotides.

5. The method of claim 4, wherein the double-stranded RNA has 7-11 nucleotides.

6. The method of claim 1, wherein the double-stranded RNA has 2-nucleotide 3′

overhangs on the sense strand and the antisense strand.

7. The method of claim 1, wherein the double-stranded RNA is conjugated to cholesterol.

8. The method of claim 1, wherein the double-stranded RNA is encapsulated in a pharmaceutically acceptable carrier selected from the group consisting of liposomes, SNALPS, nanoparticles, and dendrimers.

9. The method of claim 1, wherein the double-stranded RNA is modified to prevent RNAse degradation.

10. The method of claim 9, wherein the double-stranded RNA is modified with 2′

O-Me.

11. The method of claim 1, wherein the double-stranded RNA is a sequence-nonspecific double-stranded RNA.

12. The method of claim 1, wherein the double-stranded RNA is a sequence-specific double-stranded RNA.

13. The method of claim 1, wherein the retinal cell is a retinal pigmented epithelial cell.

14. The method of claim 1, wherein the choroidal cell is a choroidal endothelial cell.

15. The method of claim 1, wherein the double-stranded RNA is administered via a sustained release device or composition which is implanted in the vitreous humor, aqueous humor, on the sclera, in the sclera, in the suprachoroidal space, or in the subretinal space.

16. A method of treating or preventing a disease or disorder associated with activation of TLR3 comprising exposing a subject in need thereof to a toll-like receptor 3 (TLR3)-antagonistic effective amount of a double-stranded RNA of 21 nucleotides or shorter length which binds to but does not activate the activity of a TLR3.

17. The method of claim 16, wherein the disease or disorder is selected from the group consisting of diabetes, a liver disease, a neurodegenerative disease, a viral infection, a helminthic infection, and prevention of spontaneous abortion.

18. A method for increasing the specificity of a desired siRNA target knockdown, the method comprising administering an amount of a target siRNA sufficient to knockdown a target gene and an amount of a double stranded RNA of 21 nucleotides or less which prevents the target siRNA from activating TLR3.