Phagocytic cell delivery of RNAI

• US 8,815,818 B2
• Filed: 07/17/2009
• Issued: 08/26/2014
• Est. Priority Date: 07/18/2008
• Status: Active Grant

• Current Assignee: Phio Pharmaceuticals Corp.
• Original Assignee: Rxi Pharmaceuticals Corporation
• Inventors: Samarsky, Dmitry, Woolf, Tod M.
• Primary Examiner(s): WHITEMAN, BRIAN A
• Application Number:
  US13/054,696
• Publication Number:
  US 20120016005A1
• Time in Patent Office: 1,866 Days
• Field of Search: 514/44.A
• US Class Current: 514/44.A
• CPC Class Codes: A61K 47/549 Sugars, nucleosides, nucleotides or nucleic acids
• CPC Class Codes: A61K 47/61 the organic macromolecular compound being a polysaccharide or a derivative thereof
• CPC Class Codes: A61K 47/62 the modifying agent being a protein, peptide or polyamino acid
• CPC Class Codes: A61P 1/00 Drugs for disorders of the alimentary tract or the digestive system
• CPC Class Codes: A61P 11/06 Antiasthmatics
• CPC Class Codes: A61P 19/02 for joint disorders, e.g. arthritis, arthrosis
• CPC Class Codes: A61P 19/10 for osteoporosis
• CPC Class Codes: A61P 25/00 Drugs for disorders of the nervous system
• CPC Class Codes: A61P 25/28 for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
• CPC Class Codes: A61P 31/06 for tuberculosis
• CPC Class Codes: A61P 31/14 for RNA viruses
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/351 Conjugate
• CPC Class Codes: C12N 2320/32 Special delivery means, e.g. tissue-specific

Abstract

The present invention provides a particulate delivery system for delivering an RNAi construct to phagocytic cells such as macrophages, comprising various configurations of a complex comprising a phagocytic cell-targeting moiety and an RNAi construct. The invention further provides methods of making the delivery system, and their uses, such as treating phagocytic cell-associated disease conditions.

Claims

1. A delivery system for delivering an RNAi construct to a phagocytic cell and inhibiting expression of a target gene in the phagocytic cell, comprising an RNAi construct and a phagocytic cell-targeting moiety,

wherein the RNAi construct is a double-stranded RNA (dsRNA) construct of 25-30 nucleotides in length, comprising:

(1) a sense strand having a 5′

-end and a 3′

-end, wherein the 5′

-end 12 nucleotides and the 3′

-end 10 nucleotides of the sense strand comprise 2′

-modified ribose sugars and,

(2) an antisense strand having a 5′

-end and a 3′

-end, which hybridizes to the sense strand,

wherein the phagocytic cell-targeting moiety comprises beta-glucan, and wherein the RNAi construct inhibits expression of the target gene in the phagocytic cell.

2. The delivery system of claim 1, wherein the dsRNA construct is blunt-ended.

3. The delivery system of claim 1, wherein each end of the sense strand comprises non-hydrolyzable internucleotide linkages.

4. The delivery system of claim 1, wherein the 2′

-modified ribose sugars are 2′

-O-alkyl nucleotides, 2′

-deoxy-2′

-fluoro nucleotides, 2′

-deoxy nucleotides, 2′

-H (deoxyribonucleotides), or combination thereof.

5. The delivery system of claim 4, wherein the 2′

-O-alkyl nucleotides are 2′

-O-methyl nucleotides or 2′

-O-allyl nucleotides.

6. The delivery system of claim 1, wherein the antisense strand comprises a 2′

-O-methyl modified nucleotide at the 2nd nucleotide on the 5′

-end of the antisense strand and no other modified nucleotides or the antisense strand comprises at least four consecutive 2′

-O-methyl modified 3′

-end nucleotides with phosphothioate linkages.

7. The delivery system of claim 6, wherein the dsRNA has enhanced target specificity or reduced off-target silencing compared to a similar construct without the 2′

-modification at said position(s).

8. The delivery system of claim 1, wherein the sense strand of the dsRNA comprises a mismatch nucleotide at the 2nd nucleotide from the 3′

-end of the sense strand and/or the last 2nd-8th nucleotides at the 3′

-end of the sense strand mis-match their corresponding antisense strand nucleotides.

9. The delivery system of claim 1, wherein the dsRNA has improved stability in serum and/or cerebral spinal fluid compared to an unmodified dsRNA having the same sequence and/or the dsRNA does not induce interferon response in primary cells.

10. The delivery complex of claim 1, wherein the phagocytic cell-targeting moiety further comprises one or more ligands for phagocytic cell receptor(s).

11. The delivery system of claim 1, wherein the antisense strand is unmodified.

12. A pharmaceutical composition comprising the delivery system of claim 1, and a pharmaceutically acceptable excipient.

13. A method of treating an individual for a disease condition associated with a phagocytic cell defect, comprising administering to the individual a delivery system of claim 1, wherein the RNAi construct antagonizes the function of a target gene causing the disease condition.

14. The method of claim 13, wherein the disease condition is:

macular degeneration, osteoporosis, non-immune inflammation, chronic obstructive pulmonary disease (COPD), Alzheimer'"'"'s disease, sepsis and septic shock, neutrophilic alveolitis, hepatitis, inflammatory bowel disease, ischemia/reperfusion, glomerulonephritis, or rheumatoid arthritis.

15. A method for delivering an RNAi construct to a phagocytic cell in a subject, comprising:

administering a delivery system complex to the subject, wherein the delivery system complex comprises an RNAi construct and a phagocytic cell-targeting moiety,

wherein the RNAi construct is a double-stranded RNA (dsRNA) construct of 19-49 nucleotid in length, comprising;

(1) a sense strand having a 5′

-end and a 3′

-end,

wherein the 5′

-end 12 nucleotides and the 3′

-end 10 nucleotides of the sense strand are 2′

-modified sugars, and,

(2) an antisense strand having a 5′

-end and a 3′

-end, which hybridizes to the sense strand, and

wherein the phagocytic cell-targeting moiety comprises beta-glucan.

16. The method of claim 15, wherein the phagocytic cell-targeting moiety further comprises one or more ligands for phagocytic cell receptor(s).

17. The method of claim 15, wherein the dsRNA construct is blunt-ended.