Compositions for targeted delivery of siRNA

• US 9,345,775 B2
• Filed: 03/18/2015
• Issued: 05/24/2016
• Est. Priority Date: 02/24/2010
• Status: Active Grant

• Current Assignee: Arrowhead Pharmaceuticals, Inc.
• Original Assignee: Arrowhead Madison Inc (Arrowhead Pharmaceuticals, Inc.)
• Inventors: Lewis, David L, Rozema, David B, Wakefield, Darren H
• Primary Examiner(s): ANGELL, JON E
• Application Number:
  US14/661,947
• Publication Number:
  US 20150202298A1
• Time in Patent Office: 433 Days
• US Class Current: 1/1
• CPC Class Codes: A61K 31/713 Double-stranded nucleic acids or oligonucleotides
• CPC Class Codes: A61K 47/32 Macromolecular compounds obtained by reactions only involving carbon-to-carbon unsaturated bonds, e.g. carbomers , poly(meth)acrylates, or polyvinyl pyrrolidone
• CPC Class Codes: A61K 47/549 Sugars, nucleosides, nucleotides or nucleic acids
• CPC Class Codes: A61K 47/60 the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
• CPC Class Codes: A61K 47/65 Peptidic linkers, binders or spacers, e.g. peptidic enzyme-labile linkers
• CPC Class Codes: A61K 48/0041 the non-active part being polymeric
• CPC Class Codes: A61K 9/08 Solutions composition of solutions A61K47/00
• CPC Class Codes: A61P 1/16 for liver or gallbladder disorders, e.g. hepatoprotective agents, cholagogues, litholytics
• CPC Class Codes: A61P 3/00 Drugs for disorders of the metabolism of the blood or the extracellular fluid A61P7/00
• CPC Class Codes: A61P 3/06 Antihyperlipidemics
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: A61P 31/12 Antivirals
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/87 Introduction of foreign genetic material using processes not otherwise provided for, e.g. co-transformation
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/141 MicroRNAs, miRNAs
• CPC Class Codes: C12N 2310/3515 Lipophilic moiety, e.g. cholesterol
• CPC Class Codes: C12N 2320/32 Special delivery means, e.g. tissue-specific

Abstract

The present invention is directed compositions for targeted delivery of RNA interference (RNAi) polynucleotides to hepatocytes in vivo. Targeted RNAi polynucleotides are administered together with co-targeted delivery polymers. Delivery polymers provide membrane penetration function for movement of the RNAi polynucleotides from outside the cell to inside the cell. Reversible modification provides physiological responsiveness to the delivery polymers.

Claims

1. A composition for delivering an RNA interference polynucleotide to a liver cell in vivo comprising:

2. The composition of claim 1 wherein the RNA interference polynucleotide is selected from the group consisting of:

DNA, RNA, dsRNA, siRNA, and miRNA.

3. The composition of claim 1 wherein the liver cell consists of a hepatocyte.

4. The composition of claim 1 wherein the membrane active polyamine contains two or more different monomers.

5. The composition of claim 4 wherein the membrane active polyamine is composed of amine-containing monomers and lower hydrophobic group-containing monomers wherein said lower hydrophobic groups contain 1-6 carbon atoms.

6. The composition of claim 5 wherein said lower hydrophobic groups contain 2-6 carbon atoms.

7. The composition of claim 6 wherein the lower hydrophobic group consists of a butyl group.

8. The composition of claim 4 wherein the membrane active polyamine is composed of amine-containing monomers, and lower hydrophobic group-containing monomers, and higher hydrophobic group-containing monomers wherein said lower hydrophobic groups contain 1-6 carbon atoms and said lower hydrophobic groups contain 12-36 carbon atoms.

9. The composition of claim 8 wherein the lower hydrophobic group consists of a butyl group and the higher hydrophobic group consists of an octadecyl or dodecyl group.

10. The composition of claim 1 wherein the membrane active polyamine is soluble in water.

11. The composition of claim 1 wherein the reversibly masked membrane active polyamine

12. The composition of claim 1 wherein the membrane active polyamine is a random copolymer.

13. The composition of claim 12 wherein the random copolymer is selected from the group consisting of poly(vinyl ether) and poly(acrylate).

14. The composition of claim 8 wherein the amine-containing monomers, lower hydrophobic group-containing monomers, and higher hydrophobic group-containing monomers are present in a ratio of 4-8 amine-containing monomers:

3-5 lower hydrophobic group-containing monomers;

1 higher hydrophobic group-containing monomers.

15. The composition of claim 1 wherein the value y+z is greater than 70% of the number of amines on the membrane active polyamine.

16. The composition of claim 15 wherein the value y+z is greater than 80% of the number of amines on the membrane active polyamine.

17. The composition of claim 1 wherein the composition is provided in a pharmaceutically acceptable carrier or diluent.

18. The composition of claim 1 wherein N is linked to A via a physiologically labile linkage L¹.

19. The composition of claim 18 wherein L¹ is a physiologically labile covalent linkage that is orthogonal to L².

20. The composition of claim 1 wherein the ratio of galactose derivative to PEG linked to the membrane active polyamine is 1 to 0.5-2.

21. The composition of claim 1 wherein the galactose derivative consists of an N-acetylgalactosamine.

22. The composition of claim 1 wherein the hydrophobic group comprises cholesterol.

23. The composition of claim 1 wherein the hydrophobic group comprises a cholesterol derivative.

24. The composition of claim 1 wherein P-(L²-M¹) comprises the structure represented by:

25. A composition for delivering an oligonucleotide to a liver cell in vivo comprising:

a) an oligonucleotide covalently linked to a hydrophobic group having at least 20 carbon atoms; and

,

b) a reversibly masked and targeted amphipathic polymer comprising a polymer synthesized from amine-containing monomers and lower hydrophobic group-containing monomers to which a plurality of galactose derivatives are linked to said polymer via pH labile disubstituted maleamate bonds and wherein cleavage of said pH labile disubstituted maleamate bonds yields amine groups thereby generating a membrane active polyamine.

26. The composition of claim 25 wherein a plurality of polyethylene glycol moieties are linked to said polymer via pH labile disubstituted maleamate bonds.

27. A method of manufacturing an RNA oligonucleotide delivery composition comprising:

a) forming a membrane active polyamine;

b) forming a first masking agent comprising a charge neutral disubstituted maleic anhydride containing a galactose derivative;

c) optionally forming a second masking agent comprising a charge neutral disubstituted maleic anhydride containing a polyethylene glycol;

d) reversibly inhibiting membrane activity of the membrane active polyamine wherein the inhibiting consists of modifying 50% or more of the amines on the polyamine by reacting the polyamine with a plurality of first masking agents and optionally at least one second masking agent thereby linking a plurality of galactose derivatives and optionally at least one polyethylene glycol to the membrane active polymer via physiologically pH-labile disubstituted maleamate linkages; and

,

e) linking the RNA interference polynucleotide to a hydrophobic group having at least 20 carbon atoms;

f) providing the RNA interference polynucleotide and the reversibly inhibited membrane active polyamine in solution suitable for administration in vivo.