Lipid formulation

• US 8,158,601 B2
• Filed: 06/10/2010
• Issued: 04/17/2012
• Est. Priority Date: 06/10/2009
• Status: Active Grant

• Current Assignee: Arbutus Biopharma Corporation
• Original Assignee: Alnylam Pharmaceuticals Incorporated
• Inventors: Chen, Jianxin, Ansell, Steven, Akinc, Akin, Dorkin, Joseph Robert, Qin, Xiaojun, Cantley, William, Manoharan, Muthiah, Rajeev, Kallanthottathil G., Narayanannair, Jayaprakash K., Jayaraman, Muthusamy
• Primary Examiner(s): VIVLEMORE, TRACY ANN
• Application Number:
  US12/813,448
• Publication Number:
  US 20100324120A1
• Time in Patent Office: 677 Days
• Field of Search: None
• US Class Current: 514/44
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 31/22 of acyclic acids, e.g. pravastatin
• CPC Class Codes: A61K 31/7088 Compounds having three or more nucleosides or nucleotides
• CPC Class Codes: A61K 31/712 Nucleic acids or oligonucleotides having modified sugars, i.e. other than ribose or 2'-deoxyribose
• CPC Class Codes: A61K 31/713 Double-stranded nucleic acids or oligonucleotides
• CPC Class Codes: A61K 38/1709 from mammals
• CPC Class Codes: A61K 38/4846 Factor VII (3.4.21.21); Factor IX (3.4.21.22); Factor Xa (3.4.21.6); Factor XI (3.4.21.27); Factor XII (3.4.21.38)
• CPC Class Codes: A61K 47/14 Esters of carboxylic acids, e.g. fatty acid monoglycerides, medium-chain triglycerides, parabens or PEG fatty acid esters
• CPC Class Codes: A61K 47/549 Sugars, nucleosides, nucleotides or nucleic acids
• CPC Class Codes: A61K 47/551 one of the codrug's components being a vitamin, e.g. niacinamide, vitamin B3, cobalamin, vitamin B12, folate, vitamin A or retinoic acid
• CPC Class Codes: A61K 47/6911 the form being a liposome
• CPC Class Codes: A61K 48/0033 the non-active part being non-polymeric
• CPC Class Codes: A61K 9/1272 with substantial amounts of non-phosphatidyl, i.e. non-acylglycerophosphate, surfactants as bilayer-forming substances, e.g. cationic lipids with cholesterol as the only non-phosphatidyl surfactant A61K9/127; lipids as modifying agent A61K47/543
• CPC Class Codes: A61K 9/1278 Post-loading, e.g. by ion or pH gradient
• 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 9/00 Drugs for disorders of the cardiovascular system
• CPC Class Codes: C07C 227/18 by reactions involving amino or carboxyl groups, e.g. hydrolysis of esters or amides, by formation of halides, salts or esters
• CPC Class Codes: C07C 227/40 Separation; Purification
• CPC Class Codes: C07C 229/12 to carbon atoms of acyclic carbon skeletons
• CPC Class Codes: C07C 29/58 by elimination of halogen, e.g. by hydrogenolysis, splitting-off C07C29/124 takes precedence
• CPC Class Codes: C07D 475/04 : with a nitrogen atom directly attached in position 2
• 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 15/1137 : against enzymes viral enzymes C12N15/1131; receptors C12N15/1138
• CPC Class Codes: C12N 2310/14 : interfering N.A.
• CPC Class Codes: C12N 2310/315 : Phosphorothioates
• CPC Class Codes: C12N 2310/321 : 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 : 2'-R Modification
• CPC Class Codes: C12N 2310/341 : Gapmers, i.e. of the type ===---===
• CPC Class Codes: C12N 2310/346 : having a combination of backbone and sugar modifications
• CPC Class Codes: C12N 2310/351 : Conjugate
• CPC Class Codes: C12N 2310/3515 : Lipophilic moiety, e.g. cholesterol
• CPC Class Codes: C12N 2310/3521 : Methyl
• CPC Class Codes: C12N 2310/3533 : Halogen
• CPC Class Codes: C12N 2320/32 : Special delivery means, e.g. tissue-specific
• CPC Class Codes: Y02A 50/30 : Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Abstract

The invention features a cationic lipid of formula I,

• • an improved lipid formulation comprising a cationic lipid of formula I and corresponding methods of use.

Also disclosed are targeting lipids, and specific lipid formulations comprising such targeting lipids.

Claims

1. A cationic lipid of formula I:

2. A lipid formulation comprising a cationic lipid of claim 1.

3. The lipid formulation of claim 2, comprising 40-65% of cationic lipid of formula I, 5-10% of a neutral lipid, 25-40% of a sterol, and 0.5-10% of a PEG or PEG-modified lipid.

4. The lipid formulation of claim 3, wherein the neutral lipid is selected from DSPC, DPPC, DMPC, POPC, DOPE and SM.

5. The lipid formulation of claim 3, wherein the sterol is cholesterol.

6. The lipid formulation of claim 3, wherein the PEG lipid is PEG-C₁₄ to PEG-C₂₂, PEG-Cer₁₄ to PEG-C₂₀, or PEG-DSPE.

7. The lipid formulation of claim 3, wherein the formulation is prepared by an in-line mixing method.

8. The lipid formulation of claim 3, comprising about 57.5% of cationic lipid of formula I, about 7.5% of the neutral lipid, about 31.5% of the sterol, and about 3.5% of the PEG or PEG-modified lipid.

9. The lipid formulation of claim 8, wherein the formulation is prepared by an extrusion method.

10. The lipid formulation of claim 2, further comprising a therapeutic agent.

11. The lipid formulation of claim 10, wherein the therapeutic agent comprises a nucleic acid.

12. The lipid formulation of claim 11, wherein the nucleic acid is selected from the group consisting of an siRNA, an antisense nucleic acid, a microRNA, an antimicroRNA, an antagomir, a microRNA inhibitor, a microRNA activator, an immune stimulatory nucleic acid or a Ua adaptor.

13. The lipid formulation of claim 12, wherein the ratio of lipid:

nucleic acid is about 3 to about 15.

14. The lipid formulation of claim 13, wherein the ratio of lipid:

nucleic acid about 5 to about 13.

15. The lipid formulation of claim 2, further comprising at least one apolipoprotein.

16. The lipid formulation of claim 15, wherein the apolipotprotein is ApoE, active polymorphic forms, isoforms, variants and mutants, and fragments or truncated forms thereof.

17. The lipid formulation of claim 2, further comprising a targeting lipid.

18. The formulation of claim 17, wherein the targeting lipid comprises N-acetyl galactosamine.

19. The formulation of claim 18, wherein the N-acetyl galactosamide comprises at least a mono-, bi- or a triantennary sugar unit.

20. The formulation of claim 17, wherein said targeting lipid is present in the formulation in a molar amount of from about 0.001% to about 5%.

21. The formulation of claim 17, wherein said targeting lipid is the compound selected from the group consisting of Formula II, Formula III, Formula VI and Formula VII:

22. The lipid formulation of claim 3, comprising about 50% of cationic lipid of formula I, about 10% of the neutral lipid, about 38.5% of the sterol, and about 1.5% of the PEG or PEG-modified lipid.

23. The lipid formulation of claim 3, comprising about 50% of cationic lipid of formula I, about 10% of the neutral lipid, about 35% of the sterol, and about 5% of the PEG or PEG-modified lipid.

24. The lipid formulation of claim 3, comprising about 57.2% of cationic lipid of formula I, about 7.1% of the neutral lipid, about 34.3% of the sterol, and about 1.4% of the PEG or PEG-modified lipid.

25. A method of delivering a therapeutic agent to a cell comprising administering to a subject the lipid formulation of claim 10.

26. The method of claim 25, wherein the therapeutic agent is a dsRNA.

27. The method of claim 26, wherein the target gene is Factor VII.

28. The method of claim 26, further comprising comparing expression of the target gene with a preselected reference value.

29. The method of claim 26, wherein the therapeutic agent is an antisense, siRNA, ribozyme or microRNA.

30. A method of modulating the expression of a target gene in a cell, the method comprising providing to a cell the lipid formulation of claim 10.