Nucleic acid-containing lipid particles and related methods

Patent metadata

Field	Value
Publication number	US-9758795-B2
Application number	US-201213464690-A
Country	US
Kind code	B2
Filing date	May 4, 2012
Priority date	Nov 4, 2009
Publication date	Sep 12, 2017
Grant date	Sep 12, 2017

Abstract

Lipid particles containing a nucleic acid, devices and methods for making the lipid particles, and methods for using the lipid particles.

First claim

The embodiments of the invention in which an exclusive property or privilege is claimed are defined as follows:

1. A lipid particle, comprising:
(a) a substantially solid core consisting of nucleic acid, cationic lipid, and optionally second lipids; and
(b) PEG-lipid surrounding the core,
wherein the lipid particle has a core and a periphery, the core having an area averaged electron density that is not less than 20% of the maximum density of the periphery as measured by cryo transmission electron microscopy, and
wherein the 31P nuclear magnetic resonance spectrum of the lipid particle measured in solution after treatment with 150 mM ammonium acetate does not exhibit a resonance due to the nucleic acid.

2. The particle of claim 1 , comprising from about 30 to about 95 mole percent cationic lipid.

3. The particle of claim 1 , comprising from about 1.0 to about 10 mole percent PEG-lipid.

4. The particle of claim 1 having a diameter from about 15 nm to about 300 nm.

5. A method for introducing a nucleic acid into a cell, comprising contacting a cell with the lipid particle of claim 1 .

6. The particle of claim 1 , wherein the cationic lipid is an ionizable lipid.

7. The particle of claim 1 , wherein the cationic lipid is an amino lipid.

8. The particle of claim 1 , wherein the cationic lipid is selected from the group consisting of DODAC, DOTMA, DDAB, DOTAP, DOTAP.Cl, DC-Chol, DOSPA, DOGS, DOPE, DODAP, DODMA, DODMA, and DMRIE.

9. The particle of claim 1 , wherein the cationic lipid has the formula:

wherein R 1 and R 2 are either the same or different and independently optionally substituted C 10 -C 24 alkyl, optionally substituted C 10 -C 24 alkenyl, optionally substituted C 10 -C 24 alkynyl, or optionally substituted C 10 -C 24 acyl;
R 3 and R 4 are either the same or different and independently optionally substituted C 1 -C 6 alkyl, optionally substituted C 2 -C 6 alkenyl, or optionally substituted C 2 -C 6 alkynyl or R 3 and R 4 may join to form an optionally substituted heterocyclic ring of 4 to 6 carbon atoms and 1 or 2 heteroatoms chosen from nitrogen and oxygen;
R 5 is either absent or present and when present is hydrogen or C 1 -C 6 alkyl;
m, n, and p are either the same or different and independently either 0 or 1 with the proviso that m, n, and p are not simultaneously 0;
q is 0, 1 , 2, 3, or 4; and
Y and Z are either the same or different and independently O, S, or NH.

10. The particle of claim 1 , wherein the PEG-lipid is selected from the group consisting of PEG-modified phosphatidylethanolamines, PEG-modified phosphatidic acids, PEG-modified ceramides, PEG-modified dialkylamines, PEG-modified diacylglycerols, PEG-modified dialkylglycerols.

11. The particle of claim 1 , comprising from about 1 to about 5 mole percent PEG-lipid.

12. The particle of claim 1 , wherein the PEG-lipid is selected from the group consisting of PEG-c-DOMG, PEG-c-DMA, and PEG-c-DMG.

13. The particle of claim 1 , wherein the second lipids are selected from the group consisting of zwitterionic lipids, sterols, and mixtures thereof.

14. The particle of claim 1 , wherein the second lipids are selected from the group consisting of diacylphosphatidylcholines, diacylphosphatidylethanolamines, ceramides, sphingomyelins, dihydrosphingomyelins, cephalins, and cerebrosides.

15. The particle of claim 14 , wherein the diacylphosphatidylcholine is 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC).

16. The particle of claim 13 , wherein the second lipid is a sterol.

17. The particle of claim 16 , wherein the sterol is cholesterol.

18. The particle of claim 1 , wherein the nucleic acid is a DNA, an RNA, a locked nucleic acid, a nucleic acid analog, or a plasmid capable of expressing a DNA or an RNA.

19. The particle of claim 1 , wherein the nucleic acid is ssDNA or dsDNA.

20. The particle of claim 1 , wherein the nucleic acid is mRNA, siRNA, or microRNA.

21. The particle of claim 1 , wherein the nucleic acid is an antisense oligonucleotide.

22. The particle of claim 1 having a diameter from about 30 nm to about 200 nm.

23. A composition, comprising a plurality of lipid particles of claim 1 .

Assignees

• Cullis Pieter R
• Belliveau Nathan M
• Genghis Hansen Carl Lars
• Huft Jens
• Taylor James
• Wild Andre
• Malcolm Stuart
• Hafez Ismail
• Leung Alex
• Walker David
• Univ British Columbia

Inventors

• Cullis Pieter R
• Belliveau Nathan M
• Genghis Hansen Carl Lars
• Huft Jens
• Taylor James
• Wild Andre
• Malcolm Stuart
• Hafez Ismail
• Leung Alex
• Walker David

Classifications

• A61K31/7088

  Compounds having three or more nucleosides or nucleotides · CPC title

• C12N15/111

  General methods applicable to biologically active non-coding nucleic acids · CPC title

• A61K47/22

  Heterocyclic compounds, e.g. ascorbic acid, tocopherol or pyrrolidones · CPC title

• A61K9/1272 Primary

  comprising non-phosphatidyl surfactants as bilayer-forming substances, e.g. cationic lipids or non-phosphatidyl liposomes coated or grafted with polymers (lipids as modifying agents {A61K47/543}) · CPC title

• C12N2320/32

  Special delivery means, e.g. tissue-specific · CPC title