Lipophilic nucleic acid delivery vehicle and methods of use thereof
View Patent ↗Provided herein are compositions and methods for delivery of nucleic acids to individuals and to cells, including nucleic acid delivery particles that comprising a lipid-binding polypeptide, a lipid bilayer comprising one or more cationic lipids, and a nucleic acid.
1. A nucleic acid delivery particle comprising a lipid-binding polypeptide, a lipid bilayer, and a nucleic acid,
wherein the total lipid content of the lipid bilayer is between about 5% and about 100% of one or more cationic lipids,
wherein the interior of the lipid bilayer comprises a hydrophobic region,
wherein the particle
does not comprise an aqueous core,
is disc-shaped with the hydrophobic edge of the lipid bilayer circumscribed by the lipid-binding polypeptide at the periphery of the particle, and
remains disc-shaped in aqueous solution.
2. A nucleic acid delivery particle according to claim 1 , wherein the particle does not comprise a hydrophilic core.
3. A nucleic acid delivery particle according to claim 1 , wherein the disc shaped particle is between about 10 nm to about 40 nm in diameter.
4. A nucleic acid delivery particle according to claim 1 , wherein the one or more cationic lipids is selected from the group consisting of N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA), amidine, dimethyl-dioctadecyl ammonium bromide (DDAB), dimyristoyltrimethylammonium propane (DMTAP), and 1,2-dioleoyloxy-3-trimethylammoniopropane (DOTAP).
5. A nucleic acid delivery particle according to claim 1 , wherein the total lipid content of the lipid bilayer is between about 0% to about 95% of one or more phospholipids.
6. A nucleic acid delivery particle according to claim 5 , wherein the one or more phospholipids is selected from the group consisting of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG).
7. A nucleic acid delivery particle according to claim 5 , wherein the one or more phospholipids is selected from the group consisting of dipalmitoylphosphatidylcholine (DPPC) and egg phosphatidylcholine.
8. A nucleic acid delivery particle according to claim 1 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
9. A nucleic acid delivery particle according to claim 8 , wherein the nucleic acid associates with the surface of the particle by electrostatic interactions.
10. A nucleic acid delivery particle according to claim 8 , wherein the nucleic acid further comprises a lipophilic conjugate, and wherein the lipophilic conjugate is incorporated into the hydrophobic region of the lipid bilayer.
11. A nucleic acid delivery particle according to claim 10 , wherein the lipophilic conjugate is selected from the group consisting of cholesterol, oleic acid, stearic acid, palmitic acid, myristic acid, and linoleic acid.
12. A nucleic acid delivery particle according to claim 1 , wherein the lipid-binding polypeptide is an apolipoprotein.
13. A nucleic acid delivery particle according to claim 12 , wherein the apolipoprotein is an exchangeable apolipoprotein.
14. A nucleic acid delivery particle according to claim 13 , wherein the apolipoprotein is human apolipoprotein A-I.
15. A nucleic acid delivery particle according to claim 12 , wherein the apolipoprotein is a chimeric apolipoprotein that comprises a functional moiety.
16. A nucleic acid delivery particle according to claim 15 , wherein the functional moiety is a targeting moiety.
17. A nucleic acid delivery particle according to claim 12 , wherein the apolipoprotein has been modified to increase stability of the particle.
18. A nucleic acid delivery particle according to claim 17 , wherein the modification comprises introduction of cysteine residues to form intermolecular or intramolecular disulfide bonds.
19. A nucleic acid delivery particle according to claim 18 , wherein the apolipoprotein is modified to present uncharged amino acids in regions of the apolipoprotein adjacent to the head groups of the one or more cationic lipids.
20. A nucleic acid delivery particle according to claim 12 , wherein the lipid-binding polypeptide is a peptide.
21. A nucleic acid delivery particle according to claim 20 , wherein the lipid-binding polypeptide is an amphipathic peptide.
22. A nucleic acid delivery particle according to claim 20 , wherein the peptide has been modified to increase stability of the particle.
23. A nucleic acid delivery particle according to claim 22 , wherein the peptide is modified to present uncharged amino acids in regions of the peptide adjacent to the head groups of the one or more cationic lipids.
24. A pharmaceutical composition for the delivery of a nucleic acid comprising a nucleic acid delivery particle according to claim 1 and a pharmaceutically acceptable carrier.
25. A pharmaceutical composition according to claim 24 , wherein the composition is formulated for controlled release.
26. A method for in vivo administration of a nucleic acid, comprising administering an effective amount of the pharmaceutical composition of claim 24 to an individual.
27. A method according to claim 26 , wherein the pharmaceutical composition comprises a therapeutically effective amount of the nucleic acid.
28. A method according to claim 27 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
29. A method according to claim 26 , wherein the administration is parenteral.
30. A method according to claim 29 , wherein the parenteral administration is selected from the group consisting of intravenous, intramuscular, transmucosal, and intrathecal.
31. A method according to claim 26 , wherein the composition is administered as an aerosol.
32. A method according to claim 26 , wherein the composition is formulated for controlled release.
33. A process for formulating a nucleic acid delivery particle according to claim 1 , the process comprising:
(1) contacting bilayer-forming lipid vesicles with a nucleic acid to form a bilayer-forming lipid vesicle-nucleic acid mixture, wherein about 5% to about 100% of the total lipid content of the vesicles is one or more cationic lipids, and
(2) contacting the bilayer-forming lipid vesicle-nucleic acid mixture with a lipid-binding polypeptide.
34. A process according to claim 33 , wherein the one or more cationic lipids is selected from the group consisting of N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA), amidine, dimethyl-dioctadecyl ammonium bromide (DDAB), dimyristoyltrimethylammonium propane (DMTAP), and 1,2-dioleoyloxy-3-trimethylammoniopropane (DOTAP).
35. A process according to claim 33 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
36. A process according to claim 35 , wherein the nucleic acid is solubilized in dimethylsulfoxide (DMSO) prior to contacting the bilayer-forming lipid vesicles.
37. A process for formulating a nucleic acid delivery particle according to claim 1 , said process comprising the steps of:
(1) forming an aqueous dispersion of lipid vesicles, wherein said lipid vesicles comprise bilayer-forming lipids and about 5% to about 100% of total lipid content of the vesicles is one or more cationic lipids;
(2) adding a nucleic acid to the lipid vesicle dispersion to form a lipid vesicle-nucleic acid mixture;
(3) adding a lipid-binding polypeptide to the lipid vesicle-nucleic acid mixture to form a lipid-nucleic acid-lipid-binding polypeptide mixture; and
(4) incubating the mixture formed in step (3).
38. A process according to claim 37 , wherein the one or more cationic lipids is selected from the group consisting of N-[1-(2,3-dioleyloxy)propyl]-N,N,N-trimethylammonium chloride (DOTMA), amidine, dimethyl-dioctadecyl ammonium bromide (DDAB), dimyristoyltrimethylammonium propane (DMTAP), and 1,2-dioleoyloxy-3-trimethylammoniopropane (DOTAP).
39. A process according to claim 37 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
40. A process for formulating a nucleic acid delivery particle according to claim 37 , wherein the process further comprises sonicating the mixture of step (4).
41. A process according to claim 39 , wherein the nucleic acid is solubilized in DMSO prior to addition to the lipid vesicle dispersion.
42. A nucleic acid delivery particle prepared according to the process of claim 33 .
43. A nucleic acid delivery particle according to claim 42 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
44. A nucleic acid delivery particle prepared according to the process of claim 37 .
45. A nucleic acid delivery particle according to claim 44 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
46. A pharmaceutical composition comprising a nucleic acid delivery particle according to claim 42 and a pharmaceutically acceptable carrier.
47. A pharmaceutical composition according to claim 46 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
48. A pharmaceutical composition comprising a nucleic acid delivery particle according to claim 44 and a pharmaceutically acceptable carrier.
49. A pharmaceutical composition according to claim 48 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
50. A kit comprising a pharmaceutical composition according to claim 24 and instructions for use in a method for administering a nucleic acid to an individual.
51. A kit comprising a pharmaceutical composition according to claim 46 and instructions for use in a method for administering a nucleic acid to an individual.
52. A kit comprising a pharmaceutical composition according to claim 48 and instructions for use in a method for administering a nucleic acid to an individual.
53. A composition for delivery of a nucleic acid to an individual, comprising a nucleic acid delivery particle of claim 1 and a carrier.
54. A composition according to claim 53 , wherein the individual is a plant or an insect.
55. A composition according to claim 53 , wherein the nucleic acid is a short interfering RNA (siRNA), a short hairpin RNA (shRNA), a micro RNA, an antisense RNA, an antisense DNA, an aptamer, a ribozyme, or a plasmid.
56. A method for delivering a nucleic acid to a cell comprising contacting the cell with a nucleic acid delivery particle according to claim 1 .