Lipid-encapsulated polyanionic nucleic acid
Methods for the preparation of a lipid-nucleic acid composition are provided. According to the methods, a mixture of lipids containing a protonatable or deprotonatable lipid, for example an amino lipid and a lipid such as a PEG- or Polyamide oligomer-modified lipid is combined with a buffered aqueous solution of a charged therapeutic agent, for example polyanionic nucleic acids, to produce particles in which the therapeutic agent is encapsulated in a lipid vesicle. Surface charges on the lipid particles are at least partially neutralized to provide surface-neutralized lipid-encapsulated compositions of the therapeutic agents. The method permits the preparation of compositions with high ratios of therapeutic agent to lipid and with encapsulation efficiencies in excess of 50%.
1. A lipid-encapsulated polyanionic nucleic acid composition comprising lipid-nucleic acid particles, wherein each lipid-nucleic acid particle is formed from a lipid layer surrounding and encapsulating a central region containing a polyanionic nucleic acid, wherein the lipid layer comprises a titratable lipid comprising a protonatable group having a pKa of from 4 to 11 and a modified lipid selected from among lipids which prevent particle aggregation during lipid-nucleic acid particle formation.
2. The composition of claim 1 , wherein the titratable lipid is an amino lipid.
3. The composition of claim 1 , wherein the titratable lipid is DODAP.
4. The composition of claim 1 , wherein the modified lipid is a polyethylene glycol-modified or polyamide oligomer-modified lipid.
5. The composition of claim 4 , wherein the modified lipid is a polyethylene glycol-modified ceramide.
6. The composition of claim 5 , wherein the modified lipid is PEG-CerC14 or PEG-CerC20.
7. The composition of claim 6 , wherein the titratable lipid is an amino lipid.
8. The composition of claim 6 , wherein the titratable lipid is DODAP.
9. The composition of claim 1 , wherein the polyanionic nucleic acid is an oligonucleotide with a length of from 5 to 50 nucleotides.
10. The composition of claim 9 , wherein the titratable lipid is an amino lipid.
11. The composition of claim 9 , wherein the titratable lipid is DODAP.
12. The composition of claim 11 , wherein the modified lipid is PEG-CerC14 or PEG-CerC20.
13. The composition of claim 9 , wherein the polyanionic nucleic acid contains exclusively phosphodiester linkages.
14. The composition of claim 13 , wherein the titratable lipid is an amino lipid.
15. The composition of claim 13 , wherein the titratable lipid is DODAP.
16. The composition of claim 15 , wherein the modified lipid is PEG-CerC14 or PEG-CerC20.
17. The composition of claim 9 , wherein the polyanionic nucleic acid contains at least one modified intemucleotidic linkage selected from the group consisting of phosphorothjioate, phosphorothioate, phosphoroselenate, boranophosphate, methylphosphate and O-alkyl phosphotriester linkages.
18. The composition of claim 1 , wherein the lipid-nucleic acid particles are sized to a mean diameter of from about 70 to 200 nm.
19. The composition of claim 18 , wherein the lipid-nucleic acid particles are sized to a mean diameter of from about 90 to 130 nm.
20. The composition of claim 1 , wherein the protonable group of the titratable lipid has a pKa of from 4 to 7.
21. The composition of claim 1 , wherein the weight to weight ratio of the polyanionic nucleic acid to the lipids in the lipid-nucleic acid particle is greater than 0.1.