Lipid nanoparticles for delivery of nucleic acids and methods of use thereof
The present disclosure provides for improved compositions of ionizable lipid nanoparticles for the delivery of therapeutic nucleic acids to cells. Anionic phospholipids, including phosphatidylserine and phosphatidylglycerol are included in the lipid nanoparticles to increase the transfection efficiency in human dendritic cells. The further incorporation of mono-unsaturated alkyl chain analogs in dimethylaminopropyl-dioxolane or heterocyclic ketal ionizable lipids in the formulation demonstrated high levels of transfection in human dendritic cells, compared to other ionizable lipids in the same family, and demonstrated good stability to oxidative damage. Finally, the use of an ammonium salt of phosphatidylserine allows for the efficient production of PS-targeted LNPs.
1. A nucleic acid lipid nanoparticle (LNP) composition comprising:
a. a nucleic acid;
b. an ionizable cationic lipid at a N/P ratio of 3 to 8 relative to the nucleic acid in a total amount of 40-65 mol % of the total lipid content of the LNP composition;
c. a sterol in a total amount of 25-45 mol % of the total lipid content of the LNP composition;
d. one or more phospholipids in a total amount of phospholipids of 5-25 mol % of the total lipid content of the LNP composition, and comprising a phosphatidyl-L-serine lipid in a total amount of 2.5-10 mol % of the total lipid content of the LNP composition; and
e. a conjugated lipid in a total amount of 0.5-2.5 mol % of the total lipid content of the LNP composition.
2. The composition of claim 1 , wherein the sterol is cholesterol.
3. The composition of claim 2 , wherein the one or more phospholipids comprise phospholipids having mismatched acyl chain lengths.
4. The composition of claim 3 , wherein the phosphatidyl-L-serine lipid is dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS).
5. The composition of claim 4 , wherein the one or more phospholipids comprise a phospholipid selected from the group consisting of: distearoylphosphatidylcholine (DSPC) and hydrogenated soy phosphatidylcholine (HSPC).
6. The composition of claim 5 , wherein the one or more phospholipids comprise distearoylphosphatidylcholine (DSPC) and the phosphatidyl-L-serine.
7. The composition of claim 3 , wherein the ionizable cationic lipid is 3-((S)-2,2-di((Z)-octadec-9-en-1-yl)-1,3-dioxolan-4-yl)-N,N-dimethylpropan-1-amine (KC3-OA).
8. The composition of claim 6 , wherein the ionizable cationic lipid is 3-((S)-2,2-di((Z)-octadec-9-en-1-yl)-1,3-dioxolan-4-yl)-N,N-dimethylpropan-1-amine (KC3-OA).
9. The composition of claim 8 , wherein the one or more phospholipids consist of distearoylphosphatidylcholine (DSPC) and dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS).
10. The composition of claim 1 , wherein the conjugated lipid is a PEG-containing conjugated lipid, and wherein the PEG-containing conjugated lipid is selected from the group consisting of: PEG(2000)-dimyristoylglycerol (PEG-DMG), 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000] (PEG-DLPE), and PEG(2000)-dilauroylglycerol (PEG-DLG).
11. The composition of claim 2 , wherein:
a. the nucleic acid is mRNA;
b. the one or more phospholipids comprise dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS) and one or more phospholipids selected from the group consisting of:
distearoylphosphatidylcholine (DSPC), and hydrogenated soy phosphatidylcholine (HSPC); and
c. the conjugated lipid comprises a polyethylene glycol (PEG).
12. The composition of claim 11 , wherein the conjugated lipid is PEG(2000)-dimyristoylglycerol (PEG-DMG).
13. The composition of claim 12 , wherein the ionizable cationic lipid is 3-((S)-2,2-di((Z)-octadec-9-en-1-yl)-1,3-dioxolan-4-yl)-N,N-dimethylpropan-1-amine (KC3-OA).
14. The composition of claim 13 , wherein the one or more phospholipids consist of distearoylphosphatidylcholine (DSPC) and dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS).
15. The composition of claim 4 , wherein the dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS) is an ammonium salt of (L-Serine)DPPS.
16. The composition of claim 1 , wherein
a. the nucleic acid is mRNA;
b. the sterol is a cholesterol sterol in a total amount of 25-45 mol % of the total lipid content of the LNP composition;
c. the ionizable cationic lipid is at a N/P ratio of 3 to 8 relative to the nucleic acid in a total amount of 40-65 mol % of the total lipid content of the LNP composition;
d. the one or more phospholipids is in a total amount of phospholipids of 5-25 mol % of the total lipid content of the LNP composition, the one or more phospholipids consisting of:
i. dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS) lipid in a total amount of 2.5-10 mol % of the total lipid content of the LNP composition; and
ii. a distearoylphosphatidylcholine (DSPC) phospholipid in a total amount of 5-mol % of the total lipid content of the LNP composition; and
e. the conjugated lipid is a PEG-containing conjugated lipid in a total amount of 0.5-2.5 mol % of the total lipid content of the LNP composition.
17. The composition of claim 16 , wherein the PEG-containing conjugated lipid is selected from the group consisting of: PEG(2000)-dimyristoylglycerol (PEG-DMG), 1,2-dilauroyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000](PEG-DLPE), and PEG(2000)-dilauroylglycerol (PEG-DLG).
18. A nucleic acid lipid nanoparticle (LNP) human vaccine composition comprising:
a. a nucleic acid;
b. an ionizable cationic lipid at a N/P ratio of 3 to 8 relative to the nucleic acid in a total amount of 40-65 mol % of the total lipid content of the LNP composition;
c. a sterol in a total amount of 25-45 mol % of the total lipid content of the LNP composition; and
d. one or more phospholipids in a total amount of phospholipids of 5-25 mol % of the total lipid content of the LNP composition, and comprising a phosphatidylglycerol (PG) in a total amount of 1.0-10 mol % of the total lipid content of the LNP composition; and
e. a conjugated lipid in a total amount of 0.5-2.5 mol % of the total lipid content of the LNP composition.
19. A nucleic acid lipid nanoparticle (LNP) composition comprising an ionizable cationic lipid in a total amount of 40-65 mol % of the total lipid content of the LNP composition, wherein the ionizable cationic lipid is selected from the group consisting of: KC3-PA, KC3-C17 (8:1), and KC3-C15 (C8:1), KC3-OA, and KC3-01
20. The composition of claim 1 , wherein the ionizable cationic lipid is 3-((S)-2,2-di((Z)-octadec-9-en-1-yl)-1,3-dioxolan-4-yl)-N,N-dimethylpropan-1-amine (KC3-OA); and
the lipid nanoparticle (LNP) composition comprises:
a. the nucleic acid;
b. the ionizable cationic lipid at a N/P ratio of 5 to 6 relative to the nucleic acid in a total amount of 45-55 mol % of the total lipid content of the LNP composition;
c. cholesterol in a total amount of 30-40 mol % of the total lipid content of the LNP composition;
d. the one or more phospholipids in a total amount of phospholipids of 10-20 mol % of the total lipid content of the LNP composition, the one or more phospholipids consisting of:
i. dipalmitoylphosphatidyl-L-serine ((L-serine)DPPS) lipid in a total amount of 2.5-7.5 mol % of the total lipid content of the LNP composition; and
ii. a distearoylphosphatidylcholine (DSPC) phospholipid in a total amount of 5-mol % of the total lipid content of the LNP composition; and
e. the conjugated lipid in a total amount of 0.5-2.5 mol % of the total lipid content of the LNP composition.
21. The composition of claim 20 , wherein
a. the nucleic acid is mRNA; and
b. the conjugated lipid is a PEG-containing conjugated lipid, and wherein the PEG-containing conjugated lipid is selected from the group consisting of: PEG(2000)-dimyristoylglycerol (PEG-DMG), and PEG(2000)-dilauroylglycerol (PEG-DLG).
22. The composition of claim 21 , wherein the conjugated lipid is PEG(2000)-dilauroylglycerol (PEG-DLG).
23. The composition of claim 21 , wherein the conjugated lipid is PEG(2000)-dimyristoylglycerol (PEG-DMG).
24. A nucleic acid lipid nanoparticle (LNP) composition comprising a nucleic acid, a KC3 ionizable cationic lipid and one or more targeting phospholipids selected from the group consisting of: a phosphatidyl-L-serine lipid and a phosphatidylglycerol lipid, wherein the KC-3 ionizable cationic lipid has the chemical structure:
and R 1 is alkyl group of C 15 -C 19 containing one or two olefins.
25. The composition of claim 24 , wherein the nucleic acid is mRNA encoding a coronavirus or portion thereof.
26. The composition of claim 19 , wherein the ionizable cationic lipid is KC3-OA.
27. The composition of claim 1 , wherein the LNP composition consists of:
a. KC3-OA ionizable cationic lipid;
b. an antigen generating mRNA nucleic acid;
c. cholesterol;
d. one or more phospholipids comprising one or more targeting phospholipids selected from the group consisting of: a phosphatidyl-L-serine lipid, and a phosphatidylglycerol lipid; and
e. a conjugated PEG lipid.
28. The composition of claim 27 , wherein the composition is a LNP vaccine composition further comprising the LNP composition in an aqueous buffer.