Lipid formulations for nucleic acid delivery
View Patent ↗The present invention provides novel, serum-stable lipid particles comprising one or more active agents or therapeutic agents, methods of making the lipid particles, and methods of delivering and/or administering the lipid particles. More particularly, the present invention provides serum-stable nucleic acid-lipid particles (SNALP) comprising a nucleic acid (e.g., one or more interfering RNA molecules), methods of making the SNALP, and methods of delivering and/or administering the SNALP (e.g., for the treatment of cancer). In particular embodiments, the present invention provides tumor-directed lipid particles that preferentially target solid tumors. The tumor-directed formulations of the present invention are capable of preferentially delivering a payload such as a nucleic acid to cells of solid tumors compared to non-cancerous cells.
1. A nucleic acid-lipid particle comprising:
(a) an siRNA that silences polo-like kinase 1 (PLK-1) gene expression, wherein the siRNA consists of one the following pairs of sense and antisense strand sequences: SEQ ID NOS:3 and 16; SEQ ID NOS:3 and 19; SEQ ID NOS:3 and 21; SEQ ID NOS:3 and 22; SEQ ID NOS:3 and 23; SEQ ID NOS:3 and 24; SEQ ID NOS:3 and 25; SEQ ID NOS:3 and 26; SEQ ID NOS:3 and 27; SEQ ID NOS:3 and 28; SEQ ID NOS:3 and 29; SEQ ID NOS:3 and 30; SEQ ID NOS:3 and 31; SEQ ID NOS:3 and 32; SEQ ID NOS:3 and 33; SEQ ID NOS:4 and 16; SEQ ID NOS:4 and 19; SEQ ID NOS:4 and 20; SEQ ID NOS:4 and 21; SEQ ID NOS:4 and 22; SEQ ID NOS:4 and 23; SEQ ID NOS:4 and 24; SEQ ID NOS:4 and 25; SEQ ID NOS:4 and 26; SEQ ID NOS:4 and 27; SEQ ID NOS:4 and 28; SEQ ID NOS:4 and 29; SEQ ID NOS:4 and 30; SEQ ID NOS:4 and 31; SEQ ID NOS:4 and 32; SEQ ID NOS:4 and 33; SEQ ID NOS:9 and 16; SEQ ID NOS:9 and 19; SEQ ID NOS:9 and 20; SEQ ID NOS:9 and 21; SEQ ID NOS:9 and 22; SEQ ID NOS:9 and 23; SEQ ID NOS:9 and 24; SEQ ID NOS:9 and 25; SEQ ID NOS:9 and 26; SEQ ID NOS:9 and 27; SEQ ID NOS:9 and 28; SEQ ID NOS:9 and 29; SEQ ID NOS:9 and 30; SEQ ID NOS:9 and 31; SEQ ID NOS:9 and 32; SEQ ID NOS:9 and 33; SEQ ID NOS:10 and 16; SEQ ID NOS:10 and 19; SEQ ID NOS:10 and 20; SEQ ID NOS:10 and 21; SEQ ID NOS:10 and 22; SEQ ID NOS:10 and 23; SEQ ID NOS:10 and 24; SEQ ID NOS:10 and 25; SEQ ID NOS:10 and 26; SEQ ID NOS:10 and 27; SEQ ID NOS:10 and 28; SEQ ID NOS:10 and 29; SEQ ID NOS:10 and 30; SEQ ID NOS:10 and 31; SEQ ID NOS:10 and 32; and SEQ ID NOS:10 and 33;
(b) a cationic lipid comprising from about 50 mol % to about 65 mol % of the total lipid present in the particle;
(c) a non-cationic lipid comprising from about 25 mol % to about 45 mol % of the total lipid present in the particle; and
(d) a conjugated lipid that inhibits aggregation of particles comprising from about 5 mol % to about 10 mol % of the total lipid present in the particle.
2. The nucleic acid-lipid particle of claim 1 , wherein the cationic lipid comprises 1,2-dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 1,2-dilinolenyloxy-N,N-dimethylaminopropane (DLenDMA), 1,2-di-γ-linolenyloxy-N,N-dimethylaminopropane (γ-DLenDMA), 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-K-C2-DMA), 2,2-dilinoleyl-4-dimethylaminomethyl-[1,3]-dioxolane (DLin-K-DMA), or a mixture thereof.
3. The nucleic acid-lipid particle of claim 1 , wherein the cationic lipid comprises from about 50 mol % to about 60 mol % of the total lipid present in the particle.
4. The nucleic acid-lipid particle of claim 1 , wherein the non-cationic lipid is a phospholipid.
5. The nucleic acid-lipid particle of claim 1 , wherein the non-cationic lipid comprises a mixture of a phospholipid and cholesterol or a mixture of a phospholipid and a cholesterol derivative.
6. The nucleic acid-lipid particle of claim 5 , wherein the phospholipid is selected from the group consisting of dipalmitoylphosphatidylcholine (DPPC), distearoylphosphatidylcholine (DSPC), and a mixture thereof.
7. The nucleic acid-lipid particle of claim 5 , wherein the phospholipid comprises from about 5 mol % to about 10 mol % of the total lipid present in the particle.
8. The nucleic acid-lipid particle of claim 5 , wherein the cholesterol comprises from about 25 mol % to about 35 mol % of the total lipid present in the particle.
9. The nucleic acid-lipid particle of claim 1 , wherein the non-cationic lipid is a mixture of DPPC and cholesterol.
10. The nucleic acid-lipid particle of claim 1 , wherein the non-cationic lipid is cholesterol or a cholesterol derivative.
11. The nucleic acid-lipid particle of claim 10 , wherein the cholesterol comprises from about 30 mol % to about 40 mol % of the total lipid present in the particle.
12. The nucleic acid-lipid particle of claim 1 , wherein the conjugated lipid that inhibits aggregation of particles comprises a polyethyleneglycol (PEG)-lipid conjugate.
13. The nucleic acid-lipid particle of claim 12 , wherein the PEG has an average molecular weight of from about 550 daltons to about 1000 daltons.
14. The nucleic acid-lipid particle of claim 12 , wherein the PEG has an average molecular weight of about 750 daltons.
15. The nucleic acid-lipid particle of claim 12 , wherein the PEG-lipid conjugate is member selected from the group consisting of a PEG-diacylglycerol (PEG-DAG) conjugate, a PEG dialkyloxypropyl (PEG-DAA) conjugate, a PEG-phospholipid conjugate, a PEG-ceramide (PEG-Cer) conjugate, and a mixture thereof.
16. The nucleic acid-lipid particle of claim 15 , wherein the PEG-lipid conjugate comprises a PEG-DAA conjugate.
17. The nucleic acid-lipid particle of claim 16 , wherein the PEG-DAA conjugate comprises a PEG-dimyristyloxypropyl (PEG-DMA) conjugate.
18. The nucleic acid-lipid particle of claim 1 , wherein the conjugated lipid that inhibits aggregation of particles comprises from about 6 mol % to about 8 mol % of the total lipid present in the particle.
19. The nucleic acid-lipid particle of claim 12 , wherein the nucleic acid-lipid particle comprises about 54 mol % cationic lipid, about 7 mol % phospholipid, about 32 mol % cholesterol or a derivative thereof, and about 7 mol % PEG-lipid conjugate.
20. The nucleic acid-lipid particle of claim 12 , wherein the nucleic acid-lipid particle comprises about 58 mol % cationic lipid, about 35 mol % cholesterol or a derivative thereof, and about 7 mol % PEG-lipid conjugate.
21. The nucleic acid-lipid particle of claim 1 , wherein the siRNA is fully encapsulated in the nucleic acid-lipid particle.
22. A pharmaceutical composition comprising a nucleic acid-lipid particle of claim 1 and a pharmaceutically acceptable carrier.
23. A method for introducing an siRNA that silences polo-like kinase 1 (PLK-1) gene expression into a cell, the method comprising:
contacting the cell with a nucleic acid-lipid particle of claim 1 .
24. The method of claim 23 , wherein the cell is in a solid tumor.
25. The method of claim 24 , wherein the solid tumor is in a mammal.
26. The method of claim 25 , wherein the mammal is a human.
27. A method for the in vivo delivery of an siRNA that silences polo-like kinase 1 (PLK-1) gene expression to a solid tumor, the method comprising:
administering to a mammal a nucleic acid-lipid particle of claim 1 .
28. The method of claim 27 , wherein the particle is administered via a systemic route.
29. The method of claim 27 , wherein the siRNA is preferentially delivered to the solid tumor as compared to other tissues.
30. The method of claim 27 , wherein the solid tumor is a liver tumor.
31. The method of claim 27 , wherein the particle reduces the size and/or volume of the solid tumor.
32. The method of claim 27 , wherein the mammal is a human.
33. The method of claim 27 , wherein the siRNA silences PLK-1 expression by at least 50%, 60%, 70%, 80%, or 90% in a test mammal relative to the level of PLK-1 expression in a control mammal not administered the siRNA.
34. The method of claim 33 , wherein the test mammal and the control mammal are both mice.
35. A method for treating a cell proliferative disorder in a mammal in need thereof, the method comprising:
administering to the mammal a therapeutically effective amount of a nucleic acid-lipid particle of claim 1 .
36. The method of claim 35 , wherein the cell proliferative disorder is cancer.
37. The method of claim 35 , wherein the mammal is a human.
38. The nucleic acid-lipid particle of claim 1 , wherein the siRNA consists of SEQ ID NOS:3 and 16.
39. The method of claim 27 , wherein the solid tumor is a tumor located outside the liver.