Cationic lipids and methods of use

Abstract

The present invention provides compositions comprising cationic lipids, liposomes and nucleic acid-lipid particles comprising the cationic lipids, and methods of using such compositions, liposomes, and nucleic acid-lipid particles.

1 .- 7 . (canceled) 
     
     
         8 . A liposome, said liposome comprising a cationic lipid of Formula I having the following structure: 
       
         
           
           
               
               
           
         
         wherein: 
         R 1  and R 2  are independently selected from the group consisting of: H and C 1 -C 3  alkyls; and 
         R 3  and R 4  are independently selected from the group consisting of alkyl groups having from about 10 to about 20 carbon atoms, wherein at least one of R 3  and R 4  comprises at least two sites of unsaturation. 
       
     
     
         9 . The liposome in accordance with  claim 8 , further comprising a non-cationic lipid. 
     
     
         10 . The liposome in accordance with  claim 9 , wherein said non-cationic lipid is a member selected from the group consisting of dioleoylphosphatidylethanolamine (DOPE), palmitoyloleoylphosphatidylcholine (POPC), egg phosphatidylcholine (EPC), distearoylphosphatidylcholine (DSPC), palmitoyloleyolphosphatidylglycerol (POPG), dipalmitoyl phosphatidyl ethanolamine (DPPE), dimyristoylphosphoethanolamine (DMPE), distearoyl-phosphatidyl-ethanolamine (DSPE), 16-O-monomethyl PE, 16-O-dimethyl PE, 18-1-trans PE, palmitoyloleoyl-phosphatidylethanolamine (POPE), 1-stearoyl-2-oleoyl-phosphatidyethanolamine (SOPE), cholesterol, and a mixture thereof. 
     
     
         11 . The liposome in accordance with  claim 8 , further comprising a PEG-lipid. 
     
     
         12 . The liposome in accordance with  claim 10 , wherein said PEG-lipid is selected from the group consisting of: a polyethyleneglycol-diacylglycerol (PEG-DAG) conjugate, a polyethyleneglycol-dialkyloxypropyl (PEG-DAA) conjugate, a PEG-ceramide, a PEG-PE, and mixtures thereof. 
     
     
         13 . The liposome in accordance with  claim 8 , further comprising a bioactive agent. 
     
     
         14 . The liposome in accordance with  claim 13 , wherein said bioactive agent is a member selected from the group consisting of: an antineoplastic agent, an antibiotic, an immunomodulator, an anti-inflammatory agent, an agent acting on the central nervous system, a polypeptide, and a nucleic acid. 
     
     
         15 . A method of delivering a bioactive agent to a cell, said method comprising contacting said cell with a liposome of  claim 8 , wherein said bioactive agent is encapsulated in said liposome. 
     
     
         16 . The method of  claim 15 , wherein said is in a mammal. 
     
     
         17 . A nucleic acid-lipid particle comprising:
 (a) a nucleic acid;   (b) a cationic lipid of Formula I and having the following structure:   
       
         
           
           
               
               
           
         
         wherein: 
         R 1  and R 2  are independently selected from the group consisting of: H and C 1 -C 3  alkyls; and 
         R 3  and R 4  are independently selected from the group consisting of alkyl groups having from about 10 to about 20 carbon atoms, wherein at least one of R 3  and R 4  comprises at least two sites of unsaturation; 
         (c) a non-cationic lipid; and 
         (d) a PEG-lipid conjugate. 
       
     
     
         18 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid is selected from the group consisting of: 1,2-DiLinoleyloxy-N,N-dimethylaminopropane (DLinDMA) and 1,2-Dilinolenyloxy-N,N-dimethylaminopropane (DLenDMA). 
     
     
         19 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid is DLinDMA. 
     
     
         20 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said non-cationic lipid is a member selected from the group consisting of dioleoylphosphatidylethanolamine (DOPE), palmitoyloleoylphosphatidylcholine (POPC), egg phosphatidylcholine (EPC), distearoylphosphatidylcholine (DSPC), palmitoyloleyolphosphatidylglycerol (POPG), dipalmitoyl phosphatidyl ethanolamine (DPPE), dimyristoylphosphoethanolamine (DMPE), distearoyl-phosphatidyl-ethanolamine (DSPE), 16-O-monomethyl PE, 16-O-dimethyl PE, 18-1-trans PE, palmitoyloleoyl-phosphatidylethanolamine (POPE), 1-stearoyl-2-oleoyl-phosphatidyethanolamine (SOPE), cholesterol, and a mixture thereof. 
     
     
         21 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said non-cationic lipid is DSPC. 
     
     
         22 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said PEG-lipid is a member selected from the group consisting of: a polyethyleneglycol-diacylglycerol (PEG-DAG) conjugate, a polyethyleneglycol-dialkyloxypropyl (PEG-DAA) conjugate, a PEG-ceramide, a PEG-PE, and mixtures thereof. 
     
     
         23 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said PEG-lipid is a PEG-DAA conjugate. 
     
     
         24 . The nucleic acid-lipid particle in accordance with  claim 23 , wherein the PEG-DAA conjugate is PEG-dimyristyloxypropyl (C14). 
     
     
         25 . The nucleic acid-lipid particle in accordance with  claim 17 , further comprising a sterol. 
     
     
         26 . The nucleic acid-lipid particle in accordance with  claim 25 , wherein the sterol is cholesterol. 
     
     
         27 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said nucleic acid is a member selected from the group consisting of: DNA, RNA, siRNA, a plasmid, an antisense oligonucleotide, a ribozyme. 
     
     
         28 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said nucleic acid encodes a therapeutic product of interest. 
     
     
         29 . The nucleic acid-lipid particle in accordance with  claim 28 , wherein said therapeutic product of interest is a member selected from the group consisting of: a polypeptide and a small interfering RNA (siRNA). 
     
     
         30 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein the nucleic acid in said nucleic acid-lipid particle is not substantially degraded after exposure of said particle to a nuclease at 37° C. for 20 minutes. 
     
     
         31 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein the nucleic acid is fully encapsulated in said nucleic acid-lipid particle. 
     
     
         32 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid comprises from about 2% to about 60% of the total lipid present in said particle. 
     
     
         33 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid comprises from about 5% to about 45% of the total lipid present in said particle. 
     
     
         34 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid comprises from about 15% to about 40% of the total lipid present in said particle. 
     
     
         35 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid comprises about 30% of the total lipid present in said particle. 
     
     
         36 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said cationic lipid comprises from about 40% to about 50% of the total lipid present in said particle. 
     
     
         37 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said non-cationic lipid comprises from about 5% to about 90% of the total lipid present in said particle. 
     
     
         38 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said non-cationic lipid comprises from about 20% to about 85% of the total lipid present in said particle. 
     
     
         39 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said PEG-lipid comprises from 1% to about 20% of the total lipid present in said particle. 
     
     
         40 . The nucleic acid-lipid particle in accordance with  claim 17 , wherein said PEG-lipid comprises from 4% to about 15% of the total lipid present in said particle. 
     
     
         41 . The nucleic acid-lipid particle in accordance with  claim 25 , wherein the sterol comprises from about 10% to about 60% of the total lipid present in said particle. 
     
     
         42 . The nucleic acid-lipid particle in accordance with  claim 25 , wherein the sterol comprises from about 20% to about 45% of the total lipid present in said particle. 
     
     
         43 . A pharmaceutical composition comprising a nucleic acid-lipid particle in accordance with  claim 17  and a pharmaceutically acceptable carrier. 
     
     
         44 . A method of introducing a nucleic acid into a cell, said method comprising contacting said cell with a nucleic acid-lipid particle comprising:
 (a) a cationic lipid of Formula I and having the following structure:   
       
         
           
           
               
               
           
         
         wherein: 
         R 1  and R 2  are independently selected from the group consisting of: H and C 1 -C 3  alkyls; and 
         R 3  and R 4  are independently selected from the group consisting of alkyl groups having from about 10 to about 20 carbon atoms, wherein at least one of R 3  and R 4  comprises at least two sites of unsaturation. 
         (b) a non-cationic lipid; 
         (c) a PEG-lipid conjugate; and 
         (d) a nucleic acid.