Lipid encapsulating interfering rna

Abstract

The present invention provides lipid-based formulations for delivering, e.g., introducing, nucleic acid-lipid particles comprising an interference RNA molecule to a cell, and assays for optimizing the delivery efficiency of such lipid-based formulations.

1 - 37 . (canceled) 
     
     
         38 . A method of introducing a nucleic acid into a tumor cell, said method comprising contacting said tumor cell with a nucleic acid-lipid particle comprising
 (a) said 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 conjugated lipid that inhibits aggregation of particles. 
       
     
     
         39 . The method of  claim 38 , wherein said nucleic acid in said nucleic acid-lipid particle is resistant in aqueous solution to degradation with a nuclease. 
     
     
         40 . The method of  claim 38 , wherein said particle has a median diameter of less than about 150 nm. 
     
     
         41 . The method of  claim 38 , wherein nucleic acid is a small interfering RNA (siRNA). 
     
     
         42 . The method of  claim 38 , wherein said nucleic acid is transcribed from a plasmid. 
     
     
         43 . The method of  claim 38 , wherein said non-cationic lipid is a member selected from the group consisting of dioleoylphosphatidylethanolamine (DOPE), palmitoyloleoylphosphatidylcholine (POPC), egg phosphatidylcholine (EPC), di stearoylphosphatidylcholine (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. 
     
     
         44 . The method of  claim 38 , wherein the conjugated lipid that inhibits aggregation of particles is a member selected from the group consisting of a polyethyleneglycol (PEG)-lipid conjugate, a polyamide (ATTA)-lipid conjugate, and a mixture thereof. 
     
     
         45 . The method of  claim 38 , wherein the conjugated lipid that inhibits aggregation of particles is a polyethyleneglycol (PEG)-lipid. 
     
     
         46 . The method of  claim 45 , wherein the PEG-lipid is member selected from the group consisting of a PEG-diacylglycerol, a PEG dialkyloxypropyl, a PEG-phospholipid, a PEG-ceramide, and a mixture thereof. 
     
     
         47 . The method of  claim 46 , wherein the conjugated lipid that inhibits aggregation of particles is a polyethyleneglycol (PEG)-dialkyloxypropyl conjugate. 
     
     
         48 . The method of  claim 47 , wherein the PEG-dialkyloxypropyl conjugate is PEG-dimyristyloxypropyl (C 14 ). 
     
     
         49 . The method of  claim 48 , wherein said cell is in a mammal. 
     
     
         50 . The method of  claim 49 , wherein the mammal is a human. 
     
     
         51 . The method of  claim 49 , wherein presence of said nucleic acid at a tumor site distal to the site of administration is detectable for at least 48 hours after administration of said particle. 
     
     
         52 . The method of  claim 49 , wherein presence of said nucleic acid at a tumor site distal to the site of administration is detectable for at least 24 hours after administration of said particle. 
     
     
         53 . A method for in vivo delivery of nucleic acid to a liver cell, said method comprising administering to a mammalian subject a nucleic acid-lipid particle comprising:
 (a) said 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 conjugated lipid that inhibits aggregation of particles. 
       
     
     
         54 . The method of  claim 53 , wherein said mammal is a human. 
     
     
         55 . The method of  claim 54 , wherein said human has a disease or disorder associated with expression of a gene and wherein expression of said gene is reduced by said nucleic acid. 
     
     
         56 . The method of  claim 53 , wherein said disease or disorder is associated with overexpression of said gene. 
     
     
         57 . The method of  claim 53 , wherein said administration is intravenous.