Compositions and methods for delivering nucleic acid to a cell

Abstract

Provided are novel compositions useful for delivering nucleic acids to cells. Also provided are methods for making and using such compositions.

1 . A method for preparing a liposome comprising a nucleic acid
 and a lipid component, the method comprising:
 combining a lipid component and a nucleic acid component in a mixture comprising:
 water; 
 a water-miscible organic solvent; 
 a polyamine that is spermine, spermidine, or putrescine; and 
 a total lipid; 
 
   wherein the lipid component comprises a combination of a neutral lipid and a cationic lipid that is less than 0.5% of the total lipid under conditions such that a liposome comprising the nucleic acid and the liposome component is formed.   
     
     
         2 - 3 . (canceled) 
     
     
         4 . The method of  claim 1 , wherein the polyamine is spermine. 
     
     
         5 . The method of  claim 1 , wherein the nucleic acid is DNA. 
     
     
         6 . The method of  claim 1 , wherein the nucleic acid is RNA. 
     
     
         7 . The method of  claim 6 , wherein the RNA is siRNA. 
     
     
         8 . The method of  claim 6 , wherein the RNA is shRNA. 
     
     
         9 . The method of  claim 1 , wherein the lipid component comprises a non-cationic lipid. 
     
     
         10 . The method of  claim 1 , wherein the neutral lipid comprises a non-cationic lipid. 
     
     
         11 . The method of  claim 1 , wherein the neutral lipid comprises DOPC, DOPE, cholesterol, or PEG-DSG. 
     
     
         12 . The method of  claim 9 , wherein the liposome further comprises a cationic lipid. 
     
     
         13 . The method of  claim 12 , wherein the cationic lipid comprises DOTAP or DOSPA. 
     
     
         14 . The method of  claim 1 , wherein the water-miscible organic solvent comprises methanol, ethanol, 1-propanol, or 2-propanol. 
     
     
         15 . The method of  claim 1 , wherein the lipid component contains no cationic lipid. 
     
     
         16 . The method of  claim 1 , wherein the ratio of water to water-miscible organic solvent is between about 2:1 and 1:2. 
     
     
         17 . The method of  claim 1 , wherein the ratio of water to water-miscible organic solvent is about 1:1 
     
     
         18 . (canceled) 
     
     
         19 . The method of  claim 17 , wherein the ratio of polyamine nitrogen to nucleic acid phosphate groups (N/P) is between about 0.8 and about 1.5. 
     
     
         20 . (canceled) 
     
     
         21 . The method of  claim 19 , wherein the step of combining is performed at a temperature between about 40° C. and about 50° C. 
     
     
         22 . The method of  claim 21 , wherein the step of combining is performed at a pH not less than about 6.5. 
     
     
         23 - 24 . (canceled) 
     
     
         25 . The method of  claim 14 , wherein the lipid component comprises a neutral phospholipid and no cationic lipid, and wherein the nucleic acid and the lipid component are present at a ratio of 5-20 nmol lipid per microgram of the nucleic acid, and wherein the liposome is from 30 to 500 nanometers in diameter. 
     
     
         26 - 51 . (canceled)