Nucleic acid-lipopolymer compositions

Abstract

Compositions, methods, and applications that increase the efficiency of nucleic acid transfection are provided. In one aspect, a pharmaceutical composition may include at least about 0.5 mg/ml concentration of a nucleic acid condensed with a cationic lipopolymer suspended in an isotonic solution, where the cationic lipopolymer includes a cationic polymer backbone having cholesterol and polyethylene glycol covalently attached thereto, and wherein the molar ratio of cholesterol to cationic polymer backbone is within a range of from about 0.1 to about 10, and the molar ratio of polyethylene glycol to cationic polymer backbone is within a range of from about 0.1 to about 10. The composition further may include a filler excipient.

1 - 52 . (canceled) 
     
     
         53 . A dry pharmaceutical composition comprising: a filler excipient, a mixture of nucleic acids, and a cationic lipopolymer, wherein said cationic lipopolymer consists of polyethyleneimine (PEI) covalently linked independently to cholesterol and/or polyethylene glycol (PEG) groups, wherein the average PEI:PEG:cholesterol molar ratio in the cationic lipopolymer is within the range of 1-5 PEG:1 PEI:0.4-1.5 cholesterol, the ratio of amine nitrogen in the cationic lipopolymer to phosphate in the nucleic acids is from about 0.1:1 to about 100:1 and the mixture of nucleic acids and the cationic lipopolymer forms a complex. 
     
     
         54 . The dry pharmaceutical composition according to  claim 53 , wherein the mixture of nucleic acids comprises a plasmid DNA. 
     
     
         55 . The dry pharmaceutical composition according to  claim 53 , wherein the mixture of nucleic acids comprises a plasmid DNA encoding for a protein or peptide. 
     
     
         56 . The dry pharmaceutical composition according to  claim 55 , wherein the mixture of nucleic acids further comprises a plasmid encoding an shRNA. 
     
     
         57 . The dry pharmaceutical composition according to  claim 55 , wherein the mixture further comprises a synthetic RNA. 
     
     
         58 . The dry pharmaceutical composition according to  claim 55  wherein the protein is a therapeutic protein selected from the group consisting of interleukin-2, interleukin-4, interleukin-7, interleukin-12, interleukin-15, interferon-α, interferon-β, interferon-γ, colony stimulating factor, granulocyte-macrophage stimulating factor, antiangiogenic agents, tumor suppressor genes, thymidine kinase, eNOS, iNOS, p53, p16, TNF-α, Fas-ligand, mutated oncogenes, tumor antigens, viral antigens or bacterial antigens. 
     
     
         59 . A reconstituted composition comprising the dry pharmaceutical composition of  claim 53  and a diluent. 
     
     
         60 . The reconstituted composition of  claim 59 , wherein the diluent is selected from an isotonic solution or water. 
     
     
         61 . The reconstituted composition of  claim 59 , wherein the concentration of nucleic acids is at least about 0.5 mg/ml. 
     
     
         62 . A method of transfecting a mammalian cell comprising contacting the mammalian cell with the reconstituted composition of  claim 59 , wherein the composition enters the cell and elicits biological activity of the nucleic acid mixture. 
     
     
         63 . A method of transfecting a mammalian cell, comprising contacting the mammalian cell with the reconstituted composition of  claim 59 , wherein the reconstituted composition enters the cell and results in the expression of a peptide and inhibition of the transcript targeted by the shRNA. 
     
     
         64 . A method of treating cancer in a subject in need thereof comprising administering to the subject the reconstituted composition of  claim 59 . 
     
     
         65 . A method of transfecting a mammalian cell, comprising contacting the mammalian cell with a reconstituted composition comprising a filler excipient, a nucleic acid, a cationic lipopolymer, and a diluent, wherein said cationic lipopolymer consists of polyethyleneimine (PEI) covalently linked independently to cholesterol and/or polyethylene glycol (PEG) groups, wherein the average PEI:PEG:cholesterol molar ratio in the cationic lipopolymer is within the range of 1-5 PEG:1:PEI0.4-1.5 cholesterol, wherein the nucleic acid and cationic lipopolymer form a complex, and wherein the complex of the reconstituted composition enters the mammalian cell and elicits an increase in Interferon-γ protein. 
     
     
         66 . The method of  claim 65 , wherein the mammalian cell is a cancer cell. 
     
     
         67 . The method of  claim 66 , wherein the nucleic acid is a plasmid DNA. 
     
     
         68 . The method of  claim 67 , wherein the plasmid DNA expresses an IL-12 gene. 
     
     
         69 . A method of treating cancer in a subject comprising administering a reconstituted composition comprising a filler excipient, a nucleic acid, a cationic lipopolymer, and a diluent, wherein said cationic lipopolymer consists of polyethyleneimine (PEI) covalently linked independently to cholesterol and/or polyethylene glycol (PEG) groups, wherein the average PEI:PEG:cholesterol molar ratio in the cationic lipopolymer is within the range of 1-5 PEG:1:PEI0.4-1.5 cholesterol, wherein the nucleic acid and cationic lipopolymer form a complex, and wherein the complex of the reconstituted composition enters the mammalian cell and elicits the biological activity of the nucleic acid, and wherein the administration is intraperitoneal. 
     
     
         70 . The method of  claim 69 , wherein the administration is at least four times weekly. 
     
     
         71 . The method of  claim 69 , wherein the nucleic acid is a plasmid DNA. 
     
     
         72 . The method of  claim 71 , wherein the plasmid DNA expresses an IL-12 gene.