Lipid-nucleic acid particles prepared via a hydrophobic lipid-nucleic acid complex intermediate and use for gene transfer

A method for the preparation of lipid-nucleic acid particles, comprising:

• (a) contacting an amount of cationic lipids with nucleic acids in a solution;
• said solution comprising of from about 15-35% water and about 65-85% organic solvent and said amount of cationic lipids being sufficient to produce a +/- charge ratio of from about 0.85 to about 2.0, to provide a hydrophobic, charge-neutralized lipid-nucleic acid complex;
• (b) contacting said hydrophobic lipid-nucleic acid complex in solution with non-cationic lipids, to provide a lipid-nucleic acid mixture;
• and
• (c) removing said organic solvents from said mixture to provide said lipid-nucleic acid particles in which said nucleic acids are protected from degradation.

The method for the preparation of serum-stable plasmid-lipid particles, comprising:

• (a) combining a plasmid with cationic lipids in a detergent solution to provide a coated plasmid-lipid complex;
• (b) contacting non-cationic lipids with said coated plasmid-lipicl complex to provide a solution comprising detergent, a plasmid-lipid complex and non-cationic lipids;
• and
• (c) removing said detergent from said solution of step (b) to provide a solution of serum-stable plasmid-lipid particles, wherein said plasmid is encapsulated in a lipid bilayer and said particles are serum-stable and have a size of from about 50 to about 150 nm.

A method for the preparation of serum-stable plasmid-lipid particles, comprising;

• (a) preparing a mixture comprising cationic lipids and non-cationic lipids in an organic solvent;
• (b) contacting an aqueous solution of plasmid with said mixture prepared in step (a) to provide a clear single phase;
• and
• (c) removing said organic solvent to provide a suspension of plasmid-lipid particles, wherein said plasmid is encapsulated in a lipid bilayer, and said particles are stable in serum and have a size of from about 50 to about 150 nm.