Lipid particles having asymmetric lipid coating and method of preparing same

Abstract

A method of preparing lipid particles having an asymmetric lipid coating is described. The lipid composition of the outer lipid coating of the particles varies from the inner to outer surfaces. The asymmetric lipid particles are formed by preparing a lipid composition containing a charged lipid and a therapeutic agent, where the particles each have an outer lipid coating with an external lipid leaflet and an internal lipid structure. The particles are then incubated under conditions effective to remove the charged lipid from the external lipid leaflet, thus rendering the lipid coating asymmetric. The particles have the ability to their regain surface charge via translocation of the lipids.

1 . A method of preparing lipid particles having an external lipid coating, comprising: 
 preparing lipid particles comprised of (i) a charged lipid and (ii) a therapeutic agent, said particles each having an external lipid leaflet; and    incubating said particles under conditions effective to remove said charged lipid from the external lipid leaflet.    
   
   
       2 . The method of  claim 1 , wherein said preparing is comprised of preparing lipid particles composed of a lipid composition containing at least one cationic lipid.  
   
   
       3 . The method of  claim 1 , wherein said preparing comprises (i) forming lipid vesicles composed of said lipid composition and (ii) complexing said lipid vesicles with said therapeutic agent.  
   
   
       4 . The method of  claim 1 , wherein said incubating comprises incubating said lipid particles in a medium containing uncharged lipid vesicles.  
   
   
       5 . The method of  claim 4 , wherein said incubating further includes adding to the medium a lipid-polymer-ligand conjugate.  
   
   
       6 . The method of  claim 1 , wherein said lipid particles are liposomes.  
   
   
       7 . The method of  claim 6 , wherein said incubating further includes adding to the medium a lipid derivatized with a hydrophilic polymer.  
   
   
       8 . The method of  claim 7  wherein said adding is comprised of adding a phospholipid derivatized with polyethyleneglycol.  
   
   
       9 . The method of  claim 1 , wherein said incubating is at a temperature of less than about 15° C.  
   
   
       10 . The method of  claim 1 , wherein said incubating is for a time of greater than about 5 hours.  
   
   
       11 . The method of  claim 1 , wherein said preparing is comprised of preparing lipid particles having an entrapped therapeutic agent selected from the group consisting of a charged drug, a protein, a peptide, and a nucleic acid.  
   
   
       12 . The method of  claim 11 , wherein said therapeutic agent is a protein or peptide.  
   
   
       13 . A composition, comprising 
 lipid particles having a lipid coating comprised of an outer lipid leaflet and an inner lipid structure, said lipid coating formed of a lipid composition (i) comprising a charged lipid and (ii) having a gel-crystalline phase transition temperature, said lipid particles having no appreciable charge at a temperature lower than said phase transition temperature but having a measurable charge after incubation at a temperature above said phase transition temperature.    
   
   
       14 . The composition of  claim 13 , wherein said lipid composition comprises a cationic lipid.  
   
   
       15 . The composition of  claim 13 , wherein said lipid particles further include a therapeutic agent having a charge.  
   
   
       16 . The composition of  claim 15 , wherein said therapeutic agent is a nucleic acid.  
   
   
       17 . The composition of  claim 15 , wherein said lipid composition has a phase transition of between about 34-38° C.  
   
   
       18 . The composition of  claim 15 , wherein said lipid particles are liposomes.