Liposomes for drug delivery and methods for preparation thereof

Abstract

The present invention provides liposomes that are useful for delivery of bioactive agents such as therapeutics. Among others, the liposomes of the invention are capable of delivering their payload at sites of increased secretory phospholipase A2 (sPLA2) activity, because phospholipase A2 (PLA2) will hydrolyse lipids of the liposome. Thus, the liposomes of the invention may e.g. be used in relation to cancer therapy. Another aspect of the invention is a liposomal formulation comprising the liposome of the invention. Still another aspect is a method of producing a liposomal formulation of the invention.

1 . A liposome comprising
 between 25% and 45% (mol/mol) of an anionic lipid,   less than 1% cholesterol (mol/mol) and   a therapeutic agent selected from the group consisting of small molecule antitumour agents, antibiotics, antifungals, and anti-inflammatory agents
 wherein the liposome has been exposed to a divalent cation at a concentration between 0.1 mM and 1 mM. 
   
     
     
         2 . The liposome of  claim 1 , wherein the anionic lipid is selected from a group consisting of PI (phosphatidyl inositol), PS (phosphatidyl serine), DPG (bisphosphatidyl glycerol), PA (phosphatidic acid), PEOH (phosphatidyl alcohol), and PG (phosphatidyl glycerol). 
     
     
         3 . The liposome of  claim 2 , wherein the anionic lipid is phosphatidyl glycerol. 
     
     
         4 . The liposome of  claim 1 , further comprising a hydrophilic polymer selected from the group of PEG [poly(ethylene glycol)], PAcM [poly(N-acryloylmorpholine)], PVP [poly(vinylpyrrolidone)], PLA [poly(lactide)], PG [poly(glycolide)], POZO [poly(2-methyl-2-oxazoline)], PVA [poly(vinyl alcohol)], HPMC (hydroxypropylmethylcellulose), PEO [poly(ethylene oxide)], chitosan [poly(D-glucosamine)], PAA [poly(aminoacid)], polyHEMA [Poly(2-hydroxyethylmethacrylate)] and co-polymers thereof. 
     
     
         5 . The liposome of  claim 4 , wherein the polymer is PEG with a molecular weight between 100 Da and 10 kDa. 
     
     
         6 . The liposome of  claim 4 , wherein polymer is conjugated to the head group of phosphatidyl ethanolamine. 
     
     
         7 . The liposome of  claim 6 , wherein the amount of polymer-conjugated lipid is between 2.5% and 7.5% (mol/mol). 
     
     
         8 . The liposome of  claim 1 , further comprising an uncharged phospholipid selected from the group consisting of PC (phosphatidyl choline) and PE (phosphatidylethanolamine). 
     
     
         9 . The liposome of  claim 1 , comprising no cholesterol. 
     
     
         10 . The liposome of  claim 1 , wherein the alkyl chains of the lipids are C18 saturated chains. 
     
     
         11 . The liposome of  claim 1 , wherein the therapeutic agent is a small molecule antitumour agent selected from the group consisting of anthracyclin derivatives, cisplatin, oxaliplatin, carboplatin, doxorubicin, paclitaxel, 5-fluoruracil, exisulind, cis-retinoic acid, suldinac sulphide, methotrexate, bleomycin and vincristine. 
     
     
         12 . The liposome of  claim 11 , wherein the therapeutic agent is oxaliplatin or cisplatin. 
     
     
         13 . The liposome of  claim 1 , wherein the liposome is a Large Unilamellar Vesicle (LUV). 
     
     
         14 . The liposome of  claim 13 , wherein the liposome has a diameter between 80 and 120 nm. 
     
     
         15 . The liposome of  claim 1 , wherein at least one of the lipids in the liposome is a substrate for sPLA 2 . 
     
     
         16 . A liposome of any of  claim 1 , further comprising between 0.1 mM and 1 mM of a divalent cation. 
     
     
         17 . The liposome of  claim 16 , wherein the divalent cation is Ca 2+ . 
     
     
         18 . The liposome formulation comprising liposomes according to  claim 17 . 
     
     
         19 . The liposome formulation of  claim 18 , wherein the Poly Dispersity Index is 0.20 or less. 
     
     
         20 . A method for the preparation of a liposomal formulation according to  claim 18  comprising the steps of:
 a. preparing a lipid mixture by dissolving selected lipids in an organic solvent 
 b. hydrating the product of step a) with an aqueous hydration solvent so as to form liposomes; and 
 c. removing the organic solvent of step a) either before addition of the aqueous hydration solvent or after the addition of the aqueous hydration solvent. 
 
     
     
         21 . The method of  claim 20 , wherein organic solvent is removed before addition of hydration solvent. 
     
     
         22 . The method of  claim 20  further comprising a step of sonicating the liposomal formulation to produce liposomes of a certain size. 
     
     
         23 . The method of  claim 20 , wherein the hydration solvent comprises a divalent cation at a concentration between 0.1 mM and 1 mM. 
     
     
         24 . The method of  claim 20  further comprising a step of changing the exterior water phase to another exterior water phase comprising a divalent cation at a concentration 0.1 mM and 1 mM. 
     
     
         25 . The method of  claim 20 , wherein the divalent cation is Ca 2+ .