Amphoteric liposome formulations

Abstract

An amphoteric liposome composed of a mixture of lipids, said mixture comprising a cationic amphiphile, an anionic amphiphile and optionally one or more neutral amphiphiles, at least one of said cationic and anionic amphiphiles being chargeable and the respective amounts of said cationic and anionic amphiphiles being selected such there is a stoichiometric excess of positively charged cationic amphiphile at a first lower pH, a stoichiometric excess of negatively charged anionic amphiphile at a second higher pH and said mixture has an isoelectric point intermediate said first and second pHs; characterised in that said positively charged cationic and negatively charged anionic amphiphiles are adapted to form a lipid salt with one another at said isoelectric point. Also disclosed are methods of predicting the fusogenicity of an amphoteric liposome at a given pH, formulating an amphoteric liposome and loading an amphoteric liposome with a cargo moiety.

1 . An amphoteric liposome formulation comprising a cationic amphiphile, an anionic amphiphile, and one or more neutral amphiphiles. 
     
     
         2 . The amphoteric liposome formulation of  claim 1 , wherein the cationic amphiphile is selected from the group consisting of 4-(2-aminoethyl)-morpholino-cholesterol-2,3-dimethylhemisuccinate (DmC4Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-2,3-dimethylhemimalonate (DmC3Mo2), 4-(2-aminobutyl)-morpholino-cholesterol-hemisuccinate (C4Mo4), 4-(2-aminopropyl)-morpholino-cholesterol-hemimalonate (C3Mo3), 4-(2-aminoethyl)-morpholino-cholesterol-hemimalonate (C3Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-hemiglutarate (C5Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-hemiadipate (C6Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-hemiadipate (C8Mo2), 4-(2-Aminoethyl)-Morpholino-Cholesterolhemisuccinate (MoChol), Cholesterol-(3-imidazol-1-yl propyl)carbamate (CHIM), and Histaminyl-Cholesterolhemisuccinate (HisChol). 
     
     
         3 . The amphoteric liposome formulation of  claim 1 , wherein the anionic amphiphile is selected from the group consisting of cholesterol hemisuccinate CHEMS, Dimyristoylglycerolhemisuccinate DMGS, Dimyristoylglycerolhemimalonate DMGM, Dimyristoylglycerolhemiglutarate DMGG, Dimyristoylglycerolhemiadipate DMGA, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxobutanoic acid DMAS, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxopropanoic acid DMAM, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxopentanoic acid DMAG, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxohexanoic acid DMAA, Dioleoylglycerolhemisuccinate DOGS, Dioleoylglycerolhemimalonate DOGM, Dioleoylglycerolhemiglutarate DOGG, Dioleoylglycerolhemiadipate DOGA, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxobutanoic acid DOAS, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxopropanoic acid DOAM, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxopentanoic acid DOAG, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxohexanoic acid DOAA, 5,6-Dimyristoyl-hexanoic acid DMS, 4,5-Dimyristoyl-pentanoic acid DMM, 6,7-Dimyristoyl-heptanoic acid DMG, 7,8-Dioleoyl-octanoic acid DMA, 5,6-Dioleoyl-hexanoic acid DOS, 4,5-Dioleoyl-pentanoic acid DOM, 6,7-Dioleoyl-heptanoic acid DOG, 7,8-Dioleoyl-octanoic acid DOA, Cholesterolhemimalonate Chol-C3, Cholesterolhemiglutarate Chol-C5, and Cholesterolhemiadipate Chol-C6. 
     
     
         4 . The amphoteric liposome formulation of  claim 1 , wherein the neutral amphiphiles are selected from the group consisting of phosphatidylcholines, sphingomyelins, ceramides, phosphatidylethanolamines, cholesterol, and mixtures thereof. 
     
     
         5 . The amphoteric liposome formulation of  claim 1 , wherein the neutral amphiphiles are selected from phosphatidylcholines and phosphatidylethanolamines. 
     
     
         6 . The amphoteric liposome formulation of  claim 1 , wherein the neutral amphiphiles are selected from POPC and DOPE. 
     
     
         7 . The amphoteric liposome formulation of  claim 1 , wherein the formulation has an isoelectric point of from pH 4 to pH 8. 
     
     
         8 . The amphoteric liposome formulation of  claim 1 , wherein the neutral amphiphiles are less than 50 mol % of the composition and the composition has an isoelectric point of from pH 4 to pH 8. 
     
     
         9 . The amphoteric liposome formulation of  claim 1 , wherein the formulation comprises an amphoteric liposome. 
     
     
         10 . The amphoteric liposome formulation of  claim 1 , wherein the formulation has an isoelectric point of from pH 5 to pH 7. 
     
     
         11 . The amphoteric liposome formulation of  claim 1 , wherein the formulation has a stable lamellar phase at pH 7 to pH 8. 
     
     
         12 . The amphoteric liposome formulation of  claim 1 , wherein the molar ratio of the cationic amphiphile to the anionic amphiphile is greater than or equal to 1. 
     
     
         13 . The amphoteric liposome formulation of  claim 1 , wherein the neutral amphiphiles are less than 40 mol % of the formulation. 
     
     
         14 . The amphoteric liposome formulation of  claim 1 , wherein the formulation comprises liposomes that encapsulate at least one active agent. 
     
     
         15 . The amphoteric liposome formulation of  claim 14 , wherein the at least one active agent is a nucleic acid. 
     
     
         16 . The amphoteric liposome formulation of  claim 14 , wherein the at least one active agent is an oligonucleotide. 
     
     
         17 . The amphoteric liposome formulation of  claim 14 , wherein the at least one active agent is a biologically active agent. 
     
     
         18 . An amphoteric liposome-nucleic acid particle, comprising a cationic amphiphile, an anionic amphiphile, one or more neutral amphiphiles, and a nucleic acid. 
     
     
         19 . The amphoteric liposome-nucleic acid particle of  claim 18 , wherein the cationic amphiphile is selected from the group consisting of 4-(2-aminoethyl)-morpholino-cholesterol-2,3-dimethylhemisuccinate (DmC4Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-2,3-dimethylhemimalonate (DmC3Mo2), 4-(2-aminobutyl)-morpholino-cholesterol-hemisuccinate (C4Mo4), 4-(2-aminopropyl)-morpholino-cholesterol-hemimalonate (C3Mo3), 4-(2-aminoethyl)-morpholino-cholesterol-hemimalonate (C3Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-hemiglutarate (C5Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-hemiadipate (C6Mo2), 4-(2-aminoethyl)-morpholino-cholesterol-hemiadipate (C8Mo2), 4-(2-Aminoethyl)-Morpholino-Cholesterolhemisuccinate (MoChol), Cholesterol-(3-imidazol-1-yl propyl)carbamate (CHIM), and Histaminyl-Cholesterolhemisuccinate (HisChol), the anionic amphiphile is selected from the group consisting of cholesterol hemisuccinate CHEMS, Dimyristoylglycerolhemisuccinate DMGS, Dimyristoylglycerolhemimalonate DMGM, Dimyristoylglycerolhemiglutarate DMGG, Dimyristoylglycerolhemiadipate DMGA, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxobutanoic acid DMAS, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxopropanoic acid DMAM, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxopentanoic acid DMAG, 4-((1,2-Dimyristoyl-ethyl)amino)-4-oxohexanoic acid DMAA, Dioleoylglycerolhemisuccinate DOGS, Dioleoylglycerolhemimalonate DOGM, Dioleoylglycerolhemiglutarate DOGG, Dioleoylglycerolhemiadipate DOGA, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxobutanoic acid DOAS, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxopropanoic acid DOAM, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxopentanoic acid DOAG, 4-((1,2-Dioleoyl-ethyl)amino)-4-oxohexanoic acid DOAA, 5,6-Dimyristoyl-hexanoic acid DMS, 4,5-Dimyristoyl-pentanoic acid DMM, 6,7-Dimyristoyl-heptanoic acid DMG, 7,8-Dioleoyl-octanoic acid DMA, 5,6-Dioleoyl-hexanoic acid DOS, 4,5-Dioleoyl-pentanoic acid DOM, 6,7-Dioleoyl-heptanoic acid DOG, 7,8-Dioleoyl-octanoic acid DOA, Cholesterolhemimalonate Chol-C3, Cholesterolhemiglutarate Chol-C5, and Cholesterolhemiadipate Chol-C6, and the neutral amphiphiles are selected from the group consisting of phosphatidylcholines, sphingomyelins, ceramides, phosphatidylethanolamines, cholesterol, and mixtures thereof. 
     
     
         20 . The amphoteric liposome-nucleic acid particle of  claim 19 , wherein the nucleic acid is a plasmid, a linear DNA construct, an oligonucleotide, a mRNA, a shRNA, a miRNA, a ribozyme, an aptamer, or an RNA that can specifically regulate a protein expression level.