Multivescular liposomes with controlled release of encapsulated biologically active substances

A process for producing a multivesicular liposome having multiple non-concentric chambers with internal membranes distributed as a network throughout, said process comprising the steps of:

• (a) forming a water-in-oil emulsion from two immiscible components, the two immiscible components being 1) a lipid component comprising at least one organic solvent, at least one amphipathic lipid, and at least one neutral lipid lacking a hydrophilic head group, and 2) a first aqueous component;
• said water-in-oil emulsion further comprising a non-hydrohalic acid selected from the group consisting of perchloric, nitric, formic, sulfuric, phosphoric, acetic, glucuronic, citric, trichloroacetic, trifluoroacetic acid, their salts and combinations thereof, in a concentration range from about 0.1 mM to about 0.5M, and at least one biologically active substance selected from the group consisting of an antitumor agent, an anesthetic, an analgesic, an antimicrobial agent, a hormone, an antiasthmatic agent, a cardiac glycoside, an antihypertensive, a vaccine, an antiarrhythmic, an immunomodulator, a steroid, a monoclonal antibody, a neurotransmitter a radionuclide, a radio contrast agent, a nucleic acid, a protein, a herbicide, a pesticide, and suitable combinations thereof, said non-hydrohalic acid and biologically active substance being independently incorporated into either the lipid component or the first aqueous component, or into both,
• (b) mixing the water-in-oil emulsion containing the non-hydrohalic acid with a second aqueous component to form solvent spherules;
• and thereafter
• (c) removing the organic solvent from the solvent spherules to form multivesicular liposomes;
• wherein the non-hydrohalic acid concentration in the water-in-oil emulsion is selected to provide controlled release of the biologically active substance from the liposomes.