Optimization and upscaling of doxorubicin‐containing liposomes for clinical use

This work describes the optimization of a doxorubicin (DXR)‐containing liposome formulation and its upscaling for human therapy. Multilamellar vesicles (MLV) composed of egg phosphatidylcholine, egg‐derived phosphatidylglycerol, cholesterol, and the drug were prepared in 0.9% sterile, pyrogen‐free NaCl by five different hydration methods. The optimal hydration was shown to be the formation of a thin lipid film with high surface area. Alternative hydration methods based on freeze‐drying techniques of the lipids in tertiary butanol or based on “alcohol” premixing procedures of the dry DXR‐lipid mixture showed smaller DXR loading capacities and lower DXR incorporation per phospholipid. Maximal DXR entrapment was obtained at a molar concentration of phosphatidylglycerol of 30 mol % of total phospholipid. This and previous studies led to a final lipid composition of phosphatidylcholine:phosphatidylglycerol:cholesterol in a molar ratio of 7:3:4. Oligolamellar DXR‐liposomes with an average diameter in the range 0.3–0.5 μM were prepared from DXR‐MLV by extrusion through polycarbonate membranes using moderate pressures (up to 100 psi). At this size range, maximal entrapment of DXR per phospholipid was obtained. The extrusion process also ensures the sterilization of the final product. Free DXR was removed from liposome‐associated DXR (L‐DXR) by the use of a cation‐exchange resin. The L‐DXR formulation was shown to have reasonable stability on storage at 4 °C.