Vesicle-forming lipids containing PEG-PE, PG, PHEPC, and cholesterol and α-TC in a mole ratio of 0.15: 0.3: 1.85: 1: 0.2 were dissolved in chloroform to a final lipid concentration of 25 μmol phospholipid/ml. The lipid mixture was dried into a thin film under reduced pressure. The film was hydrated with a solution of 0.125 M ammonium sulfate to form MLVs. The MLV suspension was frozen in a dry ice acetone bath and thawed three times and sized to 80-100 nm by extrusion as detailed above. An ammonium ion gradient was created substantially as described in Example 10. The liposomes were loaded with epirubicin, and free (unbound drug) removed also as described in Example 10 for doxorubicin. The final concentration of entrapped drug was about 50-100 μg drug/μmol lipid. Epirubicin HCl and doxorubicin HCL, the commercial products, were obtained from the hospital pharmacy.
