LIPOSOMAL L-ASPARAGINASE - IN-VITRO EVALUATION

The purpose of this work was the development of liposomal formulations Of L-asparaginase (L-ASNase) with the following characteristics: pres ervation of active enzyme, high entrapment efficiency, prolonged serum half-life and reduced toxicity compared with the free enzyme. Several liposome formulations were developed using simplified dehydration-reh ydration vesicles (sDRV) or extruded vesicles (VET). The effect of lip id composition, vesicle size, ionic strength and osmolarity on enzyme encapsulation was investigated. Using a simplified dehydration-rehydra tion method (sDRV) we were able to achieve encapsulation efficiencies of up to 100% with full preservation (99%) of the specific activity of the encapsulated enzyme. The protein to lipid ratios of the liposomal formulations ranged from 5 to 27 mug/mumol, depending on the lipid co mposition. Extruded vesicles ranging from 85 to 250 nm in diameter wer e also tested. The encapsulation efficiency of extruded vesicles was l ower than that of large vesicles and the range of preservation of spec ific activity was dependent on the lipid composition. Lipid combinatio ns of phosphatidylcholine and cholesterol and either stearylamine, pho sphatidylinositol or monosialoganglioside resulted in a high encapsula tion efficiency (40 and 98% in VET and sDRV, respectively), high stabi lity in saline and human serum (65-90% after 48 h) and considerable pr eservation of enzymatic activity (74-98%). The liposomal formulations were significantly less toxic than the free enzyme against normal CHO cells. In vivo toxicity, pharmacokinetics, biodistribution and antitum our activity studies are planned with the best formulations described in this paper.