STUDY OF IN VITRO STABILITY OF LIPOSOMES AND IN VIVO ANTIBODY RESPONSE TO ANTIGEN ASSOCIATED WITH LIPOSOMES CONTAINING GM1 AFTER ORAL AND SUBCUTANEOUS IMMUNIZATION

ABSTRACT

In order to evaluate the usefulness of liposomes as possible vaccine vehicles (oral and subcutaneous), the stability of liposomes in buffer, plasma and saliva at 25 and 37°C was analyzed via fluorescence and enzymatic methodology. The tested mixtures included [EggPC/Chol] 1 : 1 (mixture I), [EggPC/Chol/SM] 1 : 1 : 1 (mixture II), [EggPC/Chol/SM/GM typeIII] 1 : 1 : 1 : 0.14 (mixture III), [EggPC/Chol/SM/GM1] 1 : 1 : 1 : 0.14 (mixture IV) and [DIAPC/DMPC] 1 : 1 non polymerized (mixture V) and polymerized (mixture VI); all mole ratio. Liposome mixtures I and II were more stable in buffer at 25°C. On the other hand, mixtures III and IV were more stable in plasma at 37°C; mixture VI was more stable in plasma at 37°C than in buffer or saliva. Mixtures IV and V liposomes were both stable in saliva for at least one hour. Blood and feces anti-GM1 response to antigen associated liposomes after subcutaneous and oral administration was also examined. After mixture IV mice immunization, no detectable anti-ganglioside GM1 antibody response was detected. Negative stain transmission electron microscopy, shows that liposomes containing SM, GM1, GM typeIII and DIAPC : DMPC were twice the size of those made with EggPC/Chol. The hydrophobicity factor expressed as A(570/500) was obtained using the probe merocynine 540 (MC540). The order of fluidity increased from: mixture II<mixture I<mixture III<mixture IV<mixtureV<mixture VI. Although the high hydrophobicity factor for polymerizable lipids there are other factors like stability must be taken into account according to the administration via selected. Also, the hydrophilicity of the groups protruding from the membrane interphase into the solution in the case of subcutaneous inoculation is very relevant and for oral administration stability is the property to take into account, as long as they have to last through the different fluids of the gastrointestinal tract. The results obtained suggest that liposomes that showed stability in saliva and plasma at 37°C containing GM1, GM typeIII or DIAPC/DMPC would serve effectively as a delivery vehicle for oral and subcutaneous non-viral vaccines.

Keywords: :

• Immunoliposomes
• Gangliosides
• Polymerizable liposomes
• Stability
• Antigenicity