LIPOSOMES AS VACCINE CARRIERS - INCORPORATION OF SOLUBLE AND PARTICULATE ANTIGENS IN GIANT VESICLES

Giant liposomes (mean diameter 5.5 mu m) composed of egg phosphatidylc holine or distearoyl phosphatidylcholine, phosphatidyl glycerol, chole sterol and triolein were prepared by a double emulsion technique. They were then mixed with model particulate (killed Bacillus subtilis, and killed Bacille calmette-Guerin) and soluble (tetanus toroid) vaccines and freeze-dried. Rehydration of the powder resulted in the generatio n of vesicles of similar mean diameter and diameter range, containing up to 27% (mean value) of the materials used for entrapment. Separatio n of entrapped from non-entrapped material was carried out by sucrose gradient centrifugation (B. subtilis and BCG) or centrifugation at 600 xg (toxoid). Light microscopy of liposomes containing B. subtilis lab elled with fluorescein isothiocyanate revealed the presence of bacteri a in individual vesicles which, in separate studies, were also found t o entrap later particles (0.5 and 1.0 mu m diameter). Bacteria-contain ing liposomes could be freeze-dried in the presence of trehalose with most (83-87%) of the entrapped material recovered within the vesicles on reconstitution with saline. Liposomes were also shown to retain qua ntitatively their content of B. subtilis and, to a lesser extent, toro id in the presence of mouse plasma at 37 degrees C and in situ after i ntramuscular injection into mice, for up to 24 h. Since liposomes are known (Gregoriadis, G. (1990) Immunol. Today 11, 89) to act as immunol ogical adjuvants and vaccine carriers, giant vesicles containing micro bes (live or attenuated if needed since the conditions of entrapment a re mild) and, when appropriate, soluble antigens, could be used as mul tiple vaccines to ensure simultaneous presentation of antigens to immu nocompetent cells.