Development of liposomal antigen delivery system for synthetic MUC1 peptides

by Guan, Holly H.

Synthetic MUCI peptides are potential targets for active specific immunotherapy of cancer. This thesis describes the development of a liposomal delivery system for synthetic MUCl peptides favoring type 1 immune responses. A high performance liquid chrornatographic rnethodology was developed to quantify MUCI peptides in liposomes. Encapsulation efficiencies of hydrophilic MUCl peptides were significantly improved in charged liposomes under low ionic strength conditions. More than 75 percent of MUCl peptides were retained in liposomes during 37OC incubation in 100 percent mouse or human serum for 24 h, and 85 percent were retained at 4OC in PBS for at least 12 months. Liposomal formulations were characterized for surface expression of MUCl peptide epitopes. Liposomes encapsulating N- terminal acylated MUCl peptide, but not unmodified MUCl peptide or C- terminal acylated peptide, showed MUCl epitopes on the liposome surface. The nature of MUCl peptide formulation containing monophosphoryl lipid A (MPLA) profoundly influenced both cellular and humoral immune responses. Strong and MUC1 -specific T cell proliferative responses were elicited by liposome-associated rather than non-associated peptides, suggesting a requirement of peptide association with liposomes for effective delivery to antigen presenting cells and induction of T cell responses. The exact location of the peptide in liposomes was not critical for the type and magnitude of T cell responses. 60th masked and surface-exposed peptide liposome formulations elicited strong and MUC1-specific T cell responses, accompanied with high levels of IFN-y, but not IL-4. However, the location of peptide in liposomes significantly influenced the humoral immune response. Only surface-exposed MUCl liposomes induced high levels of anti-MUCI antibodies dominated by the lgG2b isotype. Over the dose range of the peptide tested, human MUCl specific T cell responses did not exhibit dose-dependency in BALBlc or C57BU6 mice. The effect of phospholipid composition and size of liposomes on T cell responses was studied using a mouse MUCI peptide (tandem repeat consensus sequences) as the antigen. This peptide induced antigenspecific T cell responses in a dose-dependent fashion (range 12-100 pg per mouse) in BALB/c mice. This homologous T cell response was independent of phospholipid composition and liposome size. In summary, strong and MUCl specific type 1 associated immune responses were elicited by MPLA containing MUCl liposomes. A liposomal delivery system can be tailored to preferentially induce cellular and/or humoral immune responses. The results of this study are expected to advance knowledge in vaccine design.