Investigations into a mucosal targeted breast cancer vaccine: Fusion of the epithelial mucin 1 antigen to a mucosal carrier peptide and its efficacy in breaking self-antigen tolerance

Most cancer vaccine studies have focused on eliciting a systemic, cellular immune response and have ignored the mucosal and humoral routes. It is plausible that mucosal induction of both cellular and humoral responses will strengthen an anti-tumor immune response. The human epithelial mucin MUC1 is a tumor-associated antigen whose altered glycosylation pattern on tumor cells reveals an immunodominant peptide sequence along its tandem repeat (TR) that has been used as a target for tumor immunotherapy. Two bacterial products with great potential to function as mucosal carriers for subunit vaccines are the B subunits of cholera toxin (CTB) and the Escherichia coli heat labile enterotoxin (LTB). In the present study we evaluated whether a MUC1 TR peptide chemically linked to CTB (CTB-MUC1) and translationally fused to LTB (LTB-MUC1) would break self-antigen tolerance in the transgenic MUC1 tolerant mouse model (MUC1.Tg) via oral or intraperitoneal immunizations.; Oral immunization with the CTB-MUC1 conjugate along with mucosal adjuvants did not break self-antigen tolerance in MUC1.Tg mice, but induced a strong humoral response in wild-type C57BL/6 mice. However, self-antigen tolerance in the MUC1.Tg mouse model was broken after intraperitoneal immunizations with different doses of the CTB-MUC1 conjugate protein alone or co-delivered with CpG oligodeoxynucleotides (ODNs). Importantly, intraperitoneal immunization with CTB-MUC1 and CpG ODNs caused decreased tumor burden when mice were challenged with a mammary tumor cell line that expresses human MUC1.; We further report the production of LTB-MUC1 in a plant expression system. We compared LTB-MUC1 expression in stably transformed plants to the magnICON(TM) deconstructed viral vector system and found significant differences in expression level and posttranslational modifications. Plant expression, particularly from the magnICON(TM) transient system, generated glycosylated forms of LTB-MUC1. We tested the magnICON(TM) derived LTB-MUC1 in MUC1.Tg mice and showed reversal of MUC1 self-antigen tolerance after intraperitoneal immunizations. This is the first study to report the expression of a human breast cancer antigen in a plant system and the first report of a plant-made protein breaking self-antigen tolerance in a transgenic mouse model.