Immunogenic Structural Features of a Breast Tumor Specific Epitope for Cancer Immunotherapy.

The objectives of the research program were to study the structure-immunogenicity relationships of a hypoglycosylated human tumor-specific mucin common to breast and other adenocarcinomas. Hypo-glycosylation of breast mucin leads to exposure of a tumor-specific epitope TSE. The structural and immunogenic properties of the TSE were examined using synthetic mucin peptides and recombinant mucin proteins that contain the TSE andor mutations in potential glycosylation sites surrounding the TSE. One TSE is the pentapeptide PDTRP of the 20 amino acid mucin tandem repeat sequence PDTRPAPGSTAPPAHGVTSA. Immunogenic cross-reactivity with tumor-specific monoclonal antibodies show that antigenicity is maximized with the 40 amino acid MUC1-mtr2. By contrast, the MUC1-mtr3 elicited a 9-fold better cellular response compared to either single or double tandem repeat peptides. The single, double and triple tandem repeat peptides show few elements of preferred conformation, based on 1H-NMR spectroscopy. A model of the distribution of the oligosaccharide side chains along the mucin core protein was developed showing that the carbohydrates may not surround the protein in a uniform coating. Thus, regions of the core protein are exposed under normal circumstances, yet tumor-specific epitopes remain masked by the sugars in the non-malignant cells.