Chemoenzymatic synthesis of derivatives of a T-cell-stimulating peptide which carry tumor-associated carbohydrate antigens

The Tn (GalNAc alpha -Ser/Thr), T [Gal beta (1 -->3)GalNAc alpha -Ser/Thr], sialyl-Tn [Neu5Ac alpha (2 -->6)GalNAc alpha -Ser/Thr] and 2,3-sialyl-T [Neu5Ac alpha (2 -->3)Gal beta (1 -->3)GalNAc alpha -Ser/Thr] antigens are examples of tumor-associated carbohydrate antigens expressed by epithelial cancers. We now describe the preparation of 2-bromoethyl glycosides corresponding to the Tn and T antigens in one and five chemical steps (51 and 15% total yield), respectively, starting from N-acetylgalactosamine. The 2-bromoethyl Tn and T glycosides were used to alkylate a homocysteine residue incorporated in a peptide that is able to bind to class I MHC molecules on antigen-presenting cells. The two neoglycopeptides were then converted into glycopeptides which carry the sialyl-Tn and 2,3-sialyl-T antigens by using recombinant sialyltransferases. Interestingly, the sialyltransferases were able to sialylate the Tn and T carbohydrate moieties even though they were linked to the peptide backbone via a spacer instead of being attached to serine or threonine. The four glycopeptides will be used in studies directed towards inducing a carbohydrate-specific T cell response against the Tn, T, sialyl-Tn, and 2,3-sialyl-T antigens.