Chemoenzymatically synthesized multimeric Tn/STn MUC1 glycopeptides elicit cancer-specific anti-MUC1 antibody responses and override tolerance

TLDR

MUC1 mucin is a prime target antigen for cancer immunotherapy because it is abundantly expressed and aberrantly glycosylated in carcinomas, yet peptide immunizations have largely failed due to tolerance. The study aimed to develop a chemoenzymatic synthesis of extended MUC1 TR glycopeptides bearing cancer‑associated O‑glycans via recombinant human glycosyltransferases. The authors synthesized glycopeptides with varying Tn/STn densities, conjugated them to KLH, and evaluated them as immunogens to identify an optimal vaccine design. Glycopeptides fully occupied with O‑glycans induced the strongest, cancer‑cell–specific antibody responses in mice, targeting both peptide and carbohydrate epitopes, and a monoclonal antibody (5E5) with comparable specificity was produced, underscoring the design’s diagnostic and immunopreventive promise.

Abstract

The MUC1 mucin represents a prime target antigen for cancer immunotherapy because it is abundantly expressed and aberrantly glycosylated in carcinomas. Attempts to generate strong humoral immunity to MUC1 by immunization with peptides have generally failed partly because of tolerance. In this study, we have developed chemoenzymatic synthesis of extended MUC1 TR glycopeptides with cancer-associated O-glycosylation using a panel of recombinant human glycosyltransferases. MUC1 glycopeptides with different densities of Tn and STn glycoforms conjugated to KLH were used as immunogens to evaluate an optimal vaccine design. MUC1 glycopeptides with complete O-glycan occupancy (five sites per repeat) elicited the strongest antibody response reacting with MUC1 expressed in breast cancer cell lines in both Balb/c and MUC1.Tg mice. The elicited humoral immune response showed remarkable specificity for cancer cells suggesting that the glycopeptide design holds promise as a cancer vaccine. The elicited immune responses were directed to combined glycopeptide epitopes, and both peptide sequence and carbohydrate structures were important for the antigen. A MAb (5E5) with similar specificity as the elicited immune response was generated and shown to have the same remarkable cancer specificity. This antibody may hold promise in diagnostic and immunopreventive measures.