The lack of
antibodies with sufficient
cancer selectivity is currently limiting the treatment of solid
tumors by
immunotherapies. Most current immunotherapeutic targets are
tumor-associated
antigens that are also found in healthy tissues and often do not display sufficient
cancer selectivity to be used as targets for potent antibody-based immunotherapeutic treatments, such as
chimeric antigen receptor (CAR) T cells. Many solid
tumors, however, display aberrant glycosylation that results in expression of
tumor-associated carbohydrate antigens that are distinct from healthy tissues. Targeting aberrantly glycosylated
glycopeptide epitopes within existing or novel
glycoprotein targets may provide the
cancer selectivity needed for
immunotherapy of solid
tumors. However, to date only a few such
glycopeptide epitopes have been targeted. Here, we used O-glycoproteomics data from multiple cell lines to identify a
glycopeptide epitope in CD44v6, a
cancer-associated CD44
isoform, and developed a
cancer-specific mAb, 4C8, through a
glycopeptide immunization strategy. 4C8 selectively binds to Tn-glycosylated CD44v6 in a site-specific manner with low nanomolar affinity. 4C8 was shown to be highly
cancer specific by IHC of sections from multiple healthy and cancerous tissues. 4C8 CAR T cells demonstrated target-specific cytotoxicity in vitro and significant
tumor regression and increased survival in vivo. Importantly, 4C8 CAR T cells were able to selectively kill target cells in a mixed organotypic
skin cancer model having abundant CD44v6 expression without affecting healthy keratinocytes, indicating tolerability and safety.