α2,3-sialylation on human naïve T cells restrains bispecific engager-mediated anti-tumor immunity.

UNLABELLED: Aberrantly elevated cell-surface sialylation, or hypersialylation, is a common feature of human cancers and contributes to immune evasion. Sialidase-based therapies have therefore emerged as a strategy to disrupt this glyco-checkpoint. Although the immunosuppressive role of tumor-associated sialylation is well established, how sialylation on human T cells shapes anti-tumor responses remains poorly defined. Here, we identify surface sialoglycans on T cells, particularly α2,3-linked structures, as a cell-intrinsic restraint on human T cell activation, proliferation, and effector function. In vitro , enzymatic desialylation enhanced T cell activation, proliferation, cytokine production, and bispecific T cell engager (TCE)-mediated tumor-cell killing in healthy donor PBMC co-cultures. In ex vivo cultures of primary chronic lymphocytic leukemia (CLL) PBMCs, sialidase treatment combined with the CD20-directed TCE glofitamab enhanced cytotoxic effector transcriptional programming in autologous T cells. Single-cell RNA sequencing combined with lectin-based CITE-seq linked treatment-induced transcriptional states to lectin-defined cell-surface glycan signatures within the same single-cell dataset. This integrated analysis revealed that naïve and, to a lesser extent, central memory T cells combined elevated baseline α2,3-sialylation signatures with the clearest transcriptional responses to glofitamab plus sialidase treatment. CD43 emerged as a major carrier of α2,3-linked sialoglycans, and its deletion attenuated sialidase-enhanced T cell activation. Together, these findings identify sialylation of the T cell surface as a subset-specific restraint on human TCE responses and provide a rationale for testing sialidase-TCE combinations designed to engage less-differentiated T cell populations. ONE-SENTENCE SUMMARY: Desialylation enhances bispecific T cell engager responses by relieving a sialoglycan-dependent restraint in human T cells.