Anti–PD-L1 glycoengineering enhances tumor killing through coordinated interactions between PD-L1⁺ DCs and CD16A⁺ cytotoxic lymphocytes

Blockade of the PD-1/PD-L1 axis has transformed cancer immunotherapy, yet many patients fail to achieve durable responses. Although Fc glycoengineering by defucosylation is well established to enhance NK cell-mediated antibody-dependent cellular cytotoxicity through increased FcγRIIIa binding, its effects on other FcγRIIIa-expressing immune cells remain poorly understood. Using complementary human in vitro dendritic cell-lymphocyte coculture models, we compared non-glycosylated, wild-type, and defucosylated anti-PD-L1 antibody variants. Defucosylation enhanced FcγRIIIa binding and increased the relative frequency and activation of cytotoxic lymphocytes, particularly CD3⁺CD56⁺ NKT-like cells. Our findings identify a previously unrecognized mechanism whereby coordinated engagement of PD-L1 on dendritic cells and FcγRIIIa on lymphocytes alters cytotoxic lymphocyte composition. The changes in cytotoxic lymphocyte composition and activation induced by defucosylated anti-PD-L1 were associated with enhanced tumor killing in both human 2D and 3D cancer models, supporting Fc glycoengineering as a potential strategy to improve cancer immunotherapy.