Abstract 3960: Genome-wide CRISPR screens for genes inhibiting T cell-tumor cell interactions identify complex N-glycans

The efficacy of immunotherapy fundamentally depends on effective T cell-tumor cell interactions, yet the molecular determinants orchestrating this engagement are not fully understood. Here, we systematically mapped and perturbed key regulators of the T cell-tumor cell interactome using innovative cell-cell interaction assays. We performed genome-wide CRISPR screens to identify genes critically required for physical and functional interactions between T cells and tumor cells. Using TurboID proximity labeling and T cell-tumor cell conjugates, we identified the complex N-glycan biosynthesis pathway as a crucial regulator of T cell-tumor cell interactions. Recognizing the role of N-glycans in protein trafficking and surface retention, we subsequently performed cell surface proteomics and TurboID-based interactome profiling on complex N-glycan-deficient T cells, revealing increased expression of cell adhesion and co-stimulatory molecules. T cells with perturbed N-glycans show a shift towards an effector memory phenotype with elevated granzyme and integrin expression. In vivo, both genetic and pharmacological interventions that decrease the number of complex N-glycans on the surface of T cells enhanced intra-tumoral infiltration. Our findings provide a comprehensive functional atlas of molecules modulating T cell-tumor cell interactions and position the complex N-glycan biosynthesis pathway as a potential therapeutic target to strengthen T cell-mediated cytotoxicity in cancer immunotherapy. Citation Format: Adriaan F. de Groot, Zowi R. Huinen, Juan Simon Nieto, Daniel S. Peeper. Genome-wide CRISPR screens for genes inhibiting T cell-tumor cell interactions identify complex N-glycans [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3960.