Detecting Tumor Specific Antigen-Reactive T cells from Tumor Infiltrating Lymphocytes via Interaction Dependent Fucosyl-biotinylation

Summary Re-activation and clonal expansion of tumor specific antigen (TSA)-reactive T cells are critical to the success of checkpoint blockade and adoptive transfer of tumor-infiltrating lymphocyte (TIL) based therapies. There are no reliable markers to specifically identify the repertoire of TSA-reactive T cells due to their heterogeneous composition. Here we introduce FucoID as a general platform to detect endogenous antigen-specific T cells and study their biology. Through this interaction dependent labeling approach, TSA-reactive T cells can be detected and separated from bystander T cells in primary tumor digests based on their cell-surface enzymatic fucosyl-biotinylation. Compared to bystander TILs, TSA-reactive TILs possess a distinct TCR repertoire and unique gene features. Though exhibiting a dysfunctional phenotype, this subset of TILs possesses substantial capabilities of proliferation and tumor specific killing. FucoID features genetic manipulation-free procedures and a quick turnover cycle, and therefore should have the potential of accelerating the pace of personalized cancer treatment. Highlights Interaction dependent fucosylation enables the detection and isolation of bona fide intratumoral tumor specific antigen-reactive T cells Tumor specific antigen-reactive CD8 + T cells possess capabilities to be expanded and adoptively transferred for tumor control Tumor specific antigen-reactive CD8 + T cells feature oligoclonal expansion and upregulate genes for the steroid biosynthesis and metabolic process Intratumoral bystander CD8 + T cells can be separated into two groups based on PD-1 expression that feature distinct gene modules