Charting of the tumor-reactive CD8+ T-cell repertoire in pancreatic ductal adenocarcinoma at the single cell level

Our prior research on the T-cell immune infiltrate in human pancreatic cancer provided initial evidence for tumor-reactive T-cells as well as in situ expanded T-cell clones (PMID 28123878 & 32303540). We therefore analyzed the T-cell fraction of 9 pancreatic tumor samples by means of single-cell sequencing and identified multiple T cell receptors (TCRs) mediating MHC-restricted T-cell reactivity against autologous tumor cells. The fraction of tumor-reactive TCRs was considerably greater in samples from genetically unstable tumors, pointing to the recognition of mutanome-encoded, neo-antigens. By comparing the transcriptome of T-cells bearing tumor-reactive versus non-reactive TCRs, we devised a gene signature that can be effectively used to distinguish between tumor-reactive and bystander T-cells in single-cell sequencing data sets, and therefore towards the targeted molecular cloning of tumor-reactive TCRs. Application of our signature, as developed on the basis of data from primary pancreatic tumors, on external data sets showed that it could be successfully applied to multiple tumor types, including lung and metastatic cancers.