Cell surface marker-based capture of neoantigen-reactive CD8+T-cell receptors from metastatic tumor digests

Abstract

BackgroundCellular immunotherapies using autologous tumor-infiltrating lymphocytes (TIL) can induce durable regression of epithelial cancers in selected patients with treatment-refractory metastatic disease. As the genetic engineering of T cells with tumor-reactive T-cell receptors (TCRs) comes to the forefront of clinical investigation, the rapid, scalable, and cost-effective detection of patient-specific neoantigen-reactive TIL remains a top priority.MethodsWe analyzed the single-cell transcriptomic states of 31 neoantigen-specific T-cell clonotypes to identify cell surface dysfunction markers that best identified the metastatic transcriptional states enriched with antitumor TIL. We developed an efficient method to capture neoantigen-reactive TCRs directly from resected human tumors based on cell surface co-expression of CD39, programmed cell death protein-1, and TIGIT dysfunction markers (CD8+TILTP).ResultsTILTPTCR isolation achieved a high degree of correlation with single-cell transcriptomic signatures that identify neoantigen-reactive TCRs, making it a cost-effective strategy using widely available resources. Reconstruction of additional TILTPTCRs from tumors identified known and novel antitumor TCRs, showing that at least 39.5% of TILTPTCRs are neoantigen-reactive or tumor-reactive. Despite their substantial enrichment for neoantigen-reactive TCR clonotypes, clonal dynamics of 24 unique antitumor TILTPclonotypes from four patients indicated that most in vitro expanded TILTPpopulations failed to demonstrate neoantigen reactivity, either by loss of neoantigen-reactive clones during TIL expansion, or through functional impairment during cognate neoantigen recognition.ConclusionsWhile direct usage of in vitro-expanded CD8+TILTPas a source for cellular therapy might be precluded by profound TIL dysfunction, isolating TILTPrepresents a streamlined effective approach to rapidly identify neoantigen-reactive TCRs to design engineered cellular immunotherapies against cancer.