Adoptive cell transfer of T-cell receptor (TCR)-engineered T cells has emerged as a powerful and potentially curative therapy for cancers and infectious diseases. However, how to rapidly identify and obtain therapeutic TCRs remains a major challenge in this field. Here, we simultaneously performed deep single-cell RNA and TCR V(D)J sequencing on 879 single T cells from in vitro stimulated CMV pp65NLV-specific CD8+ T cells. The correlation analysis of single-cell transcriptome and TCR V(D)J sequences revealed a differential distribution of T cell transcriptomic profiles among TCR clones. Two distinct cell types were identified in the T cell population. TYPE I T cell subtype, which represents the gene signatures of T cell activation and cytotoxicity, mainly includes TCR clone 1, 2 & 4, while TYPE II subtype, which represents the gene signatures of naïve status, mainly includes TCR clone 3 and other low frequency clones. Based on their temporal phases of T cell activation, the selected TCR sequences from clone 1 (TCRZWX1) and clone 3 (TCRZWX3) were functionally validated by constructing TCR-engineered T cells in vitro. The two clones showed distinct cell stages corresponding to their transcriptome patterns. TCRZWX1 transgenic T cells displayed effector functions including secretion of pro-inflammatory cytokines, IFN-γ and TNF-α, and cytotoxicity in response to CMV pp65NLV peptide. However, TCRZWX3 transgenic T cells displayed non-detectable levels of IFN-γ, which might correlate with a low affinity TCR. Our results provide a novel approach using correlative analysis of the single-cell transcriptome and TCR V(D)J sequences for rapidly identifying potential therapeutic TCRs for T cell therapy.

Citation Format: Linnan Zhu, Fei Wang, Qumiao Xu, Hai-Xi Sun, Ziyi Li, Yanling Liang, Zhenkun Zhuang, Ying Gu, Cheng-chi Chao. Rapid generation of TCR-engineered T lymphocytes by linking the single cell transcriptome to its corresponding T cell receptor in antigen specific T cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6600.