Abstract P65: Single Cell Profiling Reveals Unique TCR Repertoire and Transcriptomic Signatures of Epstein-Barr Virus-Specific T cells (EBVSTs) Armoured with Chimeric Antigen Receptors (CARs)

Abstract

Abstract Allogeneic Epstein-Barr Virus-specific T cells (EBVSTs) have thus far been a safe immunotherapy product in the clinic, with little graft-versus-host-disease reported in over 300 recipients to date. Consequently, EBVSTs armoured with chimeric antigen receptors (CARs) have emerged as a promising platform for allogeneic T cell therapies. Believing that a deep characterisation of CAR EBVST cell products can enhance insights into their clinical safety, we performed single cell T cell receptor (TCR) sequencing on CAR EBVSTs, taking CD30.CAR EBVSTs as a prototype. Non-transduced (NT) EBVSTs and CD30.CAR EBVSTs generated from three healthy donors exhibited a narrower, more focused TCR repertoire by diversity indices, compared to resting T cells and CD30.CAR-transduced activated T cells (ATCs) from the same donors. EBVSTs harboured a lower frequency of unique TCR clonotypes and a higher proportion of expanded clones, indicating a tendency toward clonal dominance. Narrower TCR repertoires corresponded to greater IFNγ and TNFα secretion in response to stimulation with EBV peptides, with no cross-reactivity toward other common viruses, and were associated with lower allogeneic reactivity in mixed lymphocyte reactions. To investigate whether the EBVST nature of the product might contribute to better CAR activity, we concurrently performed single cell transcriptomic sequencing on NT EBVSTs, CD30.CAR EBVSTs, and CD30.CAR ATCs. Transcriptomic signatures revealed that each sample type contained two major, distinct populations of CD4+ T cells: proliferative and activated. While the proliferative subsets appeared to be closely related, the activated populations had distinct transcriptional profiles. In particular, CD30.CAR EBVSTs appeared to be more polyfunctional and metabolically poised than CD30.CAR ATCs. The restricted TCR repertoire and apparent functional superiority of CD30.CAR EBVSTs may underlie their safety and efficacy observed in early-stage clinical trials and preclinical models, with the larger implication that these qualities extend to CAR EBVSTs targeting antigens beyond CD30. Citation Format: Lindsay Kua, Ting Wei Lim, Fiona Wong, Pei Yun Teo, Lionel Low, Ivan Horak, Jinmiao Chen, Kar Wai Tan. Single Cell Profiling Reveals Unique TCR Repertoire and Transcriptomic Signatures of Epstein-Barr Virus-Specific T cells (EBVSTs) Armoured with Chimeric Antigen Receptors (CARs) [abstract]. In: Proceedings of Frontiers in Cancer Science; 2023 Nov 6-8; Singapore. Philadelphia (PA): AACR; Cancer Res 2024;84(8_Suppl):Abstract nr P65.