Cord blood–derived V <sub>δ</sub> 2 <sup>+</sup> and V <sub>δ</sub> 2 <sup>−</sup> T cells acquire differential cell state compositions upon in vitro expansion
Jeremy Wee Kiat Ng, Kar Wai Tan, Dian Yan Guo, Joey Jia Hui Lai, Xiubo Fan, Zhiyong Poon, Tse Hui Lim, Alvin Soon Tiong Lim, Tony Kiat Hon Lim, William Ying Khee Hwang, Shang Li, Connie J. Eaves, Yeow Tee Goh, Alice M.S. Cheung
Abstract
Human cord blood–derived γδ T cells (CB γδ ) display a highly diverse TCR γδ repertoire and have a unique subtype composition different from fetal or adult peripheral blood counterparts. We expanded CB γδ in vitro using an irradiated Epstein-Barr virus–transformed feeder cell–based modified rapid expansion protocol (REP). Single-cell RNA sequencing tracked progressive differentiation of naïve CB γδ into cells expressing neoantigen-reactive tumor-infiltrating lymphocyte as well as tissue-resident memory precursor–like and antigen-presenting cell–like gene signatures. TCR γδ clonal tracing revealed a bias toward cytotoxic effector differentiation in a much larger proportion of V δ 2 − clones compared to V δ 2 + clones, resulting in the former being more cytotoxic at the population level. These clonotype-specific differentiation dynamics were not restricted to REP and were recapitulated upon secondary nonviral antigen stimulations. Thus, our data showed intrinsic cellular differences between major subtypes of human γδ T cells already in operation at early postnatal stage and highlighted key areas of consideration in optimizing cell manufacturing processes.