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Rapid cloning of antigen-specific T-cell receptors by leveraging the cis activation of T cells

Nature Biomedical Engineering volume 6, pages 806–818 (2022)Cite this article

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Abstract

It is commonly understood that T cells are activated via trans interactions between antigen-specific T-cell receptors (TCRs) and antigenic peptides presented on major histocompatibility complex (MHC) molecules on antigen-presenting cells. By analysing a large number of T cells at the single-cell level on a microwell array, we show that T-cell activation can occur via cis interactions (where TCRs on the T cell interact with the antigenic peptides presented on MHC class-I molecules on the same cell), and that such cis activation can be used to detect antigen-specific T cells and clone their TCR within 4 d. We used the detection-and-cloning system to clone a tumour-antigen-specific TCR from peripheral blood mononuclear cells of healthy donors. TCR cloning by leveraging the cis activation of T cells may facilitate the development of TCR-engineered T cells for cancer therapy.

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Fig. 1: Detection of cytokine-secreting T cells in a microwell-array chip.
Fig. 2: T-cell activation via cis interactions of TCRs and pMHC-I molecules on a single T cell using the microwell-array chip.
Fig. 3: Molecular analysis of T-cell activation through cis and trans interactions with pMHC molecules.
Fig. 4: Interaction of TCR and pMHC molecules in cis or trans.
Fig. 5: A model for the cis interaction between the TCR and the antigenic pMHC-I complex on the lipid-bilayer surface.
Fig. 6: T-cell activation through cis or trans interactions of TCR and pMHC molecules in human cells.
Fig. 7: Development of the T-ISAAC system for human TCR cloning based on the cis activation of T cells.
Fig. 8: Cloning of a WT1-specific TCR from PBMCs of healthy donors with T-ISAAC.

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Data availability

The main data supporting the results in this study are available within the paper and its Supplementary Information. The cDNA sequences of the TCRs are available from the DNA Data Bank of Japan, under the set of accession codes LC663613–LC663622. The raw data generated during the study are available for research purposes from the corresponding author on reasonable request.

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Acknowledgements

Melanoma-associated antigen (MAGEA4)-specific TCR (2–28) expression vectors were kindly provided by H. Shiku (Mie University). TCR sequence data of Wilms tumour 1 (WT1)-specific TCR (TAK1) were kindly provided by M. Yasukawa (Ehime University), PLAT-E cell line by T. Kitamura (University of Tokyo), human CD8-expressing TG40 cell line by T. Ueno (Kumamoto University) with permission from T. Saito (Riken), T2-A24 cell line by K. Kuzushima (Aichi Cancer Center Research Institute) and Phoenix-A cell line by G. Nolan (Stanford University). We thank T. Katagiri (University of Toyama) for helpful discussion. This research was supported by Practical Research for Innovative Cancer Control, Project for Cancer Research and Therapeutic Evolution (P-CREATE) from AMED under Grant Number JP17cm0106417 (E.K.) and JP20cm0106371 (E.K.), and JSPS KAKENHI grant numbers 18K19441 (H.K.), 16H06499 (H.K.) and 16H06500 (S.Y.), 21K18261 (H.K.), 21H02782 (E.K.) and 21H02966 (A.M.)

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Author notes

  1. These authors contributed equally: Eiji Kobayashi, Aishun Jin.

Authors and Affiliations

  1. Department of Immunology, Faculty of Medicine, Academic Assembly, University of Toyama, Toyama, Japan

    Eiji Kobayashi, Aishun Jin, Hiroshi Hamana, Kiyomi Shitaoka, Kazuto Tajiri, Tatsuhiko Ozawa, Atsushi Muraguchi & Hiroyuki Kishi

  2. Department of Immunology, Chongqing Medical University, Chongqing, China

    Aishun Jin

  3. Department of Immunology, Harbin Medical University, Harbin, China

    Aishun Jin

  4. Department of Immunology, Hiroshima University, Hiroshima, Japan

    Kiyomi Shitaoka

  5. The Third Department of Internal Medicine, Faculty of Medicine, Academic Assembly, University of Toyama, Toyama, Japan

    Kazuto Tajiri

  6. RIKEN Cluster for Science, Technology and Innovation Hub, Yokohama, Japan

    Seisuke Kusano & Shigeyuki Yokoyama

  7. Toyama Industrial Technology Research and Development Center, Takaoka, Japan

    Tsutomu Obata

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Contributions

E.K., H.K. and A.M. proposed the concept of cis interaction of TCR with pMHC molecules and of the T-ISAAC system. E.K., H.K. and A.J. designed the research studies. H.H. produced DNA constructs. T. Obata produced microwell-array chips. E.K., H.K., A.J., H.H., K.S., K.T. and T. Ozawa performed the research and analysed the data. S.K. and S.Y. built the structure model. A.J., H.K., E.K and A.M. discussed the results and implications. E.K., H.K. and A.M. wrote the paper. All authors read and reviewed the manuscript.

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Correspondence to Hiroyuki Kishi.

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H.K and A.M. are directors of SC World, Inc. The other authors declare no competing interests.

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Kobayashi, E., Jin, A., Hamana, H. et al. Rapid cloning of antigen-specific T-cell receptors by leveraging the cis activation of T cells. Nat. Biomed. Eng 6, 806–818 (2022). https://doi.org/10.1038/s41551-022-00874-6

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