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Design of T-cell receptor libraries with diverse binding properties to examine adoptive T-cell responses

Gene Therapy volume 20, pages 634–644 (2013)Cite this article

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Abstract

Adoptive T-cell therapies have shown significant promise in the treatment of cancer and viral diseases. One approach, which introduces antigen-specific T-cell receptors (TCRs) into ex vivo activated T cells, is designed to overcome central tolerance mechanisms that prevent responses by endogenous T-cell repertoires. Studies have suggested that use of higher-affinity TCRs against class I major histocompatibility complex antigens could drive the activity of both CD4+ and CD8+ T cells, but the rules that govern the TCR binding optimal for in vivo activity are unknown. Here, we describe a high-throughput platform of ‘reverse biochemistry’ whereby a library of TCRs with a wide range of binding properties to the same antigen is introduced into T cells and adoptively transferred into mice with antigen-positive tumors. Extraction of RNA from tumor-infiltrating lymphocytes (TILs) or lymphoid organs allowed high-throughput sequencing to determine which TCRs were selected in vivo. The results showed that CD8+ T cells expressing the highest-affinity TCR variants were deleted in both the TIL population and in peripheral lymphoid tissues. In contrast, these same high-affinity TCR variants were preferentially expressed within CD4+ T cells in the tumor, suggesting they had a role in antigen-specific tumor control. The findings thus revealed that the affinity of the transduced TCRs controlled the survival and tumor infiltration of the transferred T cells. Accordingly, the TCR library strategy enables rapid assessment of TCR-binding properties that promote peripheral T-cell survival and tumor elimination.

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Acknowledgements

We would like to thank Janie Frye for her assistance with RNA isolation, Tom Gajewski for his gift of B16-SIY, Barbara Pilas and Ben Montez in the University of Illinois Flow Cytometry Facility for assistance with in vitro cell sorting, and Alvaro Hernandez and Chris Wright at the University of Illinois High-Throughput Sequencing and Genotyping Unit for assistance with 454 sequence analysis. This work was supported by NIH grant CA097296 (to DMK and HS) and a grant from the Melanoma Research Alliance (to DMK). BE was supported by a Research Fellowship of the DFG; JDS was supported by the Samuel and Ruth Engelberg/Irvington Institute Fellowship of the Cancer Research Institute.

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  1. A S Chervin, J D Stone and C M Soto: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry, University of Illinois, Urbana, IL, USA

    A S Chervin, J D Stone & D M Kranz

  2. Neuroscience Program, University of Illinois, Urbana, IL, USA

    C M Soto & E J Roy

  3. Department of Pathology, Committee on Immunology, University of Chicago, Chicago, IL, USA

    B Engels & H Schreiber

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  1. A S Chervin
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  2. J D Stone
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  4. B Engels
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Correspondence to D M Kranz.

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Chervin, A., Stone, J., Soto, C. et al. Design of T-cell receptor libraries with diverse binding properties to examine adoptive T-cell responses. Gene Ther 20, 634–644 (2013). https://doi.org/10.1038/gt.2012.80

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