Identification of tumoricidal TCRs from tumor-infiltrating lymphocytes by single-cell analysis

Kiyomi Shitaoka, Hiroshi Hamana, Hiroyuki Kishi, Yoshihiro Hayakawa, Eiji Kobayashi, Kenta Sukegawa, Xiuhong Piao, Fulian Lyu, Takuya Nagata, Daisuke Sugiyama, Hiroyoshi Nishikawa, Atsushi Tanemura, Ichiro Katayama, Mutsunori Murahashi, Yasushi Takamatsu, Kenzaburo Tani, Tatsuhiko Ozawa, Atsushi Muraguchi

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Abstract

T-cell receptor (TCR) gene therapy is a promising next-generation antitumor treatment. We previously developed a single–T-cell analysis protocol that allows the rapid capture of paired TCRa and b cDNAs. Here, we applied the protocol to analyze the TCR repertoire of tumor-infiltrating lymphocytes (TIL) of various cancer patients. We found clonally expanded populations of T cells that expressed the same clonotypic TCR in 50% to 70% of CD137þCD8þ TILs, indicating that they responded to certain antigens in the tumor environment. To assess the tumor reactivity of the TCRs derived from those clonally expanded TILs in detail, we then analyzed the CD137þCD8þ TILs from the tumor of B16F10 melanoma cells in six C57BL/6 mice and analyzed their TCR repertoire. We also found clonally expanded T cells in 60% to 90% of CD137þCD8þ TILs. When the tumor reactivity of dominant clonotypic TCRs in each mouse was analyzed, 9 of 13 TCRs induced the secretion of IFNg in response to, and showed killing of, B16F10 cells in vitro, and 2 of them showed strong antitumor activity in vivo. Concerning their antigen specificity, 7 of them reacted to p15E peptide of endogenous murine leukemia virus-derived envelope glycoprotein 70, and the rest reacted to tumor-associated antigens expressed on EL4 lymphoma as well as B16 melanoma cells. These results show that our strategy enables us to simply and rapidly obtain the tumor-specific TCR repertoire with high fidelity in an antigen- and MHC haplotype–independent manner from primary TILs.

Original languageEnglish
Pages (from-to)378-388
Number of pages11
JournalCancer Immunology Research
Volume6
Issue number4
DOIs
Publication statusPublished - Apr 2018

All Science Journal Classification (ASJC) codes

  • Immunology
  • Cancer Research

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    Shitaoka, K., Hamana, H., Kishi, H., Hayakawa, Y., Kobayashi, E., Sukegawa, K., Piao, X., Lyu, F., Nagata, T., Sugiyama, D., Nishikawa, H., Tanemura, A., Katayama, I., Murahashi, M., Takamatsu, Y., Tani, K., Ozawa, T., & Muraguchi, A. (2018). Identification of tumoricidal TCRs from tumor-infiltrating lymphocytes by single-cell analysis. Cancer Immunology Research, 6(4), 378-388. https://doi.org/10.1158/2326-6066.CIR-17-0489