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Characterization of the T-cell receptor beta chain repertoire in tumor-infiltrating lymphocytes.

ABSTRACT: Tumor-infiltrating lymphocytes (TILs) are direct effectors of tumor immunity, and their characterization is important for further development of immunotherapy. Recent advances in high-throughput sequencing technologies have enabled a comprehensive analysis of T-cell receptor (TCR) complementarity-determining region 3 (CDR3) sequences, which may provide information of therapeutic importance. We developed a high-fidelity target sequencing method with the ability for absolute quantitation, and performed large-scale sequencing of TCR beta chain (TCRB) CDR3 regions in TILs and peripheral blood lymphocytes (PBLs). The estimated TCRB repertoire sizes of PBLs from four healthy individuals and TILs from four colorectal cancer tissue samples were 608,664-1,003,098 and 90,228-223,757, respectively. The usage of J- and V-regions was similar in PBLs and TILs. Proportions of CDR3 amino acid (aa) sequences occupying more than 0.01% of the total molecular population were 0.33-0.43% in PBLs and 1.3-3.6% in TILs. Additional low coverage sequencing of 15 samples identified five CDR3 aa sequences that were shared by nine patients, one sequence shared by 10 patients, and one sequence shared by 12 patients. The estimated size of the TCRB repertoire in TILs was significantly smaller than that in PBLs. The proportion of abundant species (>0.01%) in TILs was larger than that in PBLs. Shared CDR3 aa sequences represent a response to common antigens, and the identification of such CDR3 sequences may be beneficial in developing clinical biomarkers.

SUBMITTER: Nakanishi K

PROVIDER: S-EPMC5055180 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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Characterization of the T-cell receptor beta chain repertoire in tumor-infiltrating lymphocytes.

Nakanishi Katsumi K Kukita Yoji Y Segawa Hidenobu H Inoue Norimitsu N Ohue Masayuki M Kato Kikuya K

Cancer medicine 20160727 9

Tumor-infiltrating lymphocytes (TILs) are direct effectors of tumor immunity, and their characterization is important for further development of immunotherapy. Recent advances in high-throughput sequencing technologies have enabled a comprehensive analysis of T-cell receptor (TCR) complementarity-determining region 3 (CDR3) sequences, which may provide information of therapeutic importance. We developed a high-fidelity target sequencing method with the ability for absolute quantitation, and perf ...[more]

PMID: 27465739

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Project description:The successful use of expanded tumor-infiltrating lymphocytes (TIL) in adoptive TIL therapies has been reported, but the effects of the TIL expansion, immunophenotype, function, and T cell receptor (TCR) repertoire of the infused products relative to the tumor microenvironment (TME) are not well understood. In this study, we analyzed the tumor samples (n = 58) from treatment-naïve patients with renal cell carcinoma (RCC), "pre-rapidly expanded" TILs (pre-REP TIL, n = 15) and "rapidly expanded" TILs (REP TIL, n = 25) according to a clinical-grade TIL production protocol, with single-cell RNA (scRNA)+TCRαβ-seq (TCRαβ sequencing), TCRβ-sequencing (TCRβ-seq), and flow cytometry. REP TILs encompassed a greater abundance of CD4+ than CD8+ T cells, with increased LAG-3 and low PD-1 expressions in both CD4+ and CD8+ T cell compartments compared with the pre-REP TIL and tumor T cells. The REP protocol preferentially expanded small clones of the CD4+ phenotype (CD4, IL7R, KLRB1) in the TME, indicating that the largest exhausted T cell clones in the tumor do not expand during the expansion protocol. In addition, by generating a catalog of RCC-associated TCR motifs from >1,000 scRNA+TCRαβ-seq and TCRβ-seq RCC, healthy and other cancer sample cohorts, we quantified the RCC-associated TCRs from the expansion protocol. Unlike the low-remaining amount of anti-viral TCRs throughout the expansion, the quantity of the RCC-associated TCRs was high in the tumors and pre-REP TILs but decreased in the REP TILs. Our results provide an in-depth understanding of the origin, phenotype, and TCR specificity of RCC TIL products, paving the way for a more rationalized production of TILs.SignificanceTILs are a heterogenous group of immune cells that recognize and attack the tumor, thus are utilized in various clinical trials. In our study, we explored the TILs in patients with kidney cancer by expanding the TILs using a clinical-grade protocol, as well as observed their characteristics and ability to recognize the tumor using in-depth experimental and computational tools.

Dataset Information

Characterization of the T-cell receptor beta chain repertoire in tumor-infiltrating lymphocytes.

Publications

Characterization of the T-cell receptor beta chain repertoire in tumor-infiltrating lymphocytes.

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