Recognition of human gastrointestinal cancer neoantigens by circulating PD-1+ lymphocytes
Alena Gros, … , James C. Yang, Steven A. Rosenberg
Alena Gros, … , James C. Yang, Steven A. Rosenberg
Published October 14, 2019
Citation Information: J Clin Invest. 2019;129(11):4992-5004. https://doi.org/10.1172/JCI127967.
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Research Article Immunology Oncology

Recognition of human gastrointestinal cancer neoantigens by circulating PD-1+ lymphocytes

Abstract

Tumor-resident lymphocytes can mount a response against neoantigens expressed in microsatellite-stable gastrointestinal (GI) cancers, and adoptive transfer of neoantigen-specific lymphocytes has demonstrated antitumor activity in selected patients. However, whether peripheral blood could be used as an alternative minimally invasive source to identify lymphocytes targeting neoantigens in patients with GI cancer with relatively low mutation burden is unclear. We used a personalized high-throughput screening strategy to investigate whether PD-1 expression in peripheral blood could be used to identify CD8+ or CD4+ lymphocytes recognizing neoantigens identified by whole-exome sequencing in 7 patients with GI cancer. We found that neoantigen-specific lymphocytes were preferentially enriched in the CD8+PD-1+/hi or CD4+PD-1+/hi subsets, but not in the corresponding bulk or PD-1– fractions. In 6 of 7 individuals analyzed we identified circulating CD8+ and CD4+ lymphocytes targeting 6 and 4 neoantigens, respectively. Moreover, neoantigen-reactive T cells and a T cell receptor (TCR) isolated from the CD8+PD-1+ subsets recognized autologous tumor, albeit at reduced levels, in 2 patients with available cell lines. These data demonstrate the existence of circulating T cells targeting neoantigens in GI cancer patients and provide an approach to generate enriched populations of personalized neoantigen-specific lymphocytes and isolate TCRs that could be exploited therapeutically to treat cancer.

Authors

Alena Gros, Eric Tran, Maria R. Parkhurst, Sadia Ilyas, Anna Pasetto, Eric M. Groh, Paul F. Robbins, Rami Yossef, Andrea Garcia-Garijo, Carlos A. Fajardo, Todd D. Prickett, Li Jia, Jared J. Gartner, Satyajit Ray, Lien Ngo, John R. Wunderllich, James C. Yang, Steven A. Rosenberg

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Figure 3

Autologous tumor recognition by circulating neoantigen-specific lymphocytes or TCR-transduced cells from NCI-4110 and NCI-4177.

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Autologous tumor recognition by circulating neoantigen-specific lymphocy...
(A and B) Reactivity of DNAJB1-reactive lymphocytes (A) or PBLs transduced with a DNAJB1-specific TCR isolated from NCI-4110 (see Supplemental Figure 5) (B) following coculture with APCs pulsed with WT or MUT DNAJB1 25-mers, or with TC4110. Percentage of transduced cells was quantified using an antibody targeting the mouse TCRβ constant region (mTCRB). The percentage of live CD3+ cells expressing 4-1BB is shown. (C and D) Reactivity of TMG1-reactive lymphocytes isolated from the CD8+PD-1hi peripheral blood subset of patient NCI-4177 (see Supplemental Figure 4) to WT or MUT MMP14p.R158C 25-mers, irrelevant TMG RNA, or 4177-TMG1 RNA, encoding MMP14p.R158C (C), or to irrelevant TMG RNA, 4177-TMG1 RNA, or TC4177 (D). IFN-γ ELISPOT and flow cytometric analysis of 4-1BB expression on CD8+ cells are plotted. (E) 4-1BB expression on the T cell subsets following coculture with the autologous tumor. The percentage of live, CD3+ lymphocytes expressing 4-1BB is shown. (F) IFN-γ ELISPOT and flow cytometric analysis of 4-1BB expression on in vitro–expanded TC4177-reactive CD8+PD-1hi T cells isolated in E following coculture with APCs pulsed with WT and MUT MMP14p.R158C 25-mers or 9-mers, or electroporated with encoding irrelevant TMG RNA or 4177-TMG1 RNA. (G and H) Tumor recognition (intracellular IFN-γ, cell surface CD107a, and 4-1BB) (G) and autologous tumor lysis (H) by MMP14p.R158C-reactive lymphocytes generated through enrichment of 4177-TMG1–reactive lymphocytes shown in C and D (left panel of G) or enrichment of TC4177-reactive lymphocytes shown in F (right panel of G) in the treatment conditions specified. The mean percentage lysis of technical duplicates is plotted. “>500” denotes greater than 500 spots per 2 × 104 cells. Except in H, experiments were performed without duplicates. Data from A–H are representative of at least 2 independent experiments.