PD-1 or CTLA-4 blockade promotes CD86-driven Treg responses upon radiotherapy of lymphocyte-depleted cancer in mice
Elselien Frijlink, Douwe M.T. Bosma, Julia Busselaar, Thomas W. Battaglia, Mo D. Staal, Inge Verbrugge, Jannie Borst
Elselien Frijlink, Douwe M.T. Bosma, Julia Busselaar, Thomas W. Battaglia, Mo D. Staal, Inge Verbrugge, Jannie Borst
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Research Article Immunology Oncology

PD-1 or CTLA-4 blockade promotes CD86-driven Treg responses upon radiotherapy of lymphocyte-depleted cancer in mice

Abstract

Radiotherapy (RT) is considered immunogenic, but clinical data demonstrating RT-induced T cell priming are scarce. Here, we show in a mouse tumor model representative of human lymphocyte–depleted cancer that RT enhanced spontaneous priming of thymus-derived (FOXP3+Helios+) Tregs by the tumor. These Tregs acquired an effector phenotype, populated the tumor, and impeded tumor control by a simultaneous, RT-induced CD8+ cytotoxic T cell (CTL) response. Combination of RT with CTLA-4 or PD-1 blockade, which enables CD28 costimulation, further increased this Treg response and failed to improve tumor control. We discovered that upon RT, the CD28 ligands CD86 and CD80 differentially affected the Treg response. CD86, but not CD80, blockade prevented the effector Treg response, enriched the tumor-draining lymph node migratory conventional DCs that were positive for PD-L1 and CD80 (PD-L1+CD80+), and promoted CTL priming. Blockade of CD86 alone or in combination with PD-1 enhanced intratumoral CTL accumulation, and the combination significantly increased RT-induced tumor regression and OS. We advise that combining RT with PD-1 and/or CTLA-4 blockade may be counterproductive in lymphocyte-depleted cancers, since these interventions drive Treg responses in this context. However, combining RT with CD86 blockade may promote the control of such tumors by enabling a CTL response.

Authors

Elselien Frijlink, Douwe M.T. Bosma, Julia Busselaar, Thomas W. Battaglia, Mo D. Staal, Inge Verbrugge, Jannie Borst

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Figure 3

RT induces concomitant CTL and Treg responses in the TC-1 tumor model.

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RT induces concomitant CTL and Treg responses in the TC-1 tumor model. (...
(A–D) TC-1 tumor–bearing mice were treated with 20 Gy RT (n = 10) or control (0 Gy, n = 4–6) when tumors reached approximately 20 mm2 in size (day 0). FTY720 or vehicle (NaCl) was administered orally on days –1, 3, and 5. On day 8, the CD8+ T cell response was analyzed by flow cytometry in the TdLN (A and B) and tumor (C and D). (A and C) Representative concatenated flow cytometric plots showing IFN-γ+ cells among CD8+ T cells in the TdLN (A) and tumor (C). (B and D) Frequency of CD44+TCF-1–, GZB+, and IFN-γ+ cells among CD8+ T cells in the TdLN (B) and tumor (D). IFN-γ was measured after in vitro PMA/Ionomycin stimulation. (E and F) Monitoring of the (FOXP3+CD25+) Treg response to 20 Gy RT (n = 6–8) or control (0 Gy, n = 6) in TC-1 tumor–bearing mice on day 8 after treatment. (E) Treg frequency among CD4+ T cells in the non-TdLN and TdLN, or among CD45+ cells within the tumor. (F) Percentage of Tregs among live cells in blood at the indicated time points (n = 6/group). (G) Frequency of Tregs in the indicated tissues on day 8 following 20 Gy RT (n = 10) or control (0 Gy, n = 4–6) with or without FTY720 treatment. (H) CD8+ T cell/Treg ratio in the TdLN and tumor after RT. (I) OS of TC-1 tumor–bearing mice treated with 0 Gy (n = 5) or 20 Gy (n = 11–14/group) RT in combination with a CD25-depleting mAb or vehicle (PBS) administered i.p. on day –1 and on day 5 after RT. *P < 0.05 (Mantel-Cox analysis). Data are from 1 experiment and are representative of at least 2 experiments. Error bars indicate the SD. *P < 0.05 and **P < 0.01, by 2-way ANOVA with Bonferroni’s post hoc test (B, D, F and G) and Mann-Whitney U test (E and H).