PD-1 or CTLA-4 blockade promotes CD86-driven Treg responses upon radiotherapy of lymphocyte-depleted cancer in mice
Elselien Frijlink, … , Inge Verbrugge, Jannie Borst
Elselien Frijlink, … , Inge Verbrugge, Jannie Borst
Published February 13, 2024
Citation Information: J Clin Invest. 2024;134(6):e171154. https://doi.org/10.1172/JCI171154.
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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 2

Myeloid cell– and Treg-rich TC-1 tumor shows a CD8+ T cell–dependent RT response.

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Myeloid cell– and Treg-rich TC-1 tumor shows a CD8+ T cell–dependent RT ...
(A) Frequency of the indicated immune cell populations among CD45+ cells measured by flow cytometry in 50 mm2 TC-1 tumors (n = 6). (B–D) cTregs and eTregs were defined as indicated in Supplemental Figure 2, B–D, and identified in the TdLN, non-TdLN, and tumor of 100 mm2 TC-1 tumor–bearing mice (n = 6) and age-matched naive (non-tumor-bearing) mice (n = 5). FlowSOM-guided clustering was performed on 5,000 randomly selected cells per sample within the CD3+ lymphocyte population. (B and C) Representative histograms depicting expression of the indicated markers on cTreg and eTreg populations in axillary LNs of naive and TC-1 tumor–bearing mice. (D) Frequency of Helios+ cells among cTregs and eTregs in axillary LNs of naive and TC-1 tumor–bearing mice. (E) Percentage of eTregs (left) and cTregs (right) among CD3+ T cells in the indicated tissues. (F–H) Monitoring by flow cytometry of the CD8+ T cell response to 20 Gy RT (n = 8) or control (0 Gy, n = 6) in TC-1 tumors. n-TdLN, non-TdLN. (F) Absolute number of total CD8+ T cells and (G) GZB-, IFN-γ–, or TNF-α–expressing CD8+ T cells per milligram of tumor tissue on after post-RT day 8. IFN-γ and TNF-α levels were measured after in vitro PMA/ionomycin stimulation. (H) OS of TC-1 tumor–bearing mice treated with 20 Gy RT on day 0 in combination with vehicle (PBS, n = 9) or depleting mAbs specific for CD8 (n = 5) or CD4 (n = 9). αCD8, anti-CD8 mAb. ***P < 0.001 (Mantel-Cox analysis). Data are from 1 experiment and are representative of at least 2 experiments. Error bars indicate the SD. *P < 0.05, **P < 0.01, and ***P < 0.001, by Kruskal-Wallis test with uncorrected Dunn’s post hoc analysis (E) and Mann-Whitney U test (D, F, and G).