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Inhibitory functions of PD-L1 and PD-L2 in the regulation of anti-tumor immunity in murine tumor microenvironment

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Abstract

Although a role of PD-L1 in the suppression of anti-tumor immunity and its value as a predictive biomarker has been suggested by various preclinical and clinical studies, the precise mechanisms how PD-L1 and PD-L2, another ligand of PD-1, regulate anti-tumor immunity in the tumor microenvironment are yet to be fully explored. Here, we address this issue using PD-L1-deficient tumor cells, PD-L1-knockout (KO) mice, anti-PD-L1 monoclonal antibody (mAb), and anti-PD-L2 mAb. Firstly, PD-L1-deficient or competent tumor cells were inoculated into wild-type or PD-L1-KO mice. Results of tumor growth and mouse survival indicated that both tumor- and host-derived PD-L1 are functional to suppress anti-tumor immunity, while the former contributes predominantly than the latter. Experiments using bone marrow (BM) chimeric mice, generated by transferring PD-L1-KO BM cells into wild-type mice or vice versa, further suggested that PD-L1 expressed on BM-derived hematopoietic cells mediates the suppressive effects on anti-tumor immunity. Secondly, anti-PD-L2 mAb treatment demonstrated a profound synergy with anti-PD-L1 mAb therapy, whereas anti-PD-L2 mAb alone hardly induced any anti-tumor effects, suggesting that PD-L2’s function becomes evident when the effects of PD-L1 are abrogated by anti-PD-L1 mAb. Consistent with this notion, PD-L2 expression was upregulated on tumor-associated macrophages (TAM) when mice were treated with anti-PD-L1 mAb. Taken together, our study elucidated the importance of PD-L1 associated with tumor cells and non-tumor host cells, particularly BM-derived hematopoietic cells, as well as PD-L2 inducibly expressed on TAM in the suppression of anti-tumor immunity in the tumor microenvironment.

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Abbreviations

ATCC:

American Type Culture Collection

BM:

Bone marrow

i.p.:

Intraperitoneally

KO:

Knockout

mAb:

Monoclonal antibody

mAbs:

Monoclonal antibodies

PD-1:

Programmed cell death-1

PD-L1:

Programmed cell death-ligand 1

PD-L2:

Programmed cell death-ligand 2

s.c.:

Subcutaneously

TAM:

Tumor-associated macrophages

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Acknowledgements

The authors thank Shunsuke Goto, Hiromi Kurosawa and Makiko Miyamoto for excellent technical assistance.

Funding

This study was supported by research funds from Grant-in-Aid for Scientific Research 16H02474 and Ono Pharmaceutical Inc.

Author information

Authors and Affiliations

  1. Department of Immunology, Graduate School of Medicine, Yamaguchi University, 1-1-1 Minami Kogushi, Ube, Yamaguchi, 755-8505, Japan

    Daisuke Umezu, Nana Okada, Yukimi Sakoda, Keishi Adachi & Koji Tamada

  2. Department of Urology, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan

    Daisuke Umezu & Masatoshi Eto

  3. Second Department of Surgery, School of Medicine, Wakayama Medical University, Wakayama, Japan

    Toshiyasu Ojima & Hiroki Yamaue

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  1. Daisuke Umezu
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  2. Nana Okada
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  3. Yukimi Sakoda
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  4. Keishi Adachi
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  5. Toshiyasu Ojima
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  6. Hiroki Yamaue
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  7. Masatoshi Eto
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  8. Koji Tamada
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Contributions

DU, NO, YS, and KA conducted experiments. TO, HY, ME, and KT guided the conduct of experiments. DU and KT wrote the manuscript.

Corresponding author

Correspondence to Koji Tamada.

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Conflict of interest

Koji Tamada received research funds from Ono Pharmaceutical Inc. Other authors declare no conflict of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution at which the studies were conducted (Yamaguchi University, Ube, Japan). Animal research was approved by the Institutional Animal Care and Use Committee of Yamaguchi University (animal research approval number: 14-001).

Animal source

Male or female 6 to 12-week-old wild-type C57BL/6 mice were purchased from Japan SLC (Shizuoka, Japan). PD-L1-KO mice with a C57BL/6 background were kindly provided by Lieping Chen.

Cell line authentication

The MC38 mouse colon carcinoma cell line was kindly provided by F. James Primus. The 3LL mouse lung carcinoma cell line and the B16F10 mouse melanoma cell line were purchased from Japanese Collection of Research Bioresources Cell Bank and American Type Culture Collection (ATCC), respectively, who had authenticated them.

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Umezu, D., Okada, N., Sakoda, Y. et al. Inhibitory functions of PD-L1 and PD-L2 in the regulation of anti-tumor immunity in murine tumor microenvironment. Cancer Immunol Immunother 68, 201–211 (2019). https://doi.org/10.1007/s00262-018-2263-4

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  • DOI: https://doi.org/10.1007/s00262-018-2263-4

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