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Interferon regulatory factor 1 (IRF-1) and IRF-2 regulate PD-L1 expression in hepatocellular carcinoma (HCC) cells

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Abstract

The objective response rate of immune checkpoint blockade (ICB) in hepatocellular carcinoma (HCC) with anti PD-L1/PD-1 therapy is low. Discovering the signaling pathways regulating PD-L1 might help to improve ICB response rates. Here, we investigate transcription factors IRF-1 and IRF-2 signaling pathways regulating PD-L1 in HCC cells. In vivo studies show that IRF-1 and PD-L1 mRNA expression in human HCC tumors are significantly repressed compared with noncancerous background liver. IRF-1, IRF-2, and PD-L1 mRNA expression correlated positively in HCC tumors. Increased IRF-1 mRNA expression was observed in patients with well-differentiated or early stage HCC tumors. In vitro studies show that IFN-γ induces PD-L1 mRNA and protein expression through upregulation of IRF-1 in mouse and human HCC cells. IRF-1, IRF-2, and PD-L1 mRNA expression is upregulated in murine HCC by co-culture with effector T cells from spleen cells incubated with anti-CD3/CD28 antibodies. IRF-2 over-expression down-regulates IFN-γ induced PD-L1 promoter activity and protein levels in a dose-dependent manner. We identify two IRF-1 response elements (IRE1/IRE2) in the upstream 5′-flanking region of the CD274 (PD-L1) gene promoter. Site-directed mutagenesis shows both IRE1 and IRE2 are functional in transfection promoter assays. IRF-1 traditionally functions as tumor suppressor gene. However, these novel findings show a complex role for IRF-1 which upregulates PD-L1 in the inflammatory tumor microenvironment. IRF-1 antagonizes IRF-2 for binding to the IRE promoter element in PD-L1 which gives new insight to the regulation of PD-L1/PD-1 pathways in HCC ICB therapy.

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Abbreviations

ATR:

Ataxia telangiectasia and Rad3-related protein

cGAS:

Cyclic guanosine monophosphate (GMP)–adenosine monophosphate (AMP) synthase

CHEK-1:

Check point kinase 1

CTLA-4:

Cytotoxic T-lymphocyte-associated protein 4

HCC:

Hepatocellular carcinoma

ICB:

Immune checkpoint blockade

IFN-γ:

Interferon-γ

IRE:

IRF response element

IRF-1:

Interferon regulatory factor 1

IRF-2:

Interferon regulatory factor 2

JAK:

Janus kinase

MARK:

Mitogen-activated protein kinase

PD-L1:

Programmed death-ligand 1

PD-1:

Programmed death protein 1

PI3K:

Phosphatidylinositol 3-kinase

RLU:

Relative luciferase unit

STAT:

Signal transducer and activator of transcription

STING:

Stimulator of interferon gene

TCGA:

The Cancer Genome Atlas

TNM:

Tumor node metastasis classification

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Funding

DAG was supported by funding from National Institute of Health (HHSN276201200017C and P30DK120531-01). YY was supported by funding from Guangxi Natural Science Foundation (2017GXNSFAA198014) and Guangxi High Education Institute Science Foundation (2017KY0099).

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Authors and Affiliations

  1. Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh Medical Center, Pittsburgh, PA, 15260, USA

    Yihe Yan, Leting Zheng, Qiang Du, Bing Yan & David A. Geller

  2. Department of General Surgery, The Second Affiliated Hospital of Guangxi Medical University, Nanning, 530007, Guangxi, China

    Yihe Yan

  3. Department of Rheumatology and Immunology, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, Guangxi, China

    Leting Zheng

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  1. Yihe Yan
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  2. Leting Zheng
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  4. Bing Yan
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Contributions

YY and DAG designed the research. YY, LZ, QD, BY, and DAG performed research and also analyzed the data. YY and DAG wrote the paper. All authors edited and approved the submission of this work.

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Correspondence to Yihe Yan or David A. Geller.

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All individuals provided written informed consent and human tissues were obtained in accordance with the University of Pittsburgh Institutional Review Board (IRB) approved protocol (No. MOD08010372/PRO08010372).

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Yan, Y., Zheng, L., Du, Q. et al. Interferon regulatory factor 1 (IRF-1) and IRF-2 regulate PD-L1 expression in hepatocellular carcinoma (HCC) cells. Cancer Immunol Immunother 69, 1891–1903 (2020). https://doi.org/10.1007/s00262-020-02586-9

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