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RTP4 silencing provokes tumor-intrinsic resistance to immune checkpoint blockade in colorectal cancer

  • Original Article—Alimentary Tract
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Abstract

Background

Recent advances in immune checkpoint blockade (ICB) have improved patient prognosis in mismatch repair-deficient and microsatellite instability-high colorectal cancer (dMMR/MSI-H CRC); however, PD-1 blockade has faced a challenge in early progressive disease. We aimed to understand the early event in ICB resistance using an in vivo model.

Methods

We subcutaneously transplanted the MC38 colon cancer cells into C57BL/6 mice, intraperitoneally injected anti-PD-1 antibody and then isolated ICB-resistant subclones from the recurrent tumors.

Results

Comparative gene expression analysis discovered seven genes significantly downregulated in the ICB-resistant cells. Tumorigenicity assay of the MC38 cells knocked out each of the seven candidate genes into C57BL/6 mice treated with anti-PD-1 antibody and bioinformatics analysis of the relationship between the expression of the seven candidate genes and the outcome of cancer patients receiving immunotherapy identified Rtp4, an interferon-stimulated gene and a chaperon protein of G protein-coupled receptors, as a gene involved in ICB resistance. Immunohistochemical analysis of transplanted tumor tissues demonstrated that anti-PD-1 antibody failed to recruit T lymphocytes in the Rtp4-KO MC38 cells. Mouse and human RTP4 expression could be silenced via histone H3 lysine 9 (H3K9) trimethylation, and public transcriptome data indicated the high expression level of RTP4 in most but not all of dMMR/MSI-H CRC.

Conclusions

We clarified that RTP4 could be silenced by histone H3K9 methylation as the early event of ICB resistance. RTP4 expression could be a promising biomarker for predicting ICB response, and the combination of epigenetic drugs and immune checkpoint inhibitors might exhibit synergistic effects on dMMR/MSI-H CRC.

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Abbreviations

BSS:

Bisulfite sequencing

CCLE:

Cancer cell line encyclopedia

ChIP:

Chromatin immunoprecipitation

CMS:

Consensus molecular subtype

CRC:

Colorectal cancer

dMMR:

Mismatch repair-deficient

FBS:

Fetal bovine serum

GDC:

National Cancer Institute Genomic Data Commons

H3K9:

Histone H3 lysine 9

HDAC:

Histone deacetylase

ICB:

Immune checkpoint blockade

IFN:

Interferon

JAK:

Janus kinase

KO:

Knockout

MHC-I:

Major histocompatibility complex class 1

MSI-H:

Microsatellite instability-high

MSS:

Microsatellite stable

OS:

Overall survival

PFS:

Progression-free survival

pMMR:

Mismatch repair-proficient

TCGA:

Cancer Genome Atlas Research Network

TIDE:

Tumor Immune Dysfunction and Exclusion

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Acknowledgements

We thank Ms. Hiromi Nagasaki for technical assistance. The plasmid lentiCRISPR v2 (#52961; Addgene) was a gift from Dr. Feng Zhang. The plasmids psPAX2 (#12260; Addgene) and pMD2.G (#12259; Addgene) were generously provided from Dr. Didier Trono. This work was supported by Grants-in-Aid for Scientific Research (A; 19H01055) and Challenging Research (Exploratory; 22K19554) from the Ministry of Education, Culture, Sports, Science and Technology of Japan; Research Grant from the Princess Takamatsu Cancer Research Fund; P-CREATE (JP19cm0106540) from AMED (Japan Agency for Medical Research and Development).

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Authors and Affiliations

  1. Department of Molecular Oncology, Graduate School of Medicine, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-Ku, Tokyo, 113-8519, Japan

    Yudai Yamamoto, Shu Shimada, Yoshimitsu Akiyama, Shu Tsukihara, Raizo Sugimoto, Ayano Kabashima & Shinji Tanaka

  2. Department of Gastrointestinal Surgery, Graduate School of Medicine, Tokyo Medical and Dental University, Tokyo, Japan

    Yudai Yamamoto, Masanori Tokunaga & Yusuke Kinugasa

  3. Department of Surgery, The Jikei University School of Medicine, Tokyo, Japan

    Shu Tsukihara

  4. Institute for Advanced Medical Research, Division of Cellular Signaling, Keio University School of Medicine, Tokyo, Japan

    Yutaka Kawakami

  5. Department of Immunology, School of Medicine, International University of Health and Welfare, Chiba, Japan

    Yutaka Kawakami

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  1. Yudai Yamamoto
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Contributions

YY, SS, YA, ST (Tsukihara) and RS performed experiments and analyzed data. YY and SS wrote the manuscript. YA, AK, MT, YK (Kinugasa), YK (Kawakami) and ST (Tanaka) revised the manuscript. SS and ST (Tanaka) designed the study. SS performed bioinformatics analysis and visualized data. YK (Kawakami) provided suggestions for immunological assays. ST (Tanaka) supervised the study.

Corresponding authors

Correspondence to Shu Shimada or Shinji Tanaka.

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Yamamoto, Y., Shimada, S., Akiyama, Y. et al. RTP4 silencing provokes tumor-intrinsic resistance to immune checkpoint blockade in colorectal cancer. J Gastroenterol 58, 540–553 (2023). https://doi.org/10.1007/s00535-023-01969-w

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  • DOI: https://doi.org/10.1007/s00535-023-01969-w

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