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CCR8 blockade primes anti-tumor immunity through intratumoral regulatory T cells destabilization in muscle-invasive bladder cancer

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Abstract

Regulatory T cells (Tregs) play a major role in the development of an immunosuppressive tumor microenvironment. Systemic Treg depletion is not favored because of the critical role of Tregs in maintaining immune homeostasis and preventing the autoimmunity. Recently, CCR8 has been identified as an important chemokine receptor expressed on intratumoral Tregs and is known to be critical for CCR8+Treg-mediated immunosuppression. However, the inherent molecular mechanisms and clinical significance of intratumoral CCR8+Tregs remain poorly understood. In this study, a retrospective analysis of 259 muscle-invasive bladder cancer (MIBC) patients from two independent clinic centers was conducted to explore the prognostic merit of CCR8+Tregs via immunohistochemistry. Eighty-three fresh MIBC samples and data from the Cancer Genome Atlas were used to evaluate the proportion and function of immune cells via flow cytometry, ex vivo intervention experiments and bioinformatics analysis. It was found that the CCR8 expression by intratumoral Tregs maintained the stability and potentiated their suppressive function by upregulating the expression of transcript factors FOXO1 and c-MAF. High level of CCR8+Tregs was associated with the immune tolerance and predicted poor survival and inferior therapeutic responsiveness to chemotherapy. Moreover, it was revealed that CCR8 blockade could destabilize intratumoral Tregs into a fragile phenotype accompanied with reactivation of antitumor immunity and augment of anti-PD-1 therapeutic benefits in MIBC. In summary, those results suggested that CCR8+Tregs represented a stable Treg subtype and a promising therapeutic target in the immunotherapy of MIBC.

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Abbreviations

ACT:

Adjuvant chemotherapy

CI:

Confidence interval

FCM:

Flow cytometry

FUSCC:

Fudan University Shanghai Cancer Center

GSEA:

Gene set enrichment analysis

HR:

Hazard ratio

ICIs:

Immune-checkpoint inhibitors

IF:

Immunofluorescence

IHC:

Immunohistochemistry

MIBC:

Muscle-invasive bladder cancer

OS:

Overall survival

PD-1:

Programmed cell death protein 1

RC:

Radical cystectomy

RFS:

Recurrence-free survival

TCGA:

The Cancer Genome Atlas

TMA:

Tissue microarray

TME:

Tumor microenvironment

Treg:

Regulatory T cell

ZH:

Zhongshan Hospital

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Acknowledgements

We thank Dr. Lingli Chen (Department of Pathology, Zhongshan Hospital, Fudan University, Shanghai, China) and Dr. Peipei Zhang (Department of Pathology, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China) for their excellent pathological technology help.

Funding

This study was funded by grants from National Natural Science Foundation of China (81570607, 31770851, 81702496, 81702497, 81702805, 81772696, 81872082, 81902556, 81902563, 81902898, 81974393), three-year action plan for promoting clinical skills and clinical innovation in municipal hospitals of Shanghai Shenkang (16CR2003A), National Natural Science Foundation for Young Scholars of China (81902566), Shanghai Jiaotong University Medical-Engineering Cross Research Fund (YG2019QNA53), National Key R&D Program of China (2017YFC0114303), Shanghai Municipal Natural Science Foundation (16ZR1406500, 17ZR1405100, 19ZR1431800), Guide Project of Science and Technology Commission of Shanghai Municipality (17411963100), Shanghai Sailing Program (18YF1404500, 19YF1407900, 19YF1427200), Shanghai Municipal Commission of Health and Family Planning Program (20174Y0042, 201840168, 20184Y0151), Fudan University Shanghai Cancer Center for Outstanding Youth Scholars Foundation (YJYQ201802) and Shanghai Cancer Research Charity Center. All these study sponsors have no roles in the study design, in the collection, analysis and interpretation of data.

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Author notes

  1. Tao Wang, Quan Zhou, Han Zeng and Hongyu Zhang have contributed equally to this work.

Authors and Affiliations

  1. Department of Urology, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200080, China

    Tao Wang, Jialiang Shao & Xiang Wang

  2. Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China

    Quan Zhou, Han Zeng, Hongyu Zhang, Zhaopei Liu & Jiejie Xu

  3. Department of Urology, Zhongshan Hospital, Fudan University, Shanghai, China

    Zewei Wang, Ying Xiong, Jiajun Wang, Qi Bai, Yu Xia, Li Liu & Jianming Guo

  4. Department of Urology, Shanghai Ninth People’s Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

    Yiwei Wang

  5. Department of Urology, Fudan University Shanghai Cancer Center, Shanghai, 200032, China

    Yu Zhu, Bo Dai & Yuan Chang

  6. Department of Urology, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

    Le Xu

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  1. Tao Wang
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Contributions

TW, QZ, HZ and HZ for acquisition of data, analysis and interpretation of data, statistical analysis and drafting of the manuscript; ZL, JS, ZW, YX, JW, QB, YX, YW, LL, YZ, LX, BD and JG for technical and material support; YC, XW and JX for study concept and design, analysis and interpretation of data, drafting of the manuscript, obtained funding and study supervision. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Yuan Chang, Xiang Wang or Jiejie Xu.

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The authors have declared that they have no conflict of interest.

Ethics approval and consent to participate

Written informed consent was obtained from each patient included, and the protocol of all study cohorts was approved by the Clinical Research Ethics Committee of Zhongshan Hospital and the Ethics Committee of Fudan University Shanghai Cancer Center.

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All data generated that are relevant to the results presented in this article are included in this article. Other data that were not relevant for the results presented here are available from the corresponding author Dr. Xu upon reasonable request.

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Wang, T., Zhou, Q., Zeng, H. et al. CCR8 blockade primes anti-tumor immunity through intratumoral regulatory T cells destabilization in muscle-invasive bladder cancer. Cancer Immunol Immunother 69, 1855–1867 (2020). https://doi.org/10.1007/s00262-020-02583-y

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