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CD4−/CD8− double-negative tumor-infiltrating lymphocytes expanded from solid tumor tissue suppress the proliferation of tumor cells in an MHC-independent way

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Journal of Cancer Research and Clinical Oncology Aims and scope Submit manuscript

Abstract

Purpose

Tumor-infiltrating lymphocytes (TILs) have shown remarkable clinical responses in some patients with advanced solid tumors. As a rare subset of TILs, CD4−/CD8− double-negative T cells (DNTs) were poorly known. This study aims to investigate the characteristics and function of CD3+CD4−CD8− TILs (double-negative TIL, DN-TILs) derived from solid tumor.

Methods

DN-TILs were derived and expanded ex vivo from resected gastric carcinoma tissue and phenotyped by flow cytometry. The cytotoxicity of DN-TILs was determined against established tumor cell lines in vitro or through in vivo adoptive transfer into xenograft models. K562 cells were transferred with the HLA gene to verify whether the cytotoxicity of DN-TILs was MHC-independent.

Results

Flow cytometric analysis revealed a high-purity population of DN-TILs (> 97%) within CD3+ TILs, which expanded more than 800-folds in 2 weeks, consisting of a mixture of alpha–beta (αβ) and gamma–delta (γδ) T-cell receptor (TCR)—expressing cells (with the majority being αβ-TCR, > 95%). Using single-cell RNA sequencing, the expanded DN-TILs were categorized into four main subsets, Natural Killer T cells (approximately 80%, 5563 in 7028), Progenitor cells, Germ cells and T helper2 cells. DN-TILs exhibited a broad anticancer cytotoxicity in a donor-unrestricted manner against various cancer cell lines derived from pancreatic cancer (Panc-1), gastric cancer (HGC-27), ovarian cancer (SKOV-3), malignant melanoma (A375). The cytotoxicity was MHC-independent, which was not altered in K562 transferring with HLA gene or not. DN-TILs significantly reduced tumor volume in xenograft models with superior tumor-homing ability and low off-target toxicity.

Conclusion

Gastric carcinoma derived DN-TIL can target tumor cells in vitro and in vivo. DN-TILs have the potential to be used as a adoptive cell therapy for solid cancers with both the advantages of DNT and TIL.

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Data Availability

The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

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Funding

Clinical research project of Shanghai Municipal Health Commission (No. 201940492).

Author information

Author notes

  1. Jingyi Lu and Chen Huang are equal first authors of this publication.

Authors and Affiliations

  1. Departmalet of Medical Oncology, Shanghai Tenth People’s Hospital, Tongji University Cancer Center, School of Medicine, Tongji University, Shanghai, 200072, China

    Jingyi Lu, Rong He, Rongjia Xie, Xianling Guo & Qing Xu

  2. Tongji University Cancer Center, Shanghai, China

    Jingyi Lu, Rong He, Rongjia Xie, Xianling Guo & Qing Xu

  3. Shanghai Juncell Biotechnology Co., LTD, Shanghai, China

    Chen Huang

  4. Shanghai Tumor Cell Therapy Technology Innovation Center, Shanghai, China

    Yue Li

  5. Department of Biotherapy, Eastern Hepatobiliary Surgery Hospital, Shanghai, China

    Qian Zhang

Authors
  1. Jingyi Lu
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  3. Rong He
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  4. Rongjia Xie
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  7. Qian Zhang
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  8. Qing Xu
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Contributions

All authors contributed to the study conception or design. Clinical samples were provided by XG and QZ. Material preparation was performed by RX. Cell assays were performed by JL, CH and RH. Data collection and analysis were performed by JL, CH, YL and QX. JL, CH and QX contributed to the study conception and design. The first draft of the manuscript was written by JL and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript. JL and CH are equal first authors of this publication.

Corresponding author

Correspondence to Qing Xu.

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Conflict of interest

Chen Huang is an employee of Shanghai Juncell Biotechnology Co., LTD. The other authors declare that they have no conflict of interest.

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Lu, J., Huang, C., He, R. et al. CD4−/CD8− double-negative tumor-infiltrating lymphocytes expanded from solid tumor tissue suppress the proliferation of tumor cells in an MHC-independent way. J Cancer Res Clin Oncol 149, 9007–9016 (2023). https://doi.org/10.1007/s00432-023-04823-x

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  • DOI: https://doi.org/10.1007/s00432-023-04823-x

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