Proliferative exhausted CD8+ T cells exacerbate long-lasting anti-tumor effects in human papillomavirus-positive head and neck squamous cell carcinoma

  1. Danni Cheng
  2. Ke Qiu
  3. Yufang Rao
  4. Minzi Mao
  5. Li Li
  6. Yan Wang
  7. Yao Song
  8. Junren Chen
  9. Xiaowei Yi
  10. Xiuli Shao
  11. Shao Hui Huang
  12. Yi Zhang
  13. Xuemei Chen
  14. Sisi Wu
  15. Shuaishuai Yu
  16. Jun Liu
  17. Haiyang Wang
  18. Xingchen Peng
  19. Daibo Li
  20. Lin Yang
  21. Li Chen
  22. Zhiye Ying
  23. Yongbo Zheng
  24. Meijun Zheng
  25. Binwu Ying
  26. Xiaoxi Zeng
  27. Wei Zhang
  28. Wei Xu
  29. Geoffrey Liu
  30. Fei Chen
  31. Haopeng Yu
  32. Yu Zhao
  33. Jianjun Ren

  • Curated by eLife

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    eLife assessment

    This study provides fundamental insight into the functional impact of CDK4 inhibition on cells in the tumor microenvironment, which is of high importance and interest to the field. The compelling conclusion that proliferative exhausted T cells are associated with response in HPV+ head and neck cancer is supported by the cohort of 14 patients with paired tumor and adjacent normal tissue and rigorous bioinformatic analysis of nearly 50,000 single CD3+ T cell transcriptomes. This work will be of interest to researchers across tumor types and in other immunological fields of study.

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Abstract

The survival prognosis of human papillomavirus (HPV)-positive and HPV-negative head and neck squamous cell carcinoma (HNSCC) is largely different, and little is known about the anti-tumor mechanism of tumor-infiltrated exhausted CD8 + T cells (Tex) in HNSCC. We performed cell-level multi-omics sequencing on human HNSCC samples to decipher the multi-dimensional characteristics of Tex cells. A proliferative exhausted CD8 + T cell cluster (P-Tex) which was beneficial to survival outcomes of patients with HPV-positive HNSCC was identified. Interestingly, P-Tex cells expressed CDK4 genes as high as cancer cells, which could be simultaneously inhibited by CDK4 inhibitors and might be a potential reason for the ineffectiveness of CDK4 inhibitors in treating HPV-positive HNSCC. P-Tex cells could aggregate in the antigen-presenting cell niches and activate certain signaling pathways. Together, our findings suggest a promising role for P-Tex cells in the prognosis of patients with HPV-positive HNSCC by providing modest but persistent anti-tumor effects.

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  1. Version published to 10.7554/elife.82705 on eLife
  2. eLife

    Author Response

    Joint Public review:

    1. Line 215: The authors state that pairing TCRseq with RNAseq reflects the magnitude of TCR signaling. This is absolutely not the case. TCR sequencing does not reflect TCR signaling strength.

    Thanks for the comments and we apologize for the usage of this misleading description. Actually in this part, we were trying to quantitatively assess the activation states of CD8 T cells based on the average expression of previously described activation-related gene signatures1 (also shown in Supplementary file 3). Therefore, TCRseq data was not involved in this analysis and the magnitude of TCR signaling could neither be reflected. We apologize again for this mistake and have corrected the corresponding texts and figures as follows (line 210-217): "Meanwhile, the activation states of CD8 T cell subpopulations were quantitatively assessed based on the average expression of previously described activation-related gene signatures1 (also shown in Supplementary file 3). Our results showed that the T-Tex cluster was the most activated, followed by the two P-Tex clusters (Fig. 2b left). In addition, CD8 T cells in tumor tissues were more activated than those in adjacent normal tissues (Fig. 2b, right top). And no significant difference in T cell activation states was observed between HPV-positive and HPV-negative samples (Fig. 2b right bottom)."

    1. A lot of discussion around "activation" is presented, but there is no evidence to support which genes or gene programs are associated with "activation".

    Thanks for the comments. The activation states of CD8 T cell subpopulations were quantitatively assessed based on the average expression of previously described activation-related gene signatures1 (also shown in Supplementary file 3). More specifically, activation-related gene signatures are as follows: "CD69, CCR7, CD27, BTLA, CD40LG, IL2RA, CD3E, CD47, EOMES, GNLY, GZMA, GZMB, PRF1, IFNG, CD8A, CD8B, CD95L, LAMP1, LAG3, CTLA4, HLA-DRA, TNFRSF4, ICOS, TNFRSF9, TNFRSF18".

    1. Line 249: It is unclear why the authors are indicating that TCRseq was used in pseudotime analysis. This type of analysis does not take TCRs into account but rather looks at the proportion of spliced mRNA of individual genes from the DGE data.

    Thanks for the comments and we apologize for the usage of this misleading description. As acknowledged by the reviewer, pseudotime analysis has nothing to do with TCRseq data. Actually in this part, we separately performed clonality analysis of CD8 T cells based on TCRseq data and pseudotime analysis based on RNAseq data. Shared TCRs were identified among certain cell subclusters, which could partially validate the potential lineage relationships simulated by pseudotime analysis. Therefore, we have corrected the texts as follows to avoid the misunderstanding that TCRseq was used in pseudotime analysis: "Given the clonal accumulation of CD8 T cells was a result of local T cell proliferation and activation in the tumor environment2, we further conducted clonality analysis of CD8 T cells based on TCRseq data. " (line 246-248) and "To further investigate their lineage relationships, we performed pseudotime analysis for CD3+ T cells on the basis of transcriptional similarities (Fig. 3j-l, Figure 3-figure supplementary 2d)." (line 277-279).

  3. eLife

    eLife assessment

    This study provides fundamental insight into the functional impact of CDK4 inhibition on cells in the tumor microenvironment, which is of high importance and interest to the field. The compelling conclusion that proliferative exhausted T cells are associated with response in HPV+ head and neck cancer is supported by the cohort of 14 patients with paired tumor and adjacent normal tissue and rigorous bioinformatic analysis of nearly 50,000 single CD3+ T cell transcriptomes. This work will be of interest to researchers across tumor types and in other immunological fields of study.

  4. eLife

    Joint Public Review:

    In this study, the authors transcriptomically characterize TIL from head and neck cancers and associate their transcriptional programs with overall survival as a function of HPV positivity. Specifically, they study the impact of CDK4 inhibition on TIL from these tumors. They find an exhausted T cell subset that preferentially expresses CDK4. They then perform some in vitro studies to test the function of exhausted T cells and the impact of CDK4 inhibition on different TIL subsets from head and neck tumors. Understanding the functional impact of different cancer therapies on cells in the TME is of high importance and interest to the field.

    1. Line 215: The authors state that pairing TCRseq with RNAseq reflects the magnitude of TCR signaling. This is absolutely not the case. TCR sequencing does not reflect TCR signaling strength.
    2. A lot of discussion around "activation" is presented, but there is no evidence to support which genes or gene programs are associated with "activation".
    3. Line 249: It is unclear why the authors are indicating that TCRseq was used in pseudotime analysis. This type of analysis does not take TCRs into account but rather looks at the proportion of spliced mRNA of individual genes from the DGE data.
    4. There is no way to know if the differences in proliferation and cell viability shown in Figs. 4a and b, respectively, are meaningful or not. Proper controls or replicates should be provided to fully understand if this difference is biologically meaningful. Likewise, what is the evidence that P-Tex cells are self-renewing rather than expanding?

  5. Version published to 10.1101/2022.10.27.513993v1 on bioRxiv