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METTL3 inhibition induced by M2 macrophage-derived extracellular vesicles drives anti-PD-1 therapy resistance via M6A-CD70-mediated immune suppression in thyroid cancer

Cell Death & Differentiation volume 30, pages 2265–2279 (2023)Cite this article

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Abstract

The treatment options for advanced papillary thyroid cancer (PTC) and anaplastic thyroid cancer (ATC) refractory to standard therapies are limited. Although anti-PD-1 therapy has a manageable safety profile and has been effective in a small percentage of patients with advanced PTC and refractory ATC, the majority of the patients either do not respond or develop resistance to anti-PD-1 therapy. N6-methyladenosine (m6A) modification is a critical determinant of the complexity of the tumor microenvironment (TME). However, it is unclear whether and how m6A modification in tumor cells shapes the immune landscape of PTC and ATC. In this study, we performed bulk and single cell RNA sequencing analysis of PTC and ATC tissues, and found that low METTL3 expression not only correlated to poor response to immune checkpoint blockade (ICB) but was also associated with increased TNF family-related ligand-receptor interactions in the immunosuppressive Tregs and exhausted T cells. Furthermore, overexpression of METTL3 in PTC and ATC cells enhanced the efficacy of anti-PD-1 therapy in a peripheral blood mononuclear cell humanized NCG (huPBMC-NCG) mouse model. Mechanistically, M2 macrophage-derived extracellular vesicles (M2 EVs) inhibited METTL3 expression in PTC and ATC cells via miR-21-5p. Downregulation of METTL3 promoted demethylation of CD70 mRNA, which prevented YTHDF2-mediated degradation of the transcripts. The stabilization of CD70 mRNA, and the subsequent upregulation in CD70 protein levels increased the abundance of the immunosuppressive Tregs and terminally exhausted T cells, thereby inducing resistance to anti-PD-1 therapy. Furthermore, blocking CD70 using cusatuzumab, a high-affinity monoclonal antibody, reversed the anti-PD-1 therapy resistance induced by M2 EVs in vivo. Finally, we demonstrated that METTL3 expression negatively correlated with CD70 expression and M2 macrophages and Tregs infiltration in PTC and ATC tissues. Our findings provide new insights into developing novel therapies for advanced PTC and ATC.

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Fig. 1: Low METTL3 expression in PTC and ATC leads to an immunosuppressive TME and resistance to ICB therapy.
Fig. 2: METTL3 overexpression in the PTC and ATC cells enhanced the efficacy of anti-PD-1 therapy.
Fig. 3: METTL3 silencing in thyroid cancer cells promoted an immunosuppressive phenotype via CD70 demethylation.
Fig. 4: METTL3-mediated regulation of CD70 is dependent on YTHDF2.
Fig. 5: M2 macrophage-derived EVs downregulated METTL3 in PTC and ATC cells through miR-21-5p.
Fig. 6: CD70 blockade reversed αPD-1 resistance induced by M2 EVs in PTC and ATC.
Fig. 7: METTL3 expression negatively correlates with CD70 expression and M2 macrophages and Tregs infiltration in PTC and ATC tissues.
Fig. 8: The proposed model illustrates the mechanism by which M2 EVs induced METTL3 inhibition to drives anti-PD-1 therapy resistance via m6A-CD70-mediated immune suppression in thyroid cancer.

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Data availability

The data supporting the present study are available from the corresponding author upon reasonable request.

Code availability

Detailed code is available from the corresponding author upon request.

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Funding

Funding

This work was supported by grants from the National Natural Science Foundation of China (82203668, 82172821, 82103386, 81872169), Tianjin Municipal Science and Technology Project (19JCYBJC27400, 21JCZDJC00360), Beijing-Tianjin-Hebei Basic Research Cooperation Project (20JCZXJC00120), the Science &Technology Development Fund of Tianjin Education Commission for Higher Education (2021ZD033), Tianjin Medical Key Discipline (Specialty) Construction Project (TJYXZDXK-058B), Tianjin Health Research Project (TJWJ2022XK024).

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  1. These authors contributed equally: Junya Ning, Xiukun Hou, Jie Hao.

Authors and Affiliations

  1. Department of Thyroid and Breast Surgery, Tianjin Key Laboratory of General Surgery in Construction, Tianjin Union Medical Center, Tianjin, 300121, China

    Junya Ning, Jie Hao, Wei Zhang & Ming Gao

  2. Department of Thyroid and Neck Cancer, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Tianjin’s Clinical Research Center for Cancer, Tianjin, 300060, China

    Junya Ning, Xiukun Hou, Jie Hao, Wei Zhang, Yue Huang, Xianhui Ruan, Xiangqian Zheng & Ming Gao

  3. The School of Medicine, Nankai University, Tianjin, 300071, China

    Wei Zhang & Yi Shi

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Contributions

MG, XZ and XR performed study concept, design and supervision; JN, XH and JH designed and performed most of the experiments, analyzed the data, and created the figures. WZ and YH participated in some experiments. JN and XH wrote and revised the manuscript. YS helped to revise the manuscript. MG, XZ, XR, and NJ provided financial support. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Xianhui Ruan, Xiangqian Zheng or Ming Gao.

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The authors declare no competing interests.

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This study was approved by the Ethics Committee of Tianjin Medical University Cancer Institute and Hospital (No. EK2021149 for human cancer specimens and NO. AE-2022032 for in vivo mouse experiments). Written informed consent was obtained from all subjects. All experiments were performed in accordance with the principles of the Declaration of Helsinki.

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Ning, J., Hou, X., Hao, J. et al. METTL3 inhibition induced by M2 macrophage-derived extracellular vesicles drives anti-PD-1 therapy resistance via M6A-CD70-mediated immune suppression in thyroid cancer. Cell Death Differ 30, 2265–2279 (2023). https://doi.org/10.1038/s41418-023-01217-x

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