对于难治的晚期乳头状甲状腺癌（PTC）和间变甲状腺癌（ATC）来说，标准疗法的治疗选择有限。虽然抗PD-1治疗具有可控的安全性，并在少数晚期PTC和难治ATC患者中有效，但大部分患者要么不响应，要么对抗PD-1治疗产生抵抗。N6-甲基腺苷（m6A）修饰是肿瘤微环境（TME）复杂度的关键决定因素，但目前尚不清楚肿瘤细胞中m6A修饰是否以及如何塑造PTC和ATC的免疫景观。本研究通过对PTC和ATC组织进行批量和单细胞RNA测序分析，发现低METTL3表达不仅与免疫检查点阻断（ICB）治疗的差反应相关，而且与免疫抑制性调节T细胞和终末耗竭T细胞中增加的TNF家族相关配体-受体相互作用有关。此外，在PTC和ATC细胞中过表达METTL3可以增强抗PD-1治疗在外周血单个核细胞人源化NCG（huPBMC-NCG）小鼠模型中的疗效。机制上，M2巨噬细胞衍生的细胞外囊泡（M2 EVs）通过miR-21-5p抑制PTC和ATC细胞中的METTL3表达。METTL3的下调促进了CD70 mRNA的去甲基化，阻止了YTHDF2介导的转录物降解。CD70 mRNA的稳定和随后CD70蛋白水平的上调增加了免疫抑制性调节T细胞和终末耗竭T细胞的丰度，从而导致对抗PD-1治疗的抵抗。此外，使用高亲和力的单克隆抗体cusatuzumab阻断CD70能够逆转由M2 EVs诱导的抗PD-1治疗耐药性。最后，我们证明METTL3表达与PTC和ATC组织中的CD70表达以及M2巨噬细胞和调节T细胞浸润呈负相关。我们的研究结果为开发晚期PTC和ATC的新治疗方法提供了新的见解。 © 2023. 作者（们），在ADMC Associazione Differenziamento e Morte Cellulare的专属许可下。

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.© 2023. The Author(s), under exclusive licence to ADMC Associazione Differenziamento e Morte Cellulare.