Tumor-derived neutrophil extracellular trap–associated DNA impairs treatment efficacy in breast cancer via CCDC25-dependent epithelial-mesenchymal transition
Heliang Li, Yetong Zhang, Jianghua Lin, Jiayi Zeng, Xinyan Liang, Linxi Xu, Jiang Li, Xiaoming Zhong, Xu Liu, Zhou Liu, Xinyu Yang, Yunyi Zhang, Shun Wang, Erwei Song, Man Nie, Linbin Yang
Heliang Li, Yetong Zhang, Jianghua Lin, Jiayi Zeng, Xinyan Liang, Linxi Xu, Jiang Li, Xiaoming Zhong, Xu Liu, Zhou Liu, Xinyu Yang, Yunyi Zhang, Shun Wang, Erwei Song, Man Nie, Linbin Yang
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Research Article Immunology Oncology

Tumor-derived neutrophil extracellular trap–associated DNA impairs treatment efficacy in breast cancer via CCDC25-dependent epithelial-mesenchymal transition

Abstract

Neutrophil extracellular traps (NETs) are associated with cancer progression; however, the functional role and clinical importance of NET-DNA in therapeutic resistance remain unclear. Here, we show that chemotherapy and radiotherapy provoke NET-DNA formation in primary tumor and metastatic organs in breast cancer patients and mouse models, and the level of NET-DNA correlates with treatment resistance. Mechanistically, the cathepsin C in tumor debris generated by anticancer therapy is phagocytosed by macrophages and drives CXCL1/2 and complement factor B production via activating the TLR4/NF-κB signaling pathway, subsequently promoting NETosis and impairing therapeutic efficacy. Importantly, we demonstrate that NET-DNA sensor CCDC25 is indispensable in NET-mediated treatment resistance by inducing cancer cell epithelial-mesenchymal transition via pyruvate kinase isoform M2–mediated STAT3 phosphorylation. Clinically, tumoral CCDC25 abundance is closely associated with poor prognosis in patients who underwent chemotherapy. Overall, our data reveal the mechanism of NET formation and elucidate the interaction of NET-CCDC25 in therapy resistance, highlighting CCDC25 as an appealing target for anticancer interventions.

Authors

Heliang Li, Yetong Zhang, Jianghua Lin, Jiayi Zeng, Xinyan Liang, Linxi Xu, Jiang Li, Xiaoming Zhong, Xu Liu, Zhou Liu, Xinyu Yang, Yunyi Zhang, Shun Wang, Erwei Song, Man Nie, Linbin Yang

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Figure 5

The protein CTSC in tumor debris contributes to NETosis after chemotherapy.

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The protein CTSC in tumor debris contributes to NETosis after chemothera...
(A) Schematic showing qRT-PCR analysis of BMDMs stimulated with Dox-induced 4T1 debris with different pretreatments. Created with BioRender.com. (B) Cxcl1/2 and Cfb mRNA expression in BMDMs (n = 3/group, 1-way ANOVA with Tukey’s test). (C) Venn diagram showing the overlapping DEGs upregulated in scRNA-Seq of 4T1 tumors and RNA-Seq (GSE25066). (D) Cxcl1/2 and Cfb mRNA expression in BMDMs treated with or without recombinant CTSC protein (n = 5/group, Mann-Whitney test). (E and F) Tumor growth curves (E), representative NET images, and quantification (F) of 4T1-shNC or 4T1-shCtsc tumors with indicated treatments (n = 5/group). Scale bar, 20 μm. Data were analyzed by 2-way repeated measures ANOVA with Tukey’s test and compared at day 21 (E), or 2-way ANOVA with Tukey’s test (F). (G) Violin plot showing CTSC expression in breast tumors and normal breast tissues from the TNMplot database (normal: n = 242; tumor: n = 7,569; Mann-Whitney test). The lower and upper bounds of boxes indicate the 25th and 75th percentiles, and the middle lines the median values. The whiskers represent the interquartile range. (H) Kaplan-Meier curves for OS in substratified patients with breast cancer receiving chemotherapy with high (n = 617) and low (n = 578) CTSC expression from the KMPLOT database (log-rank test). (I) BMDMs were treated with PBS or recombinant CTSC protein. Immunostaining of p65 and quantification are shown (n = 4/group, Mann-Whitney test). The white arrows indicate the areas that are magnified in the next row. Scale bars, 20 μm and 10 μm. (J) Cxcl1/2 and Cfb mRNA expression in BMDMs with indicated treatments (n = 3/group, 1-way ANOVA with Tukey’s test). (K) Quantification of TLR4/RAB5 colocalization in BMDMs (n = 5/group, 1-way ANOVA with Tukey’s test). Data represent mean ± SD. ****P < 0.0001, ***P < 0.001, **P < 0.01, *P < 0.05.