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Epstein-Barr virus causes vascular abnormalities in epithelial malignancies through upregulating ANXA3-HIF-1α-VEGF pathway

Abstract

Angiogenesis is one of the characteristics of malignant tumors, and persistent generation of abnormal tumor blood vessels is an important factor contributing to tumor treatment resistance. Epstein-Barr virus (EBV) is a highly prevalent DNA oncogenic virus that is associated with the development of various epithelial malignancies. However, the relationship between EBV infection and tumor vascular abnormalities as well as its underlying mechanisms is still unclear. In this study, we found that compared to EBV-uninfected tumors, EBV-infected tumors were more angiogenic, but the neovascularization was mostly immature vessels without pericyte attachment in both clinical patient tumor samples and mouse xenograft models; These immature vessels exhibited aberrant functionality, characterized by poor blood perfusion and increased vascular permeability. The vascular abnormalities caused by EBV infection exacerbated tumor hypoxia and was responsible for accelerated tumor growth. Mechanistically, EBV infection upregulated ANXA3-HIF-1α-VEGF pathway. Silencing the ANXA3 gene or neutralizing ANXA3 with an antibody can diminish vascular abnormalities, thereby increasing immune cell infiltration and alleviating treatment resistance. Finally, a new therapy combining ANXA3 blockade and NK cell + PD1 antibody significantly inhibited the growth of EBV-infected xenografts in mice. In conclusion, our study identified a previously unrecognized role for EBV infection in tumor vascular abnormalities and revealed its underlying mechanism that upregulated the ANXA3-HIF-1α-VEGF pathway. ANXA3 is a potential therapeutic target for EBV-infected tumors and ANXA3 blockade to improve vascular conditions, in combination with NK cell + PD1 antibody therapy, holds promise as an effective treatment strategy for EBV-associated epithelial malignancies.

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Fig. 1: Vascular morphology, number and pericyte attachment in EBV-infected and EBV-uninfected nasopharyngeal carcinoma.
Fig. 2: Effect of EBV infection on nasopharyngeal carcinoma vessels in vitro.
Fig. 3: EBV infection leads to abnormal vascular function and tumor hypoxia in nasopharyngeal carcinoma xenografts.
Fig. 4: Identification of ANXA3 as a key molecule in tumor vascular abnormalities caused by EBV infection.
Fig. 5: ANXA3 upregulates PI3K-HIF-1α-VEGF pathway associated with vascular abnormalities in EBV-infected tumor.
Fig. 6: Knockdown of ANXA3 promotes vascular normalization in EBV-infected tumors.
Fig. 7: AntiANXA3 antibody synergizes with NK cell + PD1 antibody to inhibit the growth of HK1-EBV xenografts.

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Data availability

The raw data of this study would be available from corresponding author TX (email: xiangtong@sysucc.org.cn) upon reasonable request.

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Acknowledgements

We sincerely thank Yulong Han for providing guidance on vascular leakage and perfusion assay methods; we also thank Chaopin Yang for providing technical support on multiplex immunohistochemistry. This work was financially supported by funding from the National Natural Science Foundation of China (No. 82172795, No. 82272790, No. 82273305 and No. 81402560), Guangdong Basic and Applied Basic Research Foundation (No. 2024B1515020001, 2023A1515012981, 2023A1515030013, 2021A1515010443 and 2023A1515012467), the Guangdong Province Science and Technology Plan Project (No. 2017A020215029), and Guangdong Esophageal Cancer Institute Science and Technology Program (No. Q201802).

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Yuanyuan Chen designed the study and performed most of the experiments and wrote the article. Muping Di and Yan Tang collected the pathology specimens for immunohistochemical staining and analyzed the data. Jingjing Zhao and Qijing Wang revised the article. Zhixing Guo performed the experiments to measure blood perfusion in mouse xenografts using Brightness-mode ultrasound. Yongqiang Li, Dijun Ouyang, and Jieying Yang analyzed the Brightness-mode ultrasound results. Hao Chen and Yan Wang participated in the animal experiments. Qiuzhong Pan, Tong Xiang, and Jianchuan Xia participated in the design of the experiments, provided theoretical guidance, and analyzed the results.

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Correspondence to Qiuzhong Pan, Tong Xiang or Jianchuan Xia.

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Chen, Y., Di, M., Tang, Y. et al. Epstein-Barr virus causes vascular abnormalities in epithelial malignancies through upregulating ANXA3-HIF-1α-VEGF pathway. Oncogene (2024). https://doi.org/10.1038/s41388-024-03061-w

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