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Enhancer of zeste homolog 2-mediated paired box 8 methylation promotes gastrointestinal stromal tumor progression through Wnt4 downregulation

Cancer Gene Therapy volume 28, pages 1162–1174 (2021)Cite this article

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Abstract

Gastrointestinal stromal tumor (GIST) is a refractory malignant tumor without satisfactory therapy. In recent years, aberrant gene methylation has been highlighted as an inducer for tumor progression. In this study, we explored whether enhancer of zeste homolog 2 (EZH2)-mediated paired box 8 (PAX8) methylation affects GIST development through regulation of Wnt4. A total of 50 cases of GIST tissues were collected and the human GIST cell lines were cultured. PAX8 methylation was examined using MS-PCR. Following loss- and gain-function approaches, GIST cell proliferation, migration, invasion, and apoptosis were examined by CCK-8 assay, Transwell assay and flow cytometry. The expression of proliferation related factors and apoptosis related factors was determined. Finally, xenograft tumors in nude mice were observed to examine in vivo tumorigenicity of GIST cells. Downregulated PAX8 and upregulated EZH2 expression was found in GIST tissues. Overexpression of PAX8 or suppression of PAX8 methylation using DNA methyltransferase inhibitor 5-Aza-dC inhibited the proliferation, migration, and invasion of GIST cells while promoting their apoptosis (diminished PCNA, Ki67 and Bcl-2, elevated Bax, and cleaved caspase-3). EZH2 promoted PAX8 methylation to inhibit its expression. Downregulated PAX8 decreased Wnt4 expression to accelerate GIST progression both in vitro and in vivo. Collectively, EZH2 inhibits PAX8 expression by promoting its methylation, which thus downregulates Wnt4 expression, thereby promoting the development of GIST.

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Fig. 1: PAX8 is downregulated in GIST.
Fig. 2: Overexpression of PAX8 affects the biological characteristics of GIST882 cells.
Fig. 3: EZH2 promotes the methylation of PAX8.
Fig. 4: Inhibition of PAX8 methylation suppresses the progression of GIST.
Fig. 5: EZH2 participates in the progression of GIST by regulating the methylation of PAX8 to mediate Wnt4 expression.
Fig. 6: EZH2 regulates in vivo tumorigenicity of GIST by mediating the expression of PAX8/Wnt4.

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The authors confirm that the data supporting the findings of this study are available within the article.

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Acknowledgements

We would like to acknowledge the colleagues for their helpful assistance on this study.

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Authors and Affiliations

  1. Department of Abdominal Surgery, Jiangxi Tumor Hospital, 330029, Nanchang, P.R. China

    Zhiguo Miao, Haiyun Wei, Zhipeng Luo & Kun Wu

  2. Department of Pathology, Jiangxi Tumor Hospital, 330029, Nanchang, P.R. China

    Fanggeng Wu

  3. Department of General Surgery, The First Affiliated Hospital of Nanchang University, 330006, Nanchang, P.R. China

    Jiangnan Zhang

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Contributions

ZM and FW designed study and collated data. HW and ZL carried out data analyses and produced initial draft of manuscript. KW and JZ contributed to drafting and polishing manuscript. All authors read and approved final manuscript.

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Correspondence to Jiangnan Zhang.

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The authors declare that they have no conflict of interest.

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This study was approved and reviewed by the medical ethics committee of the First Affiliated Hospital of Nanchang University and carried out following the Declaration of Helsinki, with signed informed consent from each study subject. The animal experimental procedures were also approved by the animal ethics committee of the First Affiliated Hospital of Nanchang University.

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Miao, Z., Wu, F., Wei, H. et al. Enhancer of zeste homolog 2-mediated paired box 8 methylation promotes gastrointestinal stromal tumor progression through Wnt4 downregulation. Cancer Gene Ther 28, 1162–1174 (2021). https://doi.org/10.1038/s41417-020-00266-5

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