低氧诱导髓母细胞瘤DNA羟甲基化异常的分子机制研究

Medulloblastoma is the most common malignant tumor in central neural system. It is well identified that medulloblastomas have dysregulated DNA methylation which lead to abnormal gene expression. Recent studies demonstrate that ten-eleven translocation (Tet) hydroxylases can converted 5mC into 5-hydroxymethylcytosine (5hmC). This process of active DNA demethylation is crucial for the balance the DNA methylation. However, only few reports have focused on the roles of 5hmC in medulloblastoma. In our previous works, global 5hmC levels were decreased in medulloblastoma compared with normal cerebellum. Moreover, loss of 5hmC was associated with the low expression of TET1/2 and high cell proliferation in medulloblastoma. In addition, we found abnormal vessels and HIF activation in medulloblastoma. Based on these findings, we hypothesized that hypoxia may induce the aberrations of DNA hydroxymethylation in medulloblastoma. In this project, we will try to evaluate the mechanisms that hypoxia induced aberrations of DNA hydroxymethylation and to identify the interaction between HIF and TET in medulloblastomas. We also try to find whether HIF inhibitors or anti-angiogenesis treatment could improve the hypoxic microenvironment and rescue the aberrations of DNA hydroxymethylation in medulloblastoma. This study will not only elucidate the regulatory mechanisms of DNA hydroxymethylation in medulloblastoma, but also provide new insights into development of new therapies targeting on angiogenesis and epigenetic modifications for medulloblastoma.

髓母细胞瘤作为儿童中枢神经系统最常见的恶性肿瘤，存在明显的DNA甲基化异常。TET家族DNA羟化酶介导的DNA羟甲基化作为主动去甲基化的关键环节，对DNA甲基化的平衡起到关键调控作用。我们前期工作发现，髓母细胞瘤中5-羟甲基胞嘧啶（5hmC）整体水平降低，并与TET1/2异常表达以及细胞增殖密切相关；同时，髓母细胞瘤也存在明显的低氧状态，提示可能与羟甲基化异常存在密切联系。在此基础上，我们将通过免疫荧光、斑点印迹法和DNA羟甲基化测序等方法，在髓母细胞瘤中进一步研究：（1）低氧诱导DNA羟甲基化异常的主要作用机制；（2）分析HIF与Tet的相互作用以及对DNA羟甲基化水平的影响；（3）采用低氧相关通路抑制剂和/或抗血管生成药物改善肿瘤低氧微环境，进一步明确能否逆转DNA羟甲基化异常。通过本研究的实施，将为拓展以表观修饰为靶标的药物在髓母细胞瘤中的应用奠定重要基础。

髓母细胞瘤作为儿童最常见的神经系统恶性肿瘤，其临床诊疗仍面临巨大挑战。本课题通过独特角度阐释了髓母细胞瘤中的DNA羟甲基化异常的潜在机制，确定了5-羟甲基胞嘧啶（5hmC）的降低与低氧状态和疾病进展均存在密切联系。通过本项目的深入研究，我们发现：（1）髓母细胞瘤中5hmC含量显著下降，5hmC可以作为髓母细胞瘤患者预后预测的重要生物标记；（2）低氧能够诱导髓母细胞瘤5hmC含量降低和DNA羟甲基化转移酶TET（Ten-Eleven translocation）的表达异常，并促进HIF-1α/β激活和细胞的增殖；（3）通过解析低氧状态髓母细胞瘤的全基因组5hmC图谱和基因表达，首次揭示了DNA羟甲基化修饰对低氧通路激活的潜在作用。综上所述，本项目的研究成果将为进一步研究髓母细胞瘤表观遗传异常的分子机制和以表观修饰为靶标的药物研发，提供重要理论支持。同时，本项目建立的不同分子亚型的髓母细胞瘤体内、外模型，也将为今后开展髓母细胞瘤分子生物学研究和个体化药物研发提供重要的研究平台。