DNA甲基化氧化通路缺失促进垂体瘤侵袭性生长的分子机制和临床意义

It is not clear yet what causes abnormality of DNA methylation patterns, one of important epigenetic mechanisms that promote the invasive growth of pituitary adenomas. It is reported recently that 5-hydroxymethylcytosine (5hmC) is produced from 5-methylcytosine (5mC) under catalysis of TET enzymes, which provides a new thought for study of aberrant DNA methylation in tumors. Our recent work found that the expression of TET gene, as well as the content of 5hmC, was significantly decreased in invasive pituitary adenomas. It indicates that the oxidation pathway of DNA methylation might be deficient in this type of tumor. It is, however, unknown whether this deficiency is related to aberrant DNA methylation or invasive growth of pituitary adenomas. Therefore, the applicants will investigate the molecular mechanisms leading to inactivation of TET and analyze the relevance between 5hmC and 5mC; then, the applicants will study how TET inactivation impacts on the invasive growth of pituitary adenoma cells in vitro and in animal models; moreover, the applicants will explore the relationship between TET inactivation and pathological changes of tumor or post-operation recurrence through clinical analysis. Based on these investigations, we will know how deficiency in the oxidation pathway of DNA methylation influences on the invasive growth of tumors, which may provide a novel hint or strategy for the effective treatment of invasive pituitary adenomas.

DNA异常甲基化是促进垂体瘤侵袭性生长的重要机制，但造成其异常的原因却并不清楚。5-甲基胞嘧啶(5mC) 可在TET酶的催化下氧化产生5-羟甲基胞嘧啶（ 5hmC），该通路的发现为探索甲基化异常提供了新的思路。我们的前期工作发现侵袭性垂体瘤中TET的表达明显下降，而5hmC含量也显著降低，这提示在该肿瘤中可能存在着DNA甲基化氧化通路的缺失，但该通路缺失是否促使垂体瘤产生DNA异常甲基化和侵袭性生长，还需进一步研究。为此，本课题组将在侵袭性垂体瘤标本中进一步分析引起TET失活的分子机制，并比较5hmC与5mC之间的相关性；随即在细胞水平和动物模型中观察TET失活对肿瘤生长和侵袭性的影响；最后结合临床资料，研究TET失活及5hmC产生与肿瘤病理变化和术后复发的关系，从而探明DNA甲基化氧化通路缺失对垂体瘤侵袭性生长的作用和相关分子机制，为临床更有效的治疗该疾病提供新的策略。

垂体腺瘤(简称垂体瘤)是人类最常见的颅内肿瘤之一，约占颅内肿瘤的 10%~15%。垂体瘤在组织学上属于良性肿瘤，但有些垂体瘤呈侵袭性生长，包绕或侵犯周围组织结构，该部分肿瘤手术难以彻底切除，是肿瘤复发的主要原因之一。侵袭性与非侵袭性垂体腺瘤的临床表现、预后均明显不同。侵袭性垂体腺瘤的坏死、卒中、囊变发生率明显高于非侵袭性垂体腺瘤。侵袭性生长使得外科手术难以做到全切肿瘤，而常用药物溴隐停、生长抑素对这类肿瘤的疗效较差;侵袭性生长同样限制了其对放疗的敏感性，且鞍区放疗容易导致正常垂体、下丘脑及视神经等重要结构的损伤。因此，侵袭性垂体腺瘤术后复发率高，肿瘤残余组织很快增长。显而易见，对侵袭性垂体瘤的治疗是神经外科领域和肿瘤治疗领域面临的巨大挑战。因此，对侵袭性垂体瘤的分子机制进行更加深入的研究可能为彻底战胜该肿瘤供有效的方案。DNA异常甲基化是促进垂体瘤侵袭性生长的重要机制，但造成其异常的原因却并不清楚。5-甲基胞嘧啶可在TET酶的催化下氧化产生5-羟甲基胞嘧啶，该通路的发现为探索甲基化异常供了新的思路。本项目主要研究了：1、侵袭性垂体瘤中TET2表达的变化，分析TET2与垂体瘤侵袭性的关系。2、侵袭性垂体瘤中TET2失活的分子机制。3、TET2活性对垂体瘤侵袭性生长的作用及分子机制。4、裸鼠模型中TET2活性与肿瘤生长的关系。利用高通量测序分析、重亚硫酸盐转化PCR及免疫共沉淀等技术手段发现：在对垂体瘤侵袭性的调节中TET2起到关键作用，TET2能够通过促进CDH1以及长链非编码RNA MEG3抑制垂体瘤细胞的迁移，能够与WT1相互结合促进MEG3的表达。同时对转录组数据的深入分析发现侵袭性垂腺瘤中调节TET2活性的关键酶IDH1表达量显著降低，是侵袭性垂体瘤中TET2酶活性失活的主要原因。此外我们筛选并验证了12个与侵袭性垂体瘤相关的分子生物标记申报了国家发明专利，有望为侵袭性垂体瘤临床更有效诊断和评估预后提供策略。