DACT1甲基化通过Wnt通路改变WIT-AML生物学效应机制的研究

Acute myeloid leukemia (AML) is high risk in that lack of accurate indicators for disease prognosis and mechanism is unclear. Previous studies had shown that the patients with WIT-AML had worse prognosis in intermediate group, DACT1 was found that 5hmc and expression was decreased and 5mc raised in clinical WIT-AML specimens here, we hypothesized that the changes of methylation in DACT1 make AML cell function to form WIT-AML. The levels of 5mC in DACT1 can be used as an important molecular marker of WIT-AML. To verify the above hypothesis, our group intends to analyze the relationship of hydroxy methylation and expression with DACT1 in children's WIT-AML. We will set up cell of WIT-AML model, use cloning and differentiation experiments, flow cytometry, methylation inhibitors to study mechanisms of DACT1 5hmC and 5mC levels changes make the different biological effect and action in the WIT-AML. Our study is to interprete the pathogenesis of and epigenetic change with methylation in WIT-AML, find a new effective AML typing indicator and provide individualized medical therapeutic target for WIT-AML

急性髓细胞白血病(AML)病程凶险，缺乏准确的临床预后指标且发病机制不清。在本课题组临床研究中发现WIT-AML在中危组预后较差，且首次发现DACT1在临床WIT-AML标本中5hmc降低5mc增高且表达减低，因此我们推测DACT1甲基化水平改变使AML的功能改变从而形成WIT-AML，DACT1甲基化水平可作为WIT-AML的重要分子标志。本研究拟检测儿童WIT-AML DACT1羟甲基化的改变与表达的关系，建立模拟WIT-AML细胞模型，利用细胞克隆与分化实验、流式细胞术、RNA-seq等方法研究DACT1 5hmC及5mC水平改变在WIT-AML中的生物学效应及作用机制，阐释WIT-AML甲基化改变的表观遗传学机制，为DACT1甲基化成为WIT-AML分型的分子标志物提供理论依据，为WIT-AML患者的个体化医疗提供治疗靶点。

项目的背景：AML在儿童白血病中占30%，目前临床上缺乏对儿童 AML预后进行有效分组的指标，难以开展按病人病程好坏而制定治疗方案的的个体治疗，影响AML 的治疗效果。.主要研究内容:研究DACT1 在 WIT-AML 与 N-WIT-AML 中甲基化状态及表达差异, DACT1对 WIT-AML 白血病细胞的生物学效应及DACT1通过抑制c-Myc调控PD-L1表达，促进免疫杀伤的分子机制。.重要结果：DACT1a羟甲基化降低是AML预后不良的独立影响因素，DACT1通过抑制Myc调控PD-L1表达，促进免疫杀伤。.关键数据：糖基化qPCR测定WIT-AML和非WIT-AML中DACT1羟甲基及表达水平DACT1b的5hmC高于正常对照组（U=49.50，P=0.027）；在WIT-AML中DACT1 a/b的羟甲基化均与DACT1的表达水平呈正性相关（r=0.812，P=0.000；r=0.789，P=0.000）；DACT1过表达抑制了U937细胞的增殖和克隆形成能力，增加细胞凋亡，DACT1通过抑制c-Myc调控PD-L1表达，在活体实验中DACT1对肿瘤细胞有抑制作用。.科学意义:该项目为 WIT-AML 找到潜在的分子标志物，揭示 DACT1 甲基化为WIT-AML亚型分子标志物，为医生的准确诊断提供依据。研究提示DACT1可能作为WIT-AML新的治疗靶点。