骨髓增生异常综合征患者p73基因启动子区域异常甲基化的研究

Study of aberrant p73 promoter methylation in patients with myelodysplastic syndrome

目的探讨骨髓增生异常综合征（MDS）患者抑癌基因p73启动子区域甲基化情况及其在预后中的意义。方法收集135例初诊MDS患者及13名正常志愿者骨髓细胞，用甲基化特异性PCR（MS—PCR）方法检测p73基因启动子CpG岛甲基化发生情况，并利用亚硫酸盐测序法验证MS—PCR结果；荧光定量PCR法检测p73mRNA表达情况；利用地西他滨处理MDS患者原代细胞，观察p73基因去甲基化及p73mRNA表达情况；结合临床资料分析p73基因异常高甲基化在MDS中的作用。结果37．0％的MDS患者p73基因启动子呈高甲基化状态，而且高危组MDS患者（RAEB-1、RAEB-2）甲基化发生率明显高于低危组（58．8％对29．7％，P=0．002）；甲基化组患者p73mRNA表达水平显著低于非甲基化组（P=0．032）；用地西他滨处理后，可见MDS原代细胞p73去甲基化，且p73mRNA表达水平增高；p73高甲基化患者无白血病生存时间（DFS）及总体生存时间（OS）低于p73非甲基化患者（P值均〈0．01）。在COX模型中，p73基因甲基化状况和骨髓原始细胞水平为影响患者DFS及OS的独立预后因素。结论p73基因启动子高甲基化在MDS患者中较常见，且提示预后不良，可能为地西他滨治疗的靶点。

Objective To study the methylation status of p73 gene promoter in patients with myelo- dysplastic syndrome （MDS） and explore its significance with clinical prognosis. Methods Methylation of p73 promoter was detected in bone marrow cells from 135 MDS patients and 13 healthy controls by methyla- tion-specific PCR （MSP）. The results of MSP were confirmed by bisulfite sequencing. The expression of p73 mRNA was detected by real-time quantitative PCR. Primary bone marrow cells from MDS patients were treated with decitabine, the changes of p73 methylation status and p73 mRNA expression were measured, The role of p73 methylation in the prognosis of MDS and the correlated clinical data were explored. Results p73 hyper- methylation was present in 37.04% of MDS cases and patients with high risk MDS （ RAEB-1 and RAEB-2） exhibited a significantly higher frequency of p73 methylation than that of low risk MDS （58.8% vs 29.7%, P = 0. 002）. The expression of p73 mRNA in the methylated group was decreased compared to that of the un- methylated group （ P = 0.032）. Decitabine treatment decreased the level of p73 methylation and increased the level of p73 transcripts. Patients with p73 methylation progressed rapidly to AML （P 〈 0. 001 ） and had shor- ter survival （P = 0. 002） than those who did not have p73 methylation. In the multivariate Cox regression model, BM blast and p73 methylation status emerged as independent prognostic factor for overall survival and leukemia free survival. Conclusion p73 gene methylation is common in patients with MDS and may indicate poor prognosis, p73 may be a therapeutic target in MDS.