细胞分子遗传学异常是影响MM预后的最重要因素，补充新的分子预后指标，建立完善的危险度分层体系和个体化治疗策略是MM未来发展的趋势。申请人根据前期血清miRNA芯片结果，选择miRNA-33b在MM患者和正常浆细胞中进行验证，结果miRNA-33b在初治和复发患者中表达明显降低，miRNA-33b低表达的患者总生存时间明显缩短，进一步研究发现，MM细胞miRNA-33b启动子区域甲基化程度明显高于正常人，且miRNA-33b表达与MDM2负相关，与P53蛋白表达呈正相关。据此推测miRNA-33b启动子区域甲基化致其低表达，并通过靶向MDM2进而调节P53蛋白参与MM发病。本课题拟对其具体作用机制进行研究，验证MDM2为其直接作用的靶基因，初步探索去甲基化药物5-Aza 靶向miRNA-33b对MM的治疗作用，为探索以miRNA-33b及其靶基因为新靶点的生物治疗提供可靠的理论和实验依据。

Cytogenetic abnormalities are the hallmark of multiple myeloma (MM) and commonly used as the clinical predictors for determining the stage of disease and providing the guidance for therapeutic strategies. Risk-stratification system based on genetic indicators has been established and recommended by mayo clinic and International Myeloma Working Group (IMWG) in recent years. However, current prognostic factors including del(p53), t(4;14) and amp(CKS1B) can not completely explain the heterogeneity in this disease. So, identifying some novel prognostic markers to better risk stratify patients in the management of myeloma and investigating their roles in myeloma pathogenesis are becoming important issues. Our previous results showed miRNA-33b might be a novel prognostic factor according to the results of serum microRNA microarray. To further ascertain the expression of miRNA-33b in different stages of MM patients, we detected the expression of miRNA-33b in 21 newly diagnosed, 8 relapsed, 8 complete remission (CR) MM patients and 11 normal samples. Our results showed the expression of miRNA-33b was significantly lower in newly diagnosed and relapsed MM patients than CR MM patients and normal samples, and the patients with low expression of miRNA-33b had shorter overall survival (OS) time than control patients. Further studies showed that there was obviously hypermethylation in the promoter region of miRNA-33b in myeloma cells. In addition, transient overexpression of miRNA-33b in U266 and RPMI 8226 cell lines could downregulate the expression of MDM2 protein and upregulate the expression of P53 protein. So, we hypothesize the low expression of miRNA-33b via methylation of promoter CPG region contributes the pathogenesis of myeloma by targeting the p53/MDM2 signal pathway which controls the proliferation and apoptosis. In this project, we will comprehensively investigate the mechanism that miRNA-33b participates in the pathogenesis of MM in vitro and vivo and evaluate the prognostic value of miRNA-33b in MM patients. Taken together, this project will provide the clinical and experimental evidences for miRNA-33b as a novel prognostic factor and valuable therapeutic target.