FOXM1基因在piperlongumine抑制肝细胞癌增殖中的作用及机制研究

In the study of drug screening based on the gene expression profile, we have found that piperlongumine can strongly suppress the proliferation of hepatocellular carcinoma (HCC) cells and inhibit the expression of FOXM1 gene. FOXM1 signaling has been reported to be associated with carcinogenesis. Therefore, the FOXM1 has represented a novel therapeutic target for the cancer. Piperlongumine is anti-cancer drug newly found selectively targeting the cancer cells, but the mechanism is poorly understood. We hypothesize that the anti-cancer effect of piperlongumine is mediated by FOXM1 down-regulation. In this project, we shall use the methods of flow cytometry, gene chip, the inter-action of protein and drug,etc, to determine the action that piperlongumine can selectively kill the HCC cells not normal hepatic cell;to illustrate the mechanism that piperlongumine inhibits FOXM1 gene expression, the important role that FOXM1 gene may play in the suppressive effect of piperlongumine on the growth of HCC cancer. This project can contribute to increase the anti-cancer profile of piperlongumine, to provide the evidence for the clinical trial of piperlongumine and its derivants,to offer new strategy for anti-tumor based on the target of FOXM1 gene.

基于基因表达谱的药物筛选研究中，我们发现Piperlongumine具有抗肝细胞癌的作用、抑制FOXM1基因的表达。FOXM1是目前抗肿瘤靶标的热点；Piperlongumine是最近发现的靶向抗肿瘤的药物，但作用机制有待阐明。结合文献，我们提出Piperlongumine对肝细胞癌的抑制作用是由FOXM1基因的抑制介导的。本项目采用流式细胞术、基因芯片、蛋白质与药物的相互作用等实验，明确Piperlongumine选择性抑制肝细胞癌的作用，确定Piperlongumine对FOXM1基因抑制的机制，阐明FOXM1基因在Piperlongumine抑制肿瘤中起到的关键作用。通过本项目的研究有助于扩大Piperlongumine抗肿瘤谱，为piperlongumine及其衍生物的临床应用打下理论基础，明确FOXM1基因是肿瘤治疗的重要靶标，为以FOXM1基因作为药筛靶点治疗肿瘤提供新的思路。

项目背景. Piperlongumine是最近发现的靶向抗肿瘤的药物，但作用机制有待阐明。我们前期通过生物信息学C-MAP分析表明: Piperlongumine可能具有逆转肝细胞癌基因表谱的作用，发现Piperlongumine具有抗肝细胞癌增殖的作用，其中关键的转录调控因子FOXM1可能在其中发挥重要作用。.主要研究内容. 采用流式细胞术、二代测序技术、Cas-9基因敲除FOXM1、siRNA敲减FOXM1表达等实验，明确Piperlongumine具有抑制肝细胞癌的作用，确定Piperlongumine对FOXM1基因抑制的机制，阐明FOXM1基因在Piperlongumine抑制肿瘤中起到的关键作用。.主要结果.（1）.Piperlongumine具有性抑制肝细胞癌细胞株增殖的作用，流式细胞术分析使细胞停滞在G1期。.（2）.Piperlongumine分别在mRNA水平和蛋白质水平上抑制肝细胞癌细胞株FOXM1基因的表达。.（3）.应用二代测序技术测序发现：piperlongumine在多个信号通路中发挥作用。.（4）.采用Cas-9基因敲除肝细胞癌FOXM1基因，发现细胞生长停滞。.（5）.应用siRNA技术敲减肝细胞癌FOXM1基因表达，应用荧光定量PCR技术、westernblot等检测主要降低MMP9、CCND1基因表达。此结果与piperlongumine作用于肝细胞癌的结果一致。.（6）.动物成瘤实验表明，piperlongumine具有抑制移植瘤生长的作用。.科学意义. 通过本项目的研究发现：piperlongumine具有抑制肝细胞癌生长的作用，扩大Piperlongumine抗肿瘤谱。其机制为抑制FOXM1基因表达进而下调CCND1、MMP9基因的表达，从而达到使细胞发生G1期停滞、抑制肝细胞癌的侵袭。此机制的阐明为以FOXM1基因作为药筛靶点治疗肿瘤提供新的思路。