miR-106a/20b cluster介导CCND1调控乳腺癌干细胞增殖和自我更新的研究

Clarifying the mechanism of stemness regulation of breast cancer stem cells is very important for improving prognosis of breast cancer. We have previously built up the miRNAs signalling network and observed that novel miR-106a/20b cluster takes an essential role in regulating stemness of breast cancer stem cells. In this study, we will firstly verify the role of miR-106a/20b cluster in regulating biological function of the tumor stem cells and validate structurally and functionally the authenticity of regulation of CCND1 by miR-106a/20b cluster; secondly, in breast cancer stem cells, by overexpressing CCND1, we will verify the regulatory effect of miR-106a/20b cluster- mediated CCND1 on stemness of cancer stem cells; thirdly, in breast cancer cells, by gene silencing of CCND1, we will further verify the regulatory effect of miR-106a/20b cluster - mediated CCND1 on transformation of cancer cells to cancer stem cells; finally, we will investigate the relationship between miR-106a/20b cluster and breast cancer stem cells and its clinicopathological significance histologically. Our results will be of theoretical significance and have vast vistas for clinical applications with regard to exploring the cancer stem cell origin of breast cancer and its molecular targets.

阐明乳腺癌干细胞干性调控机制，对于改善乳腺癌预后具有重要意义。前期研究建立了乳腺癌干细胞miRNAs信号网络，发现新型miR-106a/20b cluster在乳腺癌干细胞干性调控中具有重要作用。拟首先确证miR-106a/20b cluster调控乳腺癌干细胞的生物学功能，并从结构学和功能学上，验证其调控CCND1的真实性；其次，在乳腺癌干细胞中，通过对CCND1的过表达，正向验证miR-106a/20b cluster介导CCND1对癌干细胞的干性特征的调控；在普通乳腺癌细胞中，通过对CCND1的基因沉默，反向验证miR-106a/20b cluster介导CCND1对癌细胞干性转化的调控；最后，从临床标本验证miR-106a/20b cluster与乳腺癌干细胞的关系及其临床病理意义。研究结果对于探索乳腺癌干细胞本质及其分子靶点，具有重要的理论意义和临床应用前景。

阐明乳腺癌干细胞干性调控机制，对于改善乳腺癌预后具有重要意义。本研究发现：从乳腺癌干细胞miRNAs表达谱网络中，筛选到具有较高调控维度的新型miR-106a/20b cluster；无血清培养可诱导MCF-7和T47D成球生长，CD44+CD24-/low细胞群比例显著升高、多个干性基因表达升高，显示出干细胞特性；T47D/MCF-7干性细胞群中miR-106a/miR-20b表达较非干性乳腺癌细胞升高；稳定过表达miR-106a/20b的T47D干性细胞群，结果显示miR-106a促进细胞增殖，miR-106a/20b抑制细胞凋亡，miR-106a/miR-20b促进细胞成球能力；在体实验显示miR-20b促进裸鼠移植瘤生长；靶基因预测显示CCND1和E2F1等为潜在靶基因，双荧光素酶报告基因实验显示，miR-106a/20b与E2F1的3'UTR结合，过表达miR-106a/20b可抑制E2F1的蛋白水平。本研究结果是乳腺癌干细胞本质的新认识，具有重要的理论意义。阐明乳腺癌干细胞干性调控机制，对于改善乳腺癌预后具有重要意义。本研究发现：从乳腺癌干细胞miRNAs表达谱网络中，筛选到具有较高调控维度的新型miR-106a/20b cluster；无血清培养可诱导MCF-7和T47D成球生长，CD44+CD24-/low细胞群比例显著升高、多个干性基因表达升高，显示出干细胞特性；T47D/MCF-7干性细胞群中miR-106a/miR-20b表达较非干性乳腺癌细胞升高；稳定过表达miR-106a/20b的T47D干性细胞群，结果显示miR-106a促进细胞增殖，miR-106a/20b抑制细胞凋亡，miR-106a/miR-20b促进细胞成球能力；在体实验显示miR-20b促进裸鼠移植瘤生长；靶基因预测显示CCND1和E2F1等为潜在靶基因，双荧光素酶报告基因实验显示，miR-106a/20b与E2F1的3'UTR结合，过表达miR-106a/20b可抑制E2F1的蛋白水平。本研究结果是乳腺癌干细胞本质的新认识，具有重要的理论意义。