Huang Ruili, Zhu Lijuan, Zhang Yali
Department of Gynaecology and Obstetrics, The First People's Hospital of Shangqiu, Henan, No. 292 Kaixuan South Road, 476100 Shangqiu, Henan People's Republic of China.
Cancer Cell Int. 2020 Sep 4;20:436. doi: 10.1186/s12935-020-01500-8. eCollection 2020.
BACKGROUND/AIMS: The expression levels of long non-coding RNA XIST are significantly associated with paclitaxel (Pac) sensitivity in ovarian cancer, but the mechanism of action remains unclear. Therefore, this experimental design was based on lncRNA XIST analysis to regulate the effect of XIST on the tumor stem cell and paclitaxel sensitivity in ovarian cancer.
Sphere assay and fluorescence activated cell sorting (FACS) were used to determine the expression levels of XIST and sensitivity to paclitaxel treatment. The effect of the proliferation was detected by MTT assay. Target gene prediction and screening, luciferase reporter assays were used to validate downstream target genes for lncRNA XIS and KMT2C. The expression of KMT2C was detected by RT-qPCR and Western blotting. RT-qPCR was used to detect the expression of cancer stem cell-associated genes SOX2, OCT4 and Nanog. The tumor changes in mice were detected by in vivo experiments in nude mice.
There was an inverse correlation between the expression of XIST and cancer stem cell (CD44 + /CD24-) population. XIST promoted methylation of histone H3 methylation at lysine 4 by enhancing the stability of lysine (K)-specific methyltransferase 2C (KMT2C) mRNA. XIST acted on the stability of KMT2C mRNA by directly targeting miR-93-5p. Overexpression of miR-93-5p can reverse the XIST overexpression-induced KMT2C decrease and sphere number increase. Overexpression of KMT2C inhibited XIST silencing-induced proliferation of cancer stem cells, and KMT2C was able to mediate paclitaxel resistance induced by XIST in ovarian cancer. The study found that XIST can affect the expression of KMT2C in the ovarian cancer via targeting miR-93-5p.
XIST promoted the sensitivity of ovarian cancer stem cells to paclitaxel in a KMT2C-dependent manner.
背景/目的：长链非编码RNA XIST的表达水平与卵巢癌中紫杉醇（Pac）敏感性显著相关，但其作用机制仍不清楚。因此，本实验设计基于lncRNA XIST分析，以调控XIST对卵巢癌肿瘤干细胞及紫杉醇敏感性的影响。
采用球囊试验和荧光激活细胞分选（FACS）来确定XIST的表达水平及对紫杉醇治疗的敏感性。通过MTT试验检测增殖效应。进行靶基因预测和筛选，利用荧光素酶报告基因试验验证lncRNA XIS和KMT2C的下游靶基因。通过RT-qPCR和蛋白质免疫印迹法检测KMT2C的表达。采用RT-qPCR检测癌症干细胞相关基因SOX2、OCT4和Nanog的表达。通过裸鼠体内实验检测小鼠的肿瘤变化。
XIST的表达与癌症干细胞（CD44 + /CD24 -）群体呈负相关。XIST通过增强赖氨酸（K）特异性甲基转移酶2C（KMT2C）mRNA的稳定性，促进组蛋白H3赖氨酸4位点的甲基化。XIST通过直接靶向miR-93-5p作用于KMT2C mRNA的稳定性。miR-93-5p的过表达可逆转XIST过表达诱导的KMT2C降低和球囊数量增加。KMT2C过表达抑制XIST沉默诱导的癌症干细胞增殖，且KMT2C能够介导XIST在卵巢癌中诱导产生的紫杉醇耐药性。研究发现，XIST可通过靶向miR-93-5p影响卵巢癌中KMT2C的表达。
XIST以KMT2C依赖的方式促进卵巢癌干细胞对紫杉醇的敏感性。
