基于转录组测序分析人大细胞肺癌NCI-H460细胞对类泛素化抑制剂MLN4924的潜在耐药机制  

Mechanisms of drug resistance to the neddylation inhibitor MLN4924 in NCI-H460 cells based on transcriptome sequencing analysis

作　　者：邢磊 史镜琪 李荣艳 刘静 刘建伟 叶玲 张明华[3] 范皎 Xing Lei;Shi Jingqi;Li Rongyan;Liu Jing;Liu Jianwei;Ye Ling;Zhang Minghua;Fan Jiao(Medical school of Chinese PLA,Institute of Geriatrics,the Second Medical Center,Chinese PLA General Hospital,Beijing 100853,China;Institute of Geriatrics,the Second Medical Center,Chinese PLA General Hospital,National Clinical Research Center for Geriatric Diseases,Beijing 100853,China;Medical Supplies Center of PLA General Hospital,Beijing 100853,China)

机构地区：[1]解放军医学院,解放军总医院第二医学中心老年医学研究所,北京100853 [2]中国人民解放军总医院第二医学中心老年医学研究所、国家老年疾病临床医学研究中心,北京100853 [3]中国人民解放军总医院医疗保障中心,北京100853

出　　处：《中华细胞与干细胞杂志（电子版）》2024年第1期1-10,共10页Chinese Journal of Cell and Stem Cell（Electronic Edition）

基　　金：北京市科技新星计划(20220484020)。

摘　　要：目的探讨人大细胞肺癌NCI-H460细胞对类泛素化抑制剂MLN4924潜在新型耐药机制。方法利用MLN4924处理NCI-H460细胞,筛选和建立耐药细胞株。对普通NCI-H460对照细胞、MLN4924处理8 h加药细胞株、48 h加药细胞株和建立成功的耐药株进行高通量转录组测序。根据测序结果筛选差异表达基因,并进行通路富集分析。多组间比较采用单因素方差分析,组间两两比较采用Dunnett-t检验。结果共构建3个NCI-H460耐药细胞株。转录组测序分析显示,与对照比较,MLN4924处理8 h加药株、处理48 h加药株、三株耐药株分别筛选出5303、4186、3388、3675和3267个差异表达基因,包括ABCA9、ABCA13、CYR61和CYP39A1等基因。RT-qRCR实验进一步验证了在瞬时加药组和耐药株中ABCA9和CYR61基因高表达。GO富集分析结果显示,差异表达基因与细胞周期、细胞凋亡、细胞间信号转导和细胞黏附等细胞生物过程高度相关,还与细胞核和细胞膜等细胞组分密切相关。KEGG通路富集分析结果显示,耐药细胞中差异表达基因集中在MAPK信号通路、PI3K/Akt信号通路和cAMP信号通路等。结论在人大细胞肺癌NCI-H460细胞中,ABCA9、CYR61、ABCC8和CYP4F12等基因的表达变化可能会提高其对MLN4924的耐药能力。这种耐药能力可能与MAPK信号通路、PI3K/Akt信号通路和cAMP信号通路有关。Objective This study aims to explore potential novel drug resistance mechanism of NCI-H460 cells to the neddylation inhibitor MLN4924.Methods NCI-H460 cells were treated by MLN4924 to screen and establish drug-resistant cell lines.High throughput transcriptome sequencing was performed on the drug-resistant cell lines,MLN4924 treated 8-hour cell lines,48-hour cell lines,and control cells.Then we screened differentially expressed genes based on sequencing results and conducted pathway enrichment analysis.The comparison among multiple groups was used by one-way ANOVA,and the pairwise comparison between two groups were performed by Dunnett-t test.Results Three MLN4924-resistant cell lines were constructed.Transcriptome sequencing analysis showed that compared to the normal control group,there were 5303,4186,3388,3675,and 3267 differentially expressed genes for MLN4924 treated 8-hour,48-hour cell lines,and three drug-resistant cell lines,respectively,including genes such as ABCA9,ABCA13,CYR61,CYP39A1,etc.The RT-qPCR experiment further confirmed the significant high expression of ABCA9 and CYR61 in the transient treated cells and drug-resistant cells.The GO enrichment analysis results showed that differentially expressed genes were highly correlated with cellular biological processes such as cell cycle,apoptosis,intercellular signal transduction,and cell adhesion,as well as with cell components such as nucleus and cell membrane.KEGG pathway enrichment analysis showed that differentially expressed genes in drug-resistant cells were concentrated in MAPK signaling pathway,PI3K/Akt signaling pathway and cAMP signaling pathway.Conclusion In NCI-H460 human lung cancer cells,changes in the expression of genes such as ABCA9,CYR61,ABCC8,and CYP4F12 may enhance their resistance to MLN4924.This resistance may be related to the MAPK signaling pathway,PI3K Akt signaling pathway,and cAMP signaling pathway.

关 键 词：MLN4924 高通量测序 耐药机制 类泛素化 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]