背景:先前的研究已经提出转录调节因子三部分基序29(TRIM29)通过与核酸结合参与致癌作用。当癌细胞获得治疗抗性特性时,证实TRIM29高表达。我们注意到,通过从基因表达综合(GEO)基因微阵列(GSE142031;log2倍数变化>1,p<0.05)中挖掘数据信息,在耐安洛替尼的NCI-H1975(NCI-H1975/AR)细胞中TRIM29水平显着增加。
目的:我们的研究旨在探讨TRIM29在非小细胞肺癌(NSCLC)细胞对安洛替尼耐药中的作用,包括NCI-H1975和A549细胞。
方法:Real-timeRT-PCR和westernblot检测TRIM29在安洛替尼耐药NSCLC(NSCLC/AR)细胞中的表达。通过流式细胞术测定细胞凋亡,吖啶橙/溴化乙锭染色以及蛋白质印迹。ELISA用于测量C-X3-C基序趋化因子配体1的含量。进行免疫共沉淀测定以验证TRIM29和RAD50双链断裂修复蛋白(RAD50)之间的相互作用。
结果:结果显示,与正常NSCLC细胞相比,TRIM29在NSCLC/AR细胞的细胞质和细胞核中的表达升高。接下来,我们证明TRIM29敲低促进了NSCLC/AR细胞的凋亡并增强了对安洛替尼的敏感性.根据BioGRID数据库引用的精炼结果,证明TRIM29与RAD50相互作用。在这里,RAD50过表达减少了在安洛替尼抗性A549(A549/AR)细胞中通过沉默TRIM29诱导的促凋亡作用。
结论:最后,我们的结论是,NSCLC/AR细胞对安洛替尼的敏感性增加是通过敲低TRIM29实现的,TRIM29敲低的积极作用归因于通过与NSCLC/AR细胞核中的RAD50结合促进细胞凋亡。因此,TRIM29可能成为NSCLC治疗中克服安洛替尼耐药的潜在靶点。
Previous studies have proposed that the transcriptional regulatory factor tripartite motif containing 29 (TRIM29) is involved in carcinogenesis via binding with nucleic acid. TRIM29 is confirmed to be highly expressed when the cancer cells acquire therapy-resistant properties. We noticed that TRIM29 levels were significantly increased in anlotinib-resistant NCIH1975 (NCI-H1975/AR) cells via mining data information from gene expression omnibus (GEO) gene microarray (GSE142031; log2 fold change > 1, p < 0.05).
Our study aimed to investigate the function of TRIM29 on the resistance to anlotinib in non-small cell lung cancer (NSCLC) cells, including NCI-H1975 and A549 cells.
Real-time RT-PCR and western blot were used to detect TRIM29 expression in anlotinib- resistant NSCLC (NSCLC/AR) cells. Apoptosis were determined through flow cytometry, acridine orange/ethidium bromide staining as well as western blot. ELISA was used to measure the content of C-X3-C motif chemokine ligand 1. Co-Immunoprecipitation assay was performed to verify the interaction between TRIM29 and RAD50 double-strand break repair protein (RAD50).
TRIM29 expression was shown to be elevated in the cytoplasm and nucleus of NSCLC/ AR cells compared to normal NSCLC cells. Next, we demonstrated that TRIM29 knockdown facilitated apoptosis and enhanced the sensitivity to anlotinib in NSCLC/AR cells. Based on the refined results citing from the database BioGRID, it was proved that TRIM29 interacted with RAD50. Herein, RAD50 overexpression diminished the pro-apoptotic effect induced by silencing TRIM29 in anlotinib-resistant A549 (A549/AR) cells.
Finally, we concluded that the increased sensitivity to anlotinib in NSCLC/AR cells was achieved by knocking down TRIM29, besides, the positive effects of TRIM29 knockdown were attributed to the promotion of apoptosis via binding to RAD50 in NSCLC/AR cell nucleus. Therefore, TRIM29 might become a potential target for overcoming anlotinib resistance in NSCLC treatment.