背景:放射治疗是治疗晚期胃癌(GC)的重要策略,而抗辐射性限制了其有效性。核仁和纺锤体相关蛋白1(NUSAP1)与癌症进展和化疗耐药有关。然而,NUSAP1影响GC放射抗性的潜在机制仍然未知.
方法:系统评价NUSAP1在人类癌症中的预后价值。使用癌症基因组图谱(TCGA)和基因表达综合(GEO)数据集进行基因集富集分析(GSEA)。通过qRT-PCR和westernblot检测mRNA和蛋白的表达,分别。通过集落形成观察GC细胞的放射敏感性,流式细胞术,彗星,免疫荧光,和动物试验。使用免疫沉淀测定和质谱来鉴定蛋白质关联。通过starbase预测确定与NUSAP1结合的miRNA,荧光素酶报告基因,和RNA免疫沉淀(RIP)测定。
结果:NUSAP1高表达预测总体生存期(OS)和无病生存期(DFS)较差,通过荟萃分析没有统计学异质性。NUSAP1的下调通过抑制集落形成显著增加GC放射敏感性,DNA损伤修复,并促进照射后细胞凋亡。此外,NUSAP1沉默与辐射组合在异种移植小鼠模型中产生协同抗肿瘤作用。机械上,NUSAP1与ANXA2相互作用,通过阻碍其泛素化过程保护其免受蛋白质降解。NUSAP1被证实为miR-129-5p的靶标并被其负调控。
结论:我们的结果表明NUSAP1增强了GC细胞的辐射抗性。NUSAP1可能是增加GC放射敏感性的有希望的靶标。
BACKGROUND: Radiotherapy is an important strategy for the treatment of advanced gastric cancer (GC), while the radioresistance limits its effectiveness. Nucleolar and spindle associated protein 1 (NUSAP1) was implicated in cancer progression and chemoresistance. However, the underlying mechanisms of NUSAP1 influencing GC radioresistance remain largely unknown.
METHODS: Meta-analysis was conducted to systematically evaluate the prognostic value of NUSAP1 in human cancers. Gene set enrichment analysis (GSEA) was conducted using The Cancer Genome Atlas (TCGA) and gene expression omnibus (GEO) datasets. MRNA and protein expressions were detected by qRT-PCR and western blot, respectively. The radiosensitivity of GC cells was observed by colony formation, flow cytometry, comet, immunofluorescence, and animal assays. Immunoprecipitation assay and mass spectrometry were utilized to identify protein associations. MiRNAs binding with NUSAP1 were determined by starbase prediction, luciferase reporter, and RNA immunoprecipitation (RIP) assays.
RESULTS: NUSAP1 high expression predicted worse overall survival (OS) and disease-free survival (DFS) with no statistical heterogeneity through the meta-analysis. Downregulation of NUSAP1 significantly increased GC radiosensitivity by inhibiting colony formation, DNA damage repair, and promoting apoptosis following irradiation. Additionally, NUSAP1 silencing combined with radiation resulted in a synergistic anti-tumor effect in xenograft mouse model. Mechanistically, NUSAP1 interacted with ANXA2, protecting it against protein degradation via impeding its ubiquitination process. NUSAP1 was confirmed as a target of miR-129-5p and negatively regulated by it.
CONCLUSIONS: Our results suggested that NUSAP1 enhanced the radioresistance of GC cells. NUSAP1 could be a promising target to increase GC radiosensitivity.