拷贝数变异相关LINC01133通过结合膜联蛋白ANXA2介导肿瘤相关性巨噬细胞浸润及其在肝癌早期复发中的作用与机制研究

研究表明长链非编码RNA（LncRNA）在肿瘤发生发展过程中起至关重要的作用。然而拷贝数变异相关LncRNA在肝癌早期复发的作用及机制仍未明确。基于前期49例术后早期复发的肝癌原发瘤和对应癌旁组织的全基因组测序结果，我们筛选并验证LINC01133在肝癌中拷贝数显著扩增，且和表达量增加呈正相关；体内外试验表明LINC01133促进肝癌生长和转移，并介导肿瘤相关性巨噬细胞（TAM）浸润；RNA pulldown联合质谱分析证实其结合膜联蛋白A2（ANXA2），并稳定该蛋白表达。本课题拟首先明确LINC01133结合ANXA2在促进肝癌生长转移中的作用并揭示其介导TAM浸润的分子机制；然后明确两者结合区段及互作机理；最后在临床肝癌样本中分析LINC01133、ANXA2和TAM的表达及和患者术后早期复发的相关性。通过以上研究，明确拷贝数变异相关LINC01133促进肝癌早期复发的作用和机制。

Large numbers of studies have shown that many long non-coding RNAs (LncRNAs) play essential roles in tumorigenesis and tumor development. The function and mechanism of copy number variation (CNV) associated Intergenic Long Non-coding RNA (LincRNA) in Hepatocellular Carcinoma (HCC) is still unclear. In our previous work, whole-genome sequencing (WGS) was performed using 49 pairs of liver cancer tissue samples from HCC patients who experienced early recurrence after radical resection. And the genomic CNV panorama of HCC with early recurrence after radical resection was described. We screened and verified that the CNV and expression level of LINC01133 were increased in liver tumor. And there was a positive correlation between them. In vivo and in vitro functional experiments showed that LINC01133 promoted tumor growth and metastasis, which mediating tumor-associated macrophages (TAM) infiltration. RNA pull down and mass spectrometry analysis confirmed that LINC01133 was combined with ANXA2 and increasing this protein stability. In this study, we will firstly plan to identify the important role of ANXA2 in LINC01133-mediated HCC growth and metastasis. By phosphorylated protein array and cytokine chip, we will reveal the key mechanism of LINC01133-mediated TAM infiltration. Through RNA pull down and RNA immunocoprecipitation (IP) experiments, the binding region between LINC01133 and ANXA2 protein will be identified; the important role of LINC01133 in maintaining the stability of ANXA2 protein will be explored by RNA IP combined with protein ubiquitination level test. Finally, we will test the expression of LINC01133, ANXA2 and TAM in HCC samples and reveal their correlation with the prognosis of HCC patients. Through the above research, the mechanism of LINC01133 on promoting the growth and metastasis in early recurrence of HCC will be determined, and the experimental evidence for exploring the new intervention targets on HCC treatment will be provided.