lncRNAHULC促进HCC细胞保护性自噬的分子机制及其在HCC化疗耐受中的作用研究

Protective autophagy is an important cause leading to chemoresistance of cancer cells, which significantly reduces chemotherapeutic effect of hepatocellular carcinoma (HCC). HULC (up-regulated in liver cancer) is an important long non-coding RNA (lncRNA) for promoting HCC. However, it is not clear whether the promotion effect of HULC is related to the regulation of protective autophagy and chemoresistance. Our preliminary novel findings showed that HULC induces autophagy in HCC cells and reduces the sensitivity of chemotherapeutic drugs. Moreover, we found that HULC indirectly enhances the stability of SIRT1 protein (silent information regulator 1) and upregulates USP22 (ubiquitin-specific processing peptidase 22). Susequently, we found that two microRNAs(miRNAs) (miR-6825 and miR-6845) can downregulate USP22. Bioinformatics predicts that HULC may bind with the two miRNAs. Based on the above novel discoveries, we hypothesize that lncRNA HULC may promote protective autophagy of HCC cells via mediating the pathway “adsorbing miRNAs → upregulating USP22→ increasing the stability of SIRT1 protein”, and the protective autophagy reduces the sensitivity of chemotherapy and enhances the growth of HCC cells. In this project, we will illustrate the mechanisms by which the above pathway forms and which HULC induces protective autophagy. Furthermore, the HCC cells and tumor-bearing mice will be engagaed to evaluate the importance of this pathway in HULC-mediated growth promotion of HCC cells through inducing protective autophagy and subsequently reducing the sensitivity of chemotherapy. These studies may provide scientific evidences for exploring novel anti-HCC strategy targeted HULC and its mediated signaling pathway.

保护性自噬是肿瘤细胞产生耐药的重要原因，其显著降低了肝细胞癌(HCC)的疗效。HULC是一种促HCC的重要lncRNA，但该促进作用是否与调节保护性自噬并参与化疗耐受有关，目前未见报道。我们前期创新性发现：HULC可促进HCC细胞自噬、降低化疗药物敏感性；通过间接方式上调SIRT1蛋白稳定性和USP22水平；两种miRNA可下调USP22，预测提示HULC可能与这两种miRNA结合。据此提出假设：HULC可能通过“吸附miRNA→上调USP22→增加SIRT1蛋白稳定性”这一通路诱导保护性自噬，从而降低化疗敏感性，促进HCC细胞生长。本项目将解析上述通路的形成机制，阐明HULC诱导保护性自噬的分子机理，进而在HCC细胞及荷瘤鼠模型上探讨该通路在HULC通过诱导保护性自噬而降低化疗敏感性，进而促进HCC细胞生长中的重要性，为探索以HULC及其介导的信号通路为靶标的抗HCC新策略提供科学依据。

保护性自噬是肿瘤细胞产生耐药的重要原因，其显著降低了肝细胞癌(HCC)的疗效。HULC是一种促HCC的重要lncRNA，但该促进作用是否与调节保护性自噬并参与化疗耐受有关，目前未见报道。本项目通过体外和体内实验，率先揭示lncRNA HULC 通过诱导细胞保护性自噬而降低化疗敏感性是HULC促进HCC细胞生长的新机制，该机制由“HULC→吸附miRNA→上调USP22→增加SIRT1 蛋白稳定性→促进保护性自噬”通路所介导。为探索以该通路为靶标的增敏HCC 化疗的新措施提供科学依据。所获得的重要结果如下：（1）奥沙利铂、5-氟尿嘧啶、吡柔比星可显著诱导HULC表达和HCC细胞的保护性自噬；（2）HULC 可增加去乙酰化酶SIRT1（Silent information regulator 1） 蛋白的稳定性，从而上调SIRT1 蛋白的水平，进而促进细胞自噬；（3）HULC 上调去泛素化酶USP22( ubiquitin-specific processing peptidase 22) 蛋白的水平，减缓其降解，从而增加SIRT1 蛋白的稳定性，进而促进细胞自噬。（4）通过生物信息学分析、芯片和报告基因检测发现miR-6825-5p, miR-6845-5p、miR-6886-3p 可抑制USP22蛋白表达；（5）HULC 可抑制三个miR-6825-5p, miR-6845-5p、miR-6886-3p 的表达和活性；（6）在HCC 细胞模型上，探讨了“HULC-miRNA-USP22-SIRT1”通路在诱导HCC 细胞保护性自噬，降低化疗药物敏感性，进而促进HCC 细胞生长中的重要性；（7）在HCC 荷瘤裸鼠模型上，探讨了“HULC-miRNA-USP22-SIRT1”通路在诱导HCC细胞保护性自噬，降低化疗药物敏感性，进而促进HCC 生长中的重要作用。