PEG10调控β-catenin介导非小细胞肺癌化疗耐药的机制

Chemoresistance is the main cause of chemotherapy failure in non-small cell lung cancer (NSCLC) patients. Preliminary results indicated that the expression of PEG10 was higher in NSCLC chemotherapy patients with non remission than in chemotherapy patients with remission, while silence of PEG10 significantly enhanced the chemosensitivity of NSCLC chemoresistance cell lines. Previous studies found that PEG10 regulates β-catenin expression, an important chemoresistance gene. Additionally, PEG10 reduces stability of SIAH1 protein and inhibits the ubiquitination of β-catenin by combining with SIAH1, a ubiquitin ligase of phosphorylation nondependent ubiquitination. Therefore, we speculated that PEG10 regulates NSCLC chemoresistance through promotes expression of β-catenin and inhibits β-catenin degradation by binding with SIAH1. This project aims at defining relationship between PEG10 and NSCLC chemoresistance, and clarify the role of β-catenin on PEG10 mediated NSCLC chemoresistance at clinical samples, cells, animal levels. Finally, utilizing truncated mutations and GST pull down assays to analyze the binding sites between PEG10 and SIAH1.This study will clarify the mechanism of PEG10 on NSCLC chemoresistance, which is expected to provide a new target for overcoming NSCLC chemoresistance.

化疗耐药是非小细胞肺癌（NSCLC）患者化疗失败的重要原因。预实验结果提示PEG10基因在NSCLC化疗未缓解患者表达水平显著升高，沉默PEG10显著增强NSCLC耐药细胞化疗敏感性。前期研究发现PEG10调控耐药重要基因β-catenin表达，另发现PEG10可与β-catenin磷酸化非依赖泛素化途径泛素连接酶SIAH1结合，并降低SIAH1蛋白稳定性，抑制β-catenin泛素化。据此推测PEG10促进β-catenin表达并通过与SIAH1结合抑制β-catenin泛素化降解，进而调控NSCLC化疗耐药。本研究拟定从临床样本、细胞、动物三个水平明确PEG10与NSCLC化疗敏感性关系，并阐明β-catenin在PEG10介导NSCLC化疗耐药中的作用，最后解析PEG10与SIAH1结合位点。本研究将阐明PEG10介导NSCLC化疗耐药机制，有望为克服NSCLC化疗耐药提供新靶点。

目前临床上尚无逆转NSCLC化疗耐药的有效靶点，因此探索NSCLC耐药机制，寻找克服NSCLC化疗耐药的新靶点十分重要。本课题探究了PEG10表达水平与 NSCLC化疗敏感性的关系，β-catenin在PEG10调控NSCLC化疗耐药中的作用，以及PE10与SIAH1互作调控β-catenin表达的分子机制。我们研究发现PEG10在化疗耐受NSCLC患者肿瘤组织中高表达，且与患者不良预后密切相关。过表达PEG10显著增强NSCLC细胞株化疗耐药性，而沉默PEG10则相反。抑制β-catenin显著削弱PEG10过表达诱导的NSCLC细胞化疗耐药，而激活β-catenin则出现相反的趋势。抑制蛋白质合和蛋白酶体功能后，沉默PEG10仍然降低NSCLC细胞β-catenin表达，PEG10与β-catenin表达水平呈正相关。过表达PEG10显著降低SIAH1的表达水平，并削弱过表达SIAH1诱导的β-catenin泛素化。过表达PEG10显著增强β-catenin表达和核定位，过表达SIAH1显著抑制表达和β-catenin核定位，过表达PEG10可中和过表达SIAH1对β-catenin表达和核定位的抑制。SIAH1与PEG10关键结合区域为：V(17)—T(61)、 L(92)—G(136)、 P(157)—G(171)、K(227)—(286) 。过表达PEG10显著增强多药耐药蛋白MRP1的表达，抑制β-catenin则显著削弱PEG10过表达诱导的MRP1表达，沉默PEG10或激活β-catenin则相反。我们的研究表明PEG10通过与SIAH1互作上调β-catenin表达水平，并促进耐药相关蛋白MRP1表达，促进NSCLC细胞的药物外排，抑制化疗诱导的细胞凋亡，进而促使NSCLC化疗耐药。本研究有望为克服NSCLC化疗耐药问题提供新的有效靶点。