转录偶联修复基因在非小细胞肺癌异常表达的分子机制研究

Lung cancer is widespread all over the world, with high incidence rate and leading lethality. Non-small cell lung cancer accounts for about 85% total lung cancer. Lung cancer can be initiated by DNA damage caused by environmental carcinogens. However, not all exposed individuals develop lung cancer, suggesting that the genetic makeup is also important in the development of this malignancy. Transcription coupled repair (LCR) is one of important mechanisms, which repair DNA damage resulting from environmental carcinogens. The genetic variation and epigenetic change in transcription coupled repair genes which purpose to determine the level of expression of mRNAs, may be responsible for the inter-individual diversity in DNA repair capacity and cancer susceptibility. In this project, we will identify abnormal expressed LCR genes in lung cancer and discover the genetic and epigenetic mechanism of abnormal expression of these genes. Based on this we will conduct the case-control study to investigate the association between identified functional genetic variation and the risk for developing lung cancer. These observations will represent enhanced understanding of the importance of transcription-coupled repair in the lung cancer development. The identified genetic signatures may be a powerful predictor for LCR initiated cancer phenotypes and will be of tremendous benefit for providing information on lung cancer prevention and personalized chemotherapy regiments for cancer patients.

非小细胞肺癌是全世界最常见的恶性肿瘤之一，其发生和发展与环境致癌物密切相关。然而，暴露于相同剂量同一致癌物的不同个体，发生肺癌的风险及其严重程度存在着巨大差异，这说明个体遗传背景在这一过程中发挥着至关重要的作用。转录偶联修复通路是机体清除外来致癌物所致DNA损伤的主要方式之一。位于转录偶联修复关键基因的遗传变异和表遗传学变化可能通过调节基因转录而影响DNA修复能力，从而导致人群中肿瘤易感性的个体差异。本项目拟对非小细胞肺癌中异常表达的转录偶联修复基因（如CSA、CSB和XPG）进行研究，探讨它们的遗传变异和表遗传学变化如何影响基因表达以及它们对非小细胞肺癌发病的影响。本研究若能获得资助，将从整体上阐明转录偶联修复基因在非小细胞肺癌发生发展过程中的重要作用；所识别的遗传学和表遗传学指标可能作为潜在的分子标志物用于高风险个体的筛查和早期检测，也为非小细胞肺癌的预防和个体化治疗提供潜在的靶点。

该项目的目的在于探讨转录偶联修复关键基因在肺癌发生发展中的作用及机制。我们发现转录偶联修复基因XAB2遗传变异rs794078 G>A变异和非小细胞肺癌发病风险相关，AA或AG基因型携带者发生肺癌风险的OR(95%CI)为0.12(0.03-0.54);发现rs4134816 C等位基因携带者发生非小细胞肺癌的风险显著降低(OR=0.46; 95%CI=0.26-0. 84)。对转录偶联修复基因标签遗传变异与非小细胞肺癌铂类药物化疗后的预后分析，发现ERCC6 rs2228526 AA/rs4253101 AA基因型携带者中位生存期(29个月)显著高于rs2228526 AG/rs4253101 AC基因型携带者的中位生存期(21个月)，多因素Cox回归分析显示ERCC6 rs2228526 AG/rs4253101变异是非小细胞肺癌铂类药物化疗后的独立预后因素。我们用TargetScan(Release7.0)和PicTar等线上软件预测ERCC6 3’UTR上miRNA靶点 ，发现has-let-7是可能结合于ERCC6靶基因3ˊUTR上的靶点。进一步细胞研究结果表明 ERCC6的表达受Hsa-let-7f调节，该miRNA可抑制荧光素酶报告基因活性约60%。我们了构建siRNA-ERCC5 A慢病毒载体(GV248-ERCC5-RNAi)，并将其稳定转染肺癌细胞，结果发现转染ERCC5 siRNA的肺癌细胞，ERCC5 mRNA水平降低约67%。铂类药物治疗是肺癌主要治疗手段之一，我们将转染siRNA-ERCC5和对照siRNA 的H23细胞用不同浓度顺铂处理，然后用CCK-8检测，结果发现肺癌细胞对顺铂的半数抑制浓度IC50在siRNA-ERCC5组(6.52μmol/L)明显高于对照组(4.16μmol/L)。这些结果对于理解ERCC6和ERCC5在肺癌发生中的作用及调节机制具有重要作用，对非小细胞肺癌的发病风险评估以及预后预测也具有重要意义。该项目共发表研究论文4篇，培养硕士研究生5名。