LINC02085竞争结合miRNA-595调控ABCB1表达影响卵巢癌铂耐药的机制研究

Platinum resistance is seriously adverse to chemotherapeutic efficacy and prognosis of ovarian cancer patients. Unfortunately, the exact mechanism of platinum resistance is still unknown. In our previous work, we have confirmed that downregulated miRNA-595 reduces the inhibitory effect on ABCB1, which aggravates platinum resistance in ovarian cancer. Furthermore, we predicted that LINC02085 might be a ceRNA of ABCB1 by integrating the level of sharing miRNAs and the level of expression correlations. Meanwhile, the results of pre-experiments showed that LINC02085 was overexpressed in platinum-resistant ovarian cancer tissues. In addition, the expression level of LINC02085 significantly affected the sensitivity of ovarian cancer cell line to platinum. In terms of expression level in ovarian cancer tissues, we also confirmed that LINC02085 was negatively correlated with miRNA-595 and positively correlated with ABCB1. Therefore, it is assumed that LINC02085 may competitively targeting miRNA-595 and causes up-regulation of ABCB1, which further lead to platinum resistance in ovarian cancer. In order to validate this hypothesis, we are going to verify the ceRNA relationship between LINC02085, miRNA-595 and ABCB1. Next, we will confirm the effect of LINC02085 regulating ABCB1 expression by competitively targeting with miRNA-595 on platinum sensitivity phenotype of ovarian cancer. In conclusion, we will prove that LINC02085, miRNA-595 and ABCB1 can be used as platinum-resistant markers of ovarian cancer. At the same time, we will further elaborate the mechanism of platinum resistance by ceRNA regulatory mechanism. All of these conclusions are beneficial to guiding the medication choice for gynecologic oncologist and promoting the prognosis of ovarian cancer patients.

卵巢癌铂耐药严重影响病人化疗效果及预后且机制不明。我们前期研究证实，miRNA-595在卵巢癌中下调并减弱对ABCB1的抑制而导致铂耐药。课题组整合miRNA共享水平及表达相关水平预测出LINC02085可能为ABCB1的ceRNA。预实验结果显示：铂耐药型卵巢癌组织中LINC02085高表达；其表达量影响卵巢癌细胞对铂的敏感性；卵巢癌组织中LINC02085表达与miRNA-595表达呈负相关，而与ABCB1表达呈正相关。由此假设LINC02085可能与ABCB1竞争结合miRNA-595导致ABCB1上调引起铂耐药。本项目拟利用分子生物学实验验证三者的ceRNA关系，证实LINC02085竞争结合miRNA-595调控ABCB1表达对铂敏感性的影响。通过上述分析证实三者可作为卵巢癌铂耐药标志物，并深入阐述ceRNA相关铂耐药机制，为指导妇瘤科医生临床用药和促进卵巢癌预后提供理论基础。

卵巢癌铂耐药严重影响病人化疗效果及预后且机制不明。按照课题设计拟在细胞水平上证实LINC02085，miRNA-595和ABCB1三元组的ceRNA关系。课题执行过程中遇到难以解决的问题且未能寻找到关键突破口。项目组利用TCGA数据库中卵巢癌患者用药信息，预测了铂耐药和铂敏感患者的差异表达基因。进一步，我们将差异表达基因作为种子节点注释到蛋白互作网中，并通过随机游走算法进行分析，最终筛选出PRSS1进行后续实验。我们利用Oncomine和cBioportal for Cancer Genomics等数据库研究了PRSS1的表达模式，揭示了其在卵巢癌中的关键作用。RT-PCR和Western blot检测结果显示铂耐药组织或细胞中PRSS1表达水平显著高于铂敏感样品。PRSS1基因敲减或过表达分别降低和增加了卵巢癌细胞对顺铂的耐药性。我们发现了一个新的与卵巢癌铂耐药相关的风险基因PRSS1，这一成果初步识别了PRSS1作为卵巢癌铂耐药标志物和新的靶向治疗策略的可能性，有望改善卵巢癌铂耐药棘手的现状和延长卵巢癌患者的生命。此外，课题组全面描述了33种癌症中10种高血压相关基因的参与情况。每一种类型的癌症都鉴定出每个基因的体细胞拷贝数改变和单核苷酸变异。我们发现高血压相关基因在14种癌症中均有异常表达且有的与患者的总生存期或癌症分期有关。同时，我们也发现某些高血压相关基因与临床可作用基因相关。我们也识别了高血压相关基因的相关通路。与课题组研究方向密切相关的是我们发现高血压相关基因均与癌症对大多数抗癌药物的耐药性呈正相关或负相关。这些结果强调了高血压相关基因在癌症中的重要性。课题组尤其关注高血压相关基因与抗癌药物药敏的相关性，这也将为未来本课题组肿瘤耐药相关研究奠定一定基础。课题负责人在项目资助期间曾去美国加州大学圣地亚哥分校访学。在该校访学期间参与了RSPO1相关靶向药物的研制及其在卵巢癌中的应用的研究。归国后对RSPO蛋白及其受体LGR4-6在泛癌及卵巢癌中的表达和其对卵巢癌表型的影响机制进行了深入的探索并取得实质性的成果，正总结成文。