RPA1基因多态性在奥沙利铂治疗结直肠癌敏感性中的研究

发布时间：2018-06-23 17:20

本文选题：核酸切除修复基因 + 遗传变异　；参考：《南京中医药大学》2017年硕士论文

【摘要】：背景:结直肠癌(包括结肠癌与直肠癌)是世界上常见的消化系统恶性肿瘤之一,在我国,结直肠癌的发病率目前已呈逐年上升趋势,严重威胁着人类的健康。尽管目前筛查早期结直肠癌的方法为数不少,但由于结直肠癌起病隐匿,大部分患者确诊时已发展至中晚期,无法进行手术治疗,因而辅助化疗已成为结直肠癌治疗中不可或缺的手段之一。奥沙利铂(oxaliplatin)是第三代铂类化合物,是目前针对中晚期结直肠癌化疗联合方案的标准药物之一,其主要原理是通过与脱氧核糖核酸双链共价结合形成链间交联,阻断肿瘤细胞DNA的复制和转录,从而发挥抗肿瘤作用。核酸切除修复系统(nucleotide excision repair,NER)是DNA修复的基本通路,修复紫外线所致嘧啶二聚体、脱氧核糖核酸加合物、光化合物等累积性损伤,是清除大剂量铂类药所引起的脱氧核糖核酸损伤的重要系统。人类基因组计划已经确认,基因的遗传与变异会影响基因的结构、功能以及表达,进而改变机体的生物学功能。单核苷酸多态性(single nucleotide polymorphisms,SNPs)是最常见的基因遗传变异。能够改变基因结构或调节基因表达量,从而调控机体的作用。研究发现,单个SNP作用或多个SNPs间的联合作用与肿瘤发生发展及预后显著相关。耐药现象主要产生于修复系统清除药物,这是减弱化疗药物疗效的主要原因,而DNA修复基因的SNP密切影响着与修复系统的修复能力。迄今为止,已有多篇DNA修复通路SNPs与肿瘤及结直肠癌发病风险和化疗敏感性研究的报道。大量研究结果证实,NER通路与结直肠癌化疗敏感性以及生存预后存在潜在的关联。方法:本课题通过对166例结直肠癌患者的临床病例研究,筛选NER通路中可能存在的重要基因遗传变异,统计其与结直肠癌化疗敏感性及生存预后的关联,寻找到相关的遗传变异rs5030740及所在基因RPA1。在此基础上,通过分子生物学方法研究关键基因RPA1及遗传变异rs5030740对肿瘤细胞的表型影响,进而揭示其在奥沙利铂化疗敏感性及生存预后中的可能机制,为预测结直肠癌化疗敏感性及进行个体化治疗提供理论依据。结果:对166个以奥沙利铂为主要治疗方式的结直肠癌患者进行化疗敏感性及生存预后与RPA1遗传变异的关联性分析,寻找到相关的遗传变异rs5030740及所在基因RPA1。课题发现rs5030740位点T等位基因突变到C等位基因与结直肠癌患者奥沙利铂化疗敏感性降低有关,并且结直肠癌患者用药后无进展生存期缩短有关。此外,本课题组通过网站预测,数据库分析,细胞学实验等验证了 rs5030740位点的突变导致了 miRNA-let-7e-5p与PPA1基因3'-UTR的结合能力的降低,促使RPA1的表达量上升。最后,课题组通过一系列细胞表型实验(细胞增殖、周期及凋亡)进一步揭示了RPA1的表达量升高可增加肿瘤细胞的增殖能力,减少细胞凋亡数目,而这一现象在奥沙利铂药物的刺激下,更为明显。结论:基因rs5030740的突变可能导致miRNA-let-7e-5p的结合能力减弱,RPA1的异常高表达,进而通过增加肿瘤细胞的增殖力及减少凋亡率来延长肿瘤的生存能力,最终影响了奥沙利铂化疗药物的效果及结直肠癌患者的生存预后。该研究提示了 RPA1基因rs5030740多态性可作为中国结直肠癌人群奥沙利铂疗效及预后的重要的潜在生物学指标。
[Abstract]:Background: colorectal cancer (including colon and rectal cancer) is one of the most common digestive malignant tumors in the world. In China, the incidence of colorectal cancer is now increasing year by year, which seriously threatens human health. Although there are many methods of screening colorectal cancer at the early stage, the majority of colorectal cancer are hidden and most of them are hidden. The patient has been developed to the middle and late stages and can not be operated on. Therefore, adjuvant chemotherapy has become one of the indispensable means in the treatment of colorectal cancer. Oxaliplatin (oxaliplatin) is the third generation of platinum compounds, which is one of the standard drugs for the combination of advanced colorectal cancer chemotherapy. The double chain covalent binding of oxyribonucleic acid forms interchain crosslinking to block the replication and transcription of tumor cell DNA and thus plays an antitumor effect. The nucleic acid excision repair system (nucleotide excision repair, NER) is the basic pathway of DNA repair, and repairs the cumulative damage of UV induced pyrimidine two polymer, deoxyribonucleic acid adducts, and light compounds. It is an important system for deoxyribonucleic acid damage caused by large doses of platinum drugs. The human genome project has confirmed that genetic and mutation of genes affect the structure, function, and expression of genes, and then change the biological function of the body. Single nucleotide polymorphisms (SNPs) is the most common gene. Genetic variation. It can change the structure of the gene or regulate the amount of gene expression, which regulates the role of the body. Studies have found that a single SNP action or a combination of multiple SNPs interactions is significantly related to the development and prognosis of the tumor. The SNP of the complex gene is closely related to the repair ability of the repair system. So far, there have been several reports of the DNA repair pathway SNPs and the risk of cancer and colorectal cancer and chemosensitivity. A large number of results have shown that there is a potential association between the NER pathway and the chemotherapy sensitivity and survival of colorectal cancer. After studying the clinical case study of 166 patients with colorectal cancer, we screened the important genetic variation that may exist in the NER pathway, statistics its association with the chemosensitivity of colorectal cancer and the survival prognosis. On the basis of finding the related genetic variation rs5030740 and the gene RPA1., the key gene RPA1 is studied by molecular biology method. And the effect of genetic variation rs5030740 on the phenotype of tumor cells, and then reveal its possible mechanism in the chemotherapeutic sensitivity and survival of oxaliplatin, providing a theoretical basis for predicting chemotherapy sensitivity and individualized treatment of colorectal cancer. Results: chemotherapy for 166 colorectal cancer patients with oxaliplatin as the main treatment method The correlation analysis of sensitivity, survival prognosis and RPA1 genetic variation, finding the related genetic variation rs5030740 and the gene RPA1., found that the mutation of the rs5030740 loci T allele to the C allele was associated with the reduction of oxaliplatin chemosensitivity in colorectal cancer patients, and the non progression survival period of colorectal cancer patients was reduced. In addition, the research group confirmed that the mutation of rs5030740 loci led to the decrease of the binding capacity of miRNA-let-7e-5p and PPA1 gene 3'-UTR by site prediction, database analysis, and cytological experiments. Finally, the group through a series of cell phenotypic experiments (cell proliferation, cycle and apoptosis) into one. The increase in the expression of RPA1 can increase the proliferation of tumor cells and reduce the number of apoptotic cells. This phenomenon is more obvious under the stimulation of oxaliplatin. Conclusion: the mutation of the gene rs5030740 may lead to the weakening of the binding capacity of miRNA-let-7e-5p, the abnormal high expression of RPA1, and the increase of the tumor cells. The effects of oxaliplatin on the survival of the tumor and the survival of the patients with colorectal cancer are ultimately affected by colonization and reducing the rate of apoptosis. This study suggests that the RPA1 gene rs5030740 polymorphism can be used as a major potential biological indicator of the efficacy and prognosis of oxaliplatin in Chinese colorectal cancer population.
【学位授予单位】：南京中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R735.34

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