摘 要

目的：研究MDM2基因rs937283 位点与湖北人群胃癌和肺癌发生的关联。

方法：提取肺癌、胃癌患者以及正常人群的外周血细胞基因组DNA，采用限制性片段长度多态性聚合酶链反应（PCR-RFLP）技术检测550 例肺癌患者、460 例胃癌患者以及 800 例正常对照组MDM2基因rs937283 位点单核苷酸多态性。经过Hardy-weinberg 定律验证各群体均是遗传平衡群体，并且应用了用于多重测试的Bonferroni校正，具有代表性。结果运用 SPSS 软件进行统计学分析。

结果： rs937283 与肺癌的发病风险显著相关，年龄、性别、吸烟状况和饮酒状况等因素均会影响这种相关性。MDM2 rs937283 在肺癌组中等位基因分布分别为G(0.182)-A(0.818)；胃癌组G(0.175)-A(0.825)；正常对照组中G(0.136)-A(0.864)。MDM2 rs937283 在肺癌组中基因型频率分布分别为GG型0.043，AG型0.278，AA型0.679；胃癌组中GG型0.041，AG型0.267，AA型0.692；正常对照组中GG型0.022，AG型0.228，AA 型0.750。Bonferroni校正多重检验后（0.05/6=0.008），仍然发现肺癌患者rs937283 G等位基因频率（18.2%，P=0.001）显著高于正常对照组（13.6%），表明等位基因G与肺癌风险增加有关（OR=1.42,95%CI=1.15-1.75）。同时，rs937283基因型的logistic回归分析显示AG和GG基因型携带者比AA基因型携带者更倾向于患肺癌（P=0.004，OR=1.42,95%CI=1.12-1.81），并且该关联在Bonferroni纠正后仍然显著。然而，在Bonferroni校正的P值（lt;0.008,0.05/6）中发现rs937283与胃癌风险之间没有显著关联。

结论：我们的研究表明，MDM2 rs937283 Agt; G变异显着增加中国人群中肺癌和胃癌的风险。 进一步的荟萃分析显示MDM2 rs937283 Agt; G变异与癌症发展风险增加相关，特别是在中国人群中。 因此，MDM2 rs937283变异可能是中国癌症患者的一个有价值的危险因素或诊断性生物标志物，需要更多证据。

关键词：MDM2，rs937283，肺癌，胃癌，meta分析

Abstract

Objective: To find the association between the rs937283 site of MDM2 gene and the occurrence of gastric cancer and lung cancer in Hubei province.

Methods: PCR-RFLP was used to analyze the allelic and genotypic polymorphisms of MDM2 rs937283 in550 patients with lung cancer, 460 patients with gastric cancer and 800 normal controls. The test for deviation from Hardy-Weinberg expectations (HWE) showed that all populations are genetically balanced, and Bonferroni corrections for multiple tests are used. Statistical analysis was performed using the SPSS software package.

Results: The results showed that rs937283 was significantly associated with the risk of LC, and the factors of age, gender, smoking status and drinking status would affect such association. For the MDM2 rs937283 site, the allelic frequencies are G(0.182)-A（0.818） in lung cancer, G（0.175）-A（0.825） in gastric cancer, and G(0.136)-A(0.864) in normal controls ；And the genotypic frequencies are GG (0.043), AG (0.278), AA (0.679) in lung cancer, GG (0.041), AG (0.267), AA (0.692) in gastric cancer, and GG (0.022), AG (0.228), AA (0.750) in normal controls. After Bonferroni correction for multiple testing (0.05/6 = 0.008), it was still found that the frequencies of rs937283 G allele were significantly higher in LC patients (18.2%, P = 0.001) than in normal controls (13.6%), indicating allele G was associated with an increased risk of LC (OR = 1.42, 95%CI = 1.15-1.75). Concordantly, the logistic regression analysis for rs937283 genotype revealed that AG and GG genotypes carriers were more inclined to suffer from LC than AA genotype carriers (P = 0.004, OR = 1.42, 95%CI = 1.12-1.81), and the association remained significant after Bonferroni correction. However, no significant association was found between rs937283 and GC risk at the Bonferroni-corrected P value (lt; 0.008, 0.05/6).

Conclusion: In conclusion, rs937283 Agt;G variant significantly increases the risk of lung and gastric cancer in Chinese population. Further meta-analysis revealed that MDM2 rs937283 Agt;G variant is associated with increased risk of cancer development particularly in Chinese population. Therefore, the MDM2 rs937283 variant may be a valuable risk factor or diagnostic biomarker for Chinese cancer patients, which needs more supporting evidence.

Keywords：MDM2, rs937283, lung cancer, gastric cancer, meta-analysis

目 录

第一章 绪论 1

1.1肺癌和胃癌的现状 1

1.2胃癌和肺癌的发病因素 2

1.3癌症与TP53基因的联系 2

1.4癌症的遗传易感性研究 4

1.4.1 SNPs( single nucleotide polymorphisms ，单核苷酸多态性).................................4

1.4.2 SNPs 的研究方法 4

1.5 研究的目的及前景展望 5

第二章 材料和方法 6

2.1 实验器材 6

2.2 主要试剂 6

2.3 样本采集 7

2.4 样本 DNA 的提取 8

2.5 DNA 浓度及纯度检测 9

2.6 MDM2基因部分序列的扩增及 PCR-RFLP 法检测 SNPs 9

2.6.1 MDM2基因单核苷酸多态性的特点 9

2.6.2 PCR 引物及酶切错配引物的设计 9

2.6.3 PCR 反应条件 9

2.6.4 SNPs 的酶切情况 10

2.6.5 DNA跑胶结果 10

第三章 结果 12

3.1 等位基因频率及基因型分布结果 12

3.2 变量分层分析结果 13

第四章 讨论 22

第五章 结论 24

参考文献 25

致谢 27

第一章 绪论

癌症一直以来都是人类生命健康的冷血杀手，肺癌和胃癌更是世界上多年来被诊断为癌症死亡的主要癌症，并且其发病率仍然迅速增加^{[1, 2]}。大量的调查和研究显示，患者的生活环境，饮食，遗传以及慢性疾病等因素与癌症的发生有关。随着分子生物学和遗传学的进步，癌症的表观遗传学研究已成为近年来的热点，但由于缺乏理想的早期诊断肺癌和胃癌的遗传生物标志物，大多数患者已进入临床中晚期阶段，并且错过了最佳治疗期^{[3, 4]}。 因此，鉴定与肺癌和胃癌易感性相关的遗传变异将有助于进行早期诊断和风险预测。已有研究表明，MDM2与MDMX（MDM4)发挥类似抑癌蛋白P53泛素化降解E3作用^[5]。因此，MDM2癌基因的改变对于阐明肿瘤的发生，发展机制和转移机制具有重要意义，对临床实践也具有指导意义^[6]。本实验旨在研究MDM2基因rs937283位点与湖北人群胃癌和肺癌发生的关联，为人类胃癌和肺癌的诊断与防治提供理论基础。

1.1肺癌和胃癌的现状