【作者】 温晖；

【导师】 丁强； 夏国伟； 方祖军；

【作者基本信息】 复旦大学 ， 外科学， 2009， 博士

【摘要】 第一部分DNA修复基因单核苷酸多态与膀胱尿路上皮癌遗传易感性研究目的：DNA修复机制在防止基因突变，维持基因组稳定性过程中起着重要的作用。DNA修复基因多态是个体间DNA修复能力差异的重要原因。初步探讨在上海地区汉族人群中DNA修复基因(XPC Lys939Gln，XPG His1104Asp和XRCC1 Arg399Gln)单核苷酸多态性与膀胱尿路上皮癌(移行细胞癌)遗传易感性的关系方法：我们采用病例一对照研究方法，以Taqman探针实时荧光定量PCR技术和DNA测序等方法对130例病理证实原发膀胱尿路上皮癌患者和304例非肿瘤对照的三个DNA修复基因(XPC Lvs939Gln，XPG His1104Asp and XRCC1 Arg399Gln)的单核苷酸多态性位点进行检测。应用非条件Logistic回归模型，调整混杂因素后，分析各基因型与膀胱癌罹患风险的关系，还分析了基因-基因、基因-环境相互作用与膀胱癌风险的关系。结果：调整年龄、性别和吸烟因素后，XPC基因Lys939Gln多态与膀胱癌风险增高相关。与携带XPC 939 Lys／Lys基因型者比较，Lys／Gln和Gln／Gln基因型者(携带Gln突变等位基因者)罹患膀胱癌的风险增加到2.8倍(校正OR=2.80；95％CI=1.79-4.37)。按年龄分层后，携带Gln突变等位基因者(XPC939Lys／Gln杂合基因型和939Gln／Gln基因型)在≥64岁的研究对象中罹患膀胱癌的风险增加4.17倍(校正OR=4.17；95％CI=2.14-8.16)。再按性别分层分析，与Lys／Lys基因型比较，发现携带Gln突变等位基因的男性发生膀胱癌的风险增加2.95倍(校正OR=2.95；95％CI=1.43-6.08)。在重度吸烟人群中(≥25包-年)，携带至少一个Gln等位基因者(Lys／Gln基因型+Gln／Gln基因型)罹患膀胱癌的风险为携带Lys／Lys基因型者的5.54倍(95％CI=1.86-16.47)。肿瘤组中，XPC 939Gln／Gln和Lys／Gln基因型频率在肌层浸润性膀胱癌患者和肿瘤多发者中明显高于非肌层浸润性膀胱癌患者和肿瘤单发患者(P＜0.05)。与携带XPG1104Asp／Asp基因型个体比较，携带His突变等位基因者(携带1104Asp／His基因型个体和携带1104His／His基因型个体)罹患膀胱癌的风险增高到2.62倍(校正OR=2.62；95％CI=1.61-4.24)。年龄不影响与XPG Asp1104His多态相关的膀胱癌风险。再按性别分层分析，与Asp／Asp基因型比较，发现携带His／His纯合突变基因型的女性发生膀胱癌的风险增高到2.48倍(OR=2.48；95％CI=1.34-4.60)。在不吸烟人群中，携带His突变等位基因(His／His+Asp／His基因型)者罹患膀胱癌的风险为携带Asp／Asp基因型者的4.31倍(95％CI=1.19-12.47)。在肿瘤组中，XPG Asp1104His多态与肿瘤的临床病理特征无关。XRCC1基因Arg399Gln多态与膀胱癌风险无关联。与野生型基因型比较，突变杂合子和纯合子的ORs(95％CIs)分别为0.70(0.43-1.13)，1.00(0.65-1.55)。吸烟可以增加膀胱癌的风险，其OR(95％CI)值为3.34(2.01-5.57)。在重度吸烟人群中(≥25包-年)，携带至少一个XPC939Gln等位基因者(Lys／Gln基因型+Gln／Gln基因型)罹患膀胱癌的风险为携带Lys／Lys基因型者的5.54倍(95％CI=1.86-16.47)。以携带XPC 939Gln突变等位基因、XPG 1104His突变等位基因和XRCC1399Gln突变等位基因为高风险型。调整混杂因素后，与不携带风险型的个体相比，携带高风险基因型数量为2个和3个位点时，膀胱癌的患病风险分别增高到1.90倍(95％CI：1.13-4.12)和3.59倍(95％CI：1.82-9.26)并且该风险增高效应有统计学意义。结论：本次以上海地区汉族人群为研究对象的病例对照研究，主要的研究结果有：吸烟可增加膀胱尿路上皮癌的患病风险；XPC基因Lys939Gln和XPG基因Asp1104His的单核苷酸多态与膀胱尿路上皮癌的发病风险有关联；重度吸烟和XPC基因939Gln的交互作用可增加膀胱尿路上皮癌的患病风险；DNA修复体系多个(＞2)高风险型SNP的积累可使膀胱尿路上皮癌的患病风险增高。第二部分毒物代谢酶基因单核苷酸多态与膀胱尿路上皮癌遗传易感性研究目的：参与环境化学物代谢的多种代谢酶具有遗传多态性，对于环境化学物质引起健康损害的个体易感性有重要的影响作用。在不同的种族和地域人群间多态性人群频率特征往往存在显著差异。初步探讨在上海地区汉族人群中Ⅰ相代谢酶基因(CYP2C9~*3，CYP2C19~*2，CYP2C19~*3，CYP2D6~*10，CYP3A4~*18和CYP3A5~*3)单核苷酸多态性与膀胱尿路上皮癌(移行细胞癌)遗传易感性的关系方法：我们采用病例一对照研究方法，以Taqman探针实时荧光定量PCR技术对130例病理证实原发膀胱尿路上皮癌患者和304例非肿瘤对照的6个Ⅰ相代谢酶基因基因(CYP2C9~*3，CYP2C19~*2，CYP2C19~*3，CYP2D6~*10，CYP3A4~*18和CYP3A5~*3)的单核苷酸多态性位点进行检测。应用非条件Logistic回归模型，调整混杂因素后，分析各基因型与膀胱尿路上皮癌罹患风险的关系，还分析了基因-基因、基因-环境相互作用与膀胱尿路上皮癌风险的关系。结果：这六个代谢酶基因单核苷酸多态各种基因型在正常对照及病例组中的分布均符合Hardy-Weinberg平衡定律。调整性别、年龄和吸烟情况后，无论是CYP2C9~*3，CYP2C19~*2，CYP2D6~*10，CYP3A4~*18和CYP3A5~*3的突变杂合基因型还是突变纯合基因型，均未发现与膀胱尿路上皮癌的发病风险有关联(调整OR值95％CI分别为突变杂合体1.02[0.61-1.68]，0.85[0.47-1.54]，1.09[0.67-1.77]，1.12[0.67-1.86]和0.58[0.24-1.38]；突变纯合体1.02[0.63-1.63]，0.98[0.65-1.59]，1.50[1.00-2.25]，0.92[0.54-1.57]和0.96[0.65-1.40])。CYP2C19~*3的突变等位基因频率也和膀胱尿路上皮癌风险无关(调整OR：1.01；95％CI=0.42-2.45)。同时也未观察到这六个单核苷酸多态性与吸烟交互作用对膀胱尿路上皮癌罹患风险的阳性结果。基因-基因联合分析显示：病例组中的所用研究对象中至少携带1个高风险型；两组中未见有携带6个高风险型者。所以用携带1个高风险型的数据作为参照，调整混杂因素后，随着携带高风险基因型数量逐渐增多时，并未发现膀胱尿路上皮癌的患病风险随之增高。结论：本次以上海地区汉族人群为研究对象的病例对照研究结果：代谢酶基因CYP2C9~*3，CYP2C19~*2，CYP2C19~*3，CYP2D6~*10，CYP3A4~*18和CYP3A5~*3的单核苷酸多态性与膀胱尿路上皮癌的发病风险无关。第三部分单核苷酸多态与膀胱尿路上皮癌遗传易感性：多位点联合分析目的：肿瘤是一类复杂性疾病，它的发生与众多低外显的肿瘤遗传易感基因之间的相互作用以及基因-环境之间的交互作用有关。本研究以130例膀胱尿路上皮癌患者和304例非肿瘤对照为研究对象，选择了9个DNA损伤修复和致癌物代谢相关的功能性基因单核苷酸多态，包括XPCLys939Gln，XPG His1104Asp，XRCC1Arg399Gln，CYP2C9~*3，CYP2C19~*2，CYP2C19~*3，CYP2D6~*10，CYP3A4~*18和CYP3A5~*3。方法：在第一章和第二章以非条件logistic回归方法分别分析了这9个SNPs单个位点与肿瘤风险的相关性以及基因和环境之间的交互作用。为了进一步了解这9个位点之间的相互作用以及具体是哪几个位点有交互作用，我们采用用多元缩减法(multifactor-dimensionality reduction，MDR)分析这9个单核苷酸多态之间的相互作用关系。结果：本研究候选的9个基因多态中，从两点模型看，XPC Lys939Gln和XPGAsp1104His多态之间的存在交互作用，其相互验证的一致性(cross-validationconsistency，CVC)达到了10次，从三点模型看，XPC Lys939Gln，CYP2D6~*10和CYP3A5~*3基因多态之间的联合效应最显著，CVC达到了8；四点模型得出XPC、XPG、CYP2D6~*10和CYP3A5~*3多态与膀胱癌发病存在联合作用，其CVC6，Permutation检验P均小于0.05。在大于4个点的交互作用模型中，没有得出有显著性差异的结果。结论：我们的研究表明，膀胱尿路上皮癌是一个多基因参与的复杂疾病；多元缩减法多位点联合分析为肿瘤易感基因的研究提供了有用的方法。更多还原

【Abstract】 Background & Objective: DNA repair has an essential role in protecting the genomefrom damage by endogenous and environmental agents. Polymorphisms in DNArepair genes may be associated with difference in the repair capacity and mayinfluence an individual’s risk of cancer. We preliminarily investigated the associationbetween bladder transitional cell carcinoma(TCC) and single nucleotidepolymorphisms(SNPs) of DNA repair genes(XPC Lys939Gln, XPG His1104Asp andXRCC1 Arg399Gln)among Han-Chinese in Shanghai.Methods: We therefore conducted a case-control study to assess the relationshipbetween single nucleotide polymorphisms in DNA repair genes XPC, XPG andXRCC 1 and Bladder Cancer risk. and we also attempted to assess gene-gene andgene-environmental interactions. A total of 130 primary bladder transitional cellcarcinoma cases and 304 cancer-free controls were recruited and frequency-matched on age and gender status. Genotypes were determined by TaqMan Probe-basedpolymerase chain reaction and DNA sequencing techniques. Adjusted odds ratios(ORs) and their 95% confidence intervals(CIs) were computed to estimate the risk ofbladder transitional cell carcinoma for each genotype. All statistical tests weretwo-side tests.Results: We found that polymorphisms in XPC and XPG but not XRCC1 wereassociated with risk of bladder transitional cell carcinoma.The individuals with bladder transitional cell carcinoma at least having one XPC939Gln had an adjusted OR of 2.80(95%CI=1.79-4.37) compared with noncarriers.Stratification analysis revealed that the increased risk was mainly confined to malewith the adjusted OR of 2.95(95%CI=1.43-6.08) and older individuals(age atdiagnosis≥64 years) with the adjusted OR of 4.17(95%CI=2.14-8.16). In the casegroup, XPC Lys939Gln polymorphisms was associated with tumor stage and tumornumber.Compared with XPG1104Asp/Asp genotype, the His/His and Asp/His genotypewas significantly associated with bladder cancer(adjusted OR=2.62; 95% CI=1.61-4.24). The risk of the(1104His/His+Asp/His)genotype appeared to be morepronounced in females(adjusted OR=2.48;95%CI=1.34-4.60) and in subjects withbladder transitional cell carcinoma who were non-smokers(adjusted OR=4.31; 95%CI=1.19-12.47). In the case group, XPG His1104Asp polymorphisms was notassociated with tumor clinicopathological features. Neither the heterozygote nor thehomozygote variants of XRCC1 Arg399Gln polymorphisms were associated withincreased bladder cancer risk.Smoking increased the risk of developing bladder transitional cell carcinoma,having an adjusted OR of 3.34(95% CI=2.01-5.57). The subjects with heavysmoking and at least having one XPC 939Gln allele had an adjusted OR of 5.54(95%CI=1.86-16.47), compared with noncarriers and heavy smoking,As compared with individuals having no putative high-risk genotypes,individuals with three or more putative high-risk genotypes had a statistical increasedrisk of bladder transitional cell carcinoma. And the adjusted ORs(95%CIs) were1.90(1.13-4.12) for individuals with two high-risk genotypes, 3.59(1.82-9.26) forindividuals with three high-risk genotypes, respectively.Conclusion: The main findings from this case-control study carried out inHan-Chinese in Shanghai were as follows: Smoking may contribute to the bladder transitional cell carcinoma predisposition. DNA repair genes XPC Lys939Gln andXPG Asp1104His single nucleotide polymorphisms may play an important role in thedevelopment of bladder transitional cell carcinoma. The gene-environment interactionbetween XPC Lys939Gln polymorphism and cigarette smoke exposure may also haveeffect on the bladder carcinogenesis. The accumulation of multi-polymorphisms inDNA repair genes has a contribution to the genetic predisposition of bladdertransitional cell carcinoma. Background & Objective: Many carcinogen-metabolizing enzyme genes displaywide ranges of genetic polymorphism. The genetic polymorphism of these enzymesmodulate individual susceptibility to certain disease. There are notable deviations onthe population frequency of carcinogen-metabolizing gene polymorphism amongdifferent ethnic groups and regional resident population. We preliminarilyinvestigated the association between bladder transitional cell carcinoma(TCC) andsingle nucleotide polymorphisms(SNPs) of carcinogen-metabolizing enzymegenes(CYP2C9~*3, CYP2C19~*2, CYP2C19~*3, CYP2D6~*10, CYP3A4~*18 andCYP3A5~*3)among Han-Chinese in Shanghai.Methods: We therefore conducted a case-control study to assess the relationshipbetween single nucleotide polymorphisms in xenobiotic metabolism genesCYP2C9~*3, CYP2C19~*2, CYP2C19~*3, CYP2D6~*10, CYP3A4~*18 and CYP3A5~*3and Bladder Cancer risk. and we also attempted to assess gene-gene andgene-environmental interactions. A total of 130 primary bladder transitional cellcarcinoma cases and 304 cancer-free controls were recruited and frequency-matchedon age and gender status. Genotypes were determined by TaqMan Probe-basedpolymerase chain reaction. Adjusted odds ratios(ORs) and their 95% confidenceintervals(CIs) were computed to estimate the risk of bladder transitional cellcarcinoma for each genotype. All statistical tests were two-side tests.Results: The distribution of genotypes of these CYP polymorphisms geneticvariants in cases and controls was consistent with Hardy-Weinberg equilibrium. Usinglogistic regression adjusting for smoking, gender and age, neither the heterozygotenor the homozygote variants of CYP2C9~*3, CYP2C19~*2,, CYP2D6~*10, CYP3A4~*18and CYP3A5~*3 polymorphisms were associated with increased bladder cancer risk(adjusted odds ratio [95% confidence interval] for heterozygote 1.02[0.61-1.68],0.85[0.47-1.54], 1.09[0.67-1.77], 1.12[0.67-1.86] and 0.58[0.24-1.38], respectivelyand homozygote variant, 1.02[0.63-1.63], 0.98[0.65-1.59], 1.50[1.00-2.25],0.92[0.54-1.57] and 0.96[0.65-1.40]).The CYP2C19~*3 polymorphisms was notassociated with bladder cancer risk either(adjusted OR:1.01; 95% CI=0.42-2.45).Moreover, we did not find any significant interaction between these CYPpolymorphisms and environmental exposure to cigarette smoking. As compared with individuals having one putative high-risk genotypes,individuals with two or more putative high-risk genotypes had not a statisticalincreased risk of bladder transitional cell carcinoma.Conclusion: The main findings from this case-control study carried out inHan-Chinese in Shanghai were as follows: the single nucleotide polymorphisms(SNPs) of CYP2C9~*3, CYP2C19~*2, CYP2C19~*3, CYP2D6~*10, CYP3A4~*18 andCYP3A5~*3 may not influence the risk of bladder transitional cell carcinoma. Background & Objective: Accumulating evidence shows that most cancers arecaused by interactions of many low-penetrance susceptibility genes. In this study, weinvestigated the association between risk of bladder transitional cell carcinoma andsingle nucleotide polymorphisms(SNPs) in multiple loci in a case-control study consisted of 130 patients and 304 cancer-free controls.Methods: The functional SNPs investigated were XPC Lys939Gln, XPGHis1104Asp, XRCC1Arg399Gln, CYP2C9~*3, CYP2C19~*2, CYP2C19~*3,CYP2D6~*10, CYP3A4~*18 and CYP3A5~*3. Traditional unconditional logisticregression model was used to evaluate the effect of single SNP alone in chapter oneand chapter two. In this chapter, multifactor-dimensionality reduction(MDR) wasapplied to examine the interactions between these 9 SNPs interactions.Results: It was found that, among these SNPs, MDR analysis revealed that the XPCLys939Gln and XPG His1104Asp SNPs displayed significant interaction in risk of thebladder cancer. Moreover, two sets of polymorphisms were found to nteract, whichincreased the risk of bladder cancer.Conclusion: Our results support the hypothesis that bladder cancer is a complexdisease resulting from the interactions of low-penetrance genes and MDR seems to bea useful analytical tool for complex genetic diseases.更多还原