肿瘤遗传标记-DNA修复与肿瘤发病风险的研究-NoSl

肿瘤遗传标记-DNA修复与肿瘤发病风险的研究魏庆义,M.D.,Ph.D.流行病系美国休斯顿德州医学中心MDAndersonCancerCenter肿瘤死亡病因电离辐射3%Tobacco30%酗酒5%成年人膳食和肥胖症30%其他7%免疫因素5%缺乏运动5%遗传因素5%病毒感染5%产前因素和生长因素5%HarvardSchoolofPublicHealth,1998基本数据:百万美国人口:300.0吸烟者:46.5每年癌症患者1.3每年肺癌患者0.5每年头颈部癌患者0.04吸烟与肿瘤ACS,2009遗传因素一定在起作用!PetoJ,Nature,2001吸烟在不同人群中有不同危害Heartdisease9%Strok9%TB5%Stomachcancer9%Livercancer11%Otherrespiratory27%Esophagealcancer8%Lungcancer18%Othercancer4%Heartdisease36%Strok5%Stomach+liver+TB1%Otherrespiratory12%Esophagealcancer2%Lungcancer39%Othercancer5%中国美国CellCycleArrestDNARepairCancerApoptosisTranscriptionDependentApoptosisTranscriptionIndependentApoptosisDNA损伤-修复与肿瘤发生的机制p53ProteinAccumulationDNADAMAGEAlteredExpressionBAX,Fas,Bcl2PIG3BindingtoTranscriptionReplication-RepairFactors•TFIIH(XPB,XPD)andp62bindstop53•PCNA(p21WAF1andGADD45)IncreasedExpressionp21WAF1,MDM2,cyclinG,andGADD45ModifiedfromHarris,1994DNA修复基因和肿瘤发生基因肿瘤XP(A)皮肤癌XP(B)皮肤癌XP(C)皮肤癌XP(D)皮肤癌XP(E-G)皮肤癌hMLH1结肠癌hPSM1/2结肠癌hMSH2结肠癌hMSH3结肠癌hMSH6结肠癌pRB视网膜母细胞瘤P53,hCHK2Li-Fraumeni综合症P16黑色素瘤ATM乳腺癌BRCA1/2乳腺癌…正常修复功能修复功能缺陷DNA修复的特定通路DNA断裂重组（HR）和非重组修复（NHEJ）氧化损伤多碱基剪切修复(BER)DNA加成物多碱基剪切修复(NER)铰链物NER,HR&NHEJ的综合修复人类DNA修复通路修复类型基因种类损伤种类单碱基剪切修复DNAligase(LIG3),单碱基损伤DNAglycosylase(MBD4,MPG,MYH,NTH1,OGG1,SMUG1,TDG,UNG),APE1,APE2,XRCC1,ADPRT,ADPRTL2,ADPRTL3...多碱基剪切修复XPA,XPC,XPE,XPF/ERCC4,多碱基损伤XPG/ERCC5,ERCC1,LIG1,（紫外线、吸烟）CSB/ERCC6,CSA/CKN1,XAB2,TFIIH(XPB/ERCC3XPD/ERCC2,GTF2H1,GTF2H2\1,GTF2H3,GTF2H4,CDK7,CCNH,MNAT),DDB1,DDB2,MMS19,CENN2,AD23A,RAD23B,RPA1,RPA2,RPA3...碱基错配修复MSH2,MHS3,MSH6,MSH4,碱基错配MSH5,MLH1,MLH3,PMS1,PMS2,PMS2L3,PMS2L4...Woodetal,Science,2001重组修复RAD50,RAD51,RAD51B,RAD51C,DNA双链断裂RAD51D,RAD54L,RAD54B,V(D)J重组RAD52,DMC1,MRE11A,NBS1,ERCC1,XPF/ERCC4,XRCC2XRCC3,XRCC4,XRCC5,XRCC6XRCC7,XRCC8,BRCA1,BRCA2...XP病人和正常人中皮肤癌发生的年龄分布Kraemer,PNAS,1997SkincancersinnormalpopulationSkincancersinXPpopulationXP=xerodermapigmentosum变量病例对照OR(95%CI)PvalueNo.(%)No.(%)总数895898年龄0.493≤55439(49.1)455(50.7)55456(50.9)443(49.3)性别0.231男674(75.3)654(72.8)女221(24.7)244(27.2)种族0.859白人767(85.7)763(85.0)西裔69(7.7)70(7.8)非裔59(6.6)65(7.2)吸烟0.0001从无291(32.5)310(34.5)1.39(1.09–1.78)曾经239(26.7)441(49.1)1.00未间断365(40.8)147(16.4)3.97(3.08–5.13)喝酒0.0001从无235(26.3)398(44.3)1.00曾经206(23.0)149(16.6)2.12(1.60–2.81)未间断454(50.7)351(39.1)2.10(1.68–2.63)家族史0.748有516(57.7)511(56.9)1.00无‡379(42.3)387(43.1)1.01(0.83-1.23)头颈部肿瘤病例-对照研究中部分变量的频数分布2000-2006*Differencesbetweencasesandcontrolswereanalyzedbytwo-sided2andStudent’sttests实验室血样处理流程Wholebloodshort-termculture1ml1ml1mlMutagensensitivityassayBPDEControlBPDEGamma1mlBPDE-InduceDNAadductsassayDNAextraction1mlRT-PCRforgeneexpressionLong-termstoragecDNADNA1mlRNAextractionGenotypingLymphocyteisolation(frozen)CAT/LucassaysDNArepaircapacity2mleachHeparinized,10-30mlSamplecollection(casesandcontrols)1mlPlasmaSerum烟草致癌物诱导的DNA损伤与修复机制OHBPDEOHO过氧化酶DNA加成物DNA恢复正常Benzo[a]pyrene吸烟修复复合体连接POL/,ligasePCNA,RFCRPANERCoreProteinsERCC1XPAXPB/ERCC3XPCXPD/ERCC2XPE/DDB1/2XPE/ERCC4XPG/ERCC5Neumannetal.,MolCarcino,2005Lietal.,IntJCancer.2001BPDE-诱导DNA加成物的检测头颈部肿瘤研究CancerRes2007;67:(12).June15,2007变量病例(n=803)对照(n=839)PorOR(95%CI)*n(%)n(%)总数加成物中位数0.0001≤29.22/107295(36.7)420(50.0)1.0029.22/107508(63.3)419(50.0)1.71(1.39–2.10)白人加成物中位数0.0001≤29.22/107235(35.3)355(48.8)1.0029.22/107431(64.7)338(51.2)1.81(1.45–2.27)*Adjustedforthematchingvariables:age(inyears),sex,ethnicity,smokingandalcoholuse.染色体对DNA损伤剂的敏感性测定变量病例(n=895)对照(n=898)PorOR(95%CI)*n(%)n(%)总数中位数0.0001≤0.43347(38.8)441(49.1)1.000.43548(61.2)457(50.9)1.57(1.29–1.90)白人中位数0.0001≤0.43302(39.4)382(50.1)1.000.43465(60.6)381(49.9)1.56(1.27–1.91)*Adjustedforthematchingvariables:age(inyears),sex,ethnicity,smokingandalcoholuse.CancerRes2008;68:(11).June1,2008宿主报告基因的DNA损伤修复测定4863bpApEnhPLucBglIIXbalIPvuIBamHINarIBglIIpCMVlucpCMVcat5000bpApEnhPcatHindIIIXbalIEcoRIEcoRIBPDEBPDEQiaoetal.,MutatRes,20020123Ist2nd3rd4thCases=803Controls=839趋势检验:P0.001调整相对比数DRC(%)四分位数个体DNA修复功能与头颈部肿瘤发病分险HighLowLietal.,CanRes,20070123Ist2nd3rd4th0123Ist2nd3rd4th病例=721对照=721趋势检验:P0.0001修复功能Cases=767Controls=763趋势检验:P=0.0001染色体敏感性DNA加成物Wangetal.,CCR,2009Wangetal.,CancerRes,2007反式蛋白质芯片测定修复蛋白水平Weietal.,CEBP,200557病例，63对照0.01.02.03.04.05.06.07.0相对比数相对表达水平:ERCC1XPAXPCXPD/ERCC2XPF/ERCC4XPG/ERCC5(comparedtobeta-actin)NER修复蛋白水平与头颈部肿瘤发病风险病例=57对照=63Weietal.,CEBP,2005Genersno.Allele1Allele2CodonWtVariantVarfreERCC1NoneERCC21799793GA312ASP(D)ASN(N)0.241052559AC751LYS(K)GLN(Q)0.22ERCC3NoneERCC42020955TC662SER(S)PRO(P)0.06ERCC517655GC1104ASP(D)HIS(H)0.38ERCC62228528GA399GLY(G)ASP(D)0.222228526AG1097MET(M)VAL(V)0.182228527AG1213ARG(R)GLY(G)0.194253211GC1230ARG(R)PRO(P)0.082228529AG1413GLN(Q)ARG(R)0.19XPANoneXPC2228000CT499ALA(A)VAL(V)0.242228001AC939LYS(K)GLN(Q)0.34多碱基剪切修复基因多态位点及对其功能的影响(nsSNPsinNERgenes)多碱基剪切修复基因多态位点与头颈部肿瘤发病风险NER-SNPs829/854OR(95%CI)ERCC1C8092AAAvsCC+CA0.9(0.6-1.4)XPAG23AAAvsGG+AG0.9(0.7-1.2)XPCAla499ValVal/ValvsAla/Ala+Ala/Val1.7(1.2-2.4)Lys939GlnGln/GlnvsLys/Lys+Lys/Gln1.1(0.8-1.4)ERCC2/XPDAsp312AsnAsn/AsnvsAsp/Asp+Asp/Asn1.2(0.9-1.7)Lys751GlnGln/GlnvsLys/Lys+Gln/Gln1.1(0.8-1.4)XPG/ERCC5His1104AspAsp/Asp