肿瘤遗传标记-DNA修复与肿瘤发病风险的研究ppt课件

1、肿瘤遗传标志-DNA修复与肿瘤发病风险的研讨 魏庆义, M.D., Ph.D. 流行病系美国休斯顿德州医学中心MD Anderson Cancer Center肿瘤死亡病因电离辐射 3%Tobacco 30%酗酒 5%成年人膳食和肥胖症 30% 其他 7%免疫要素 5%缺乏运动 5%遗传要素 5%病毒感染 5% 产前要素和生长要素 5%Harvard School of Public Health, 1998 根本数据: 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, 2021遗传要素一定在起作用!Pe

2、to J, Nature, 2001吸烟在不同人群中有不同危害中国美国Cell CycleArrestDNA RepairCancer ApoptosisTranscription Dependent ApoptosisTranscription IndependentApoptosis DNA损伤-修复与肿瘤发生的机制 p53 ProteinAccumulation DNADAMAGEAltered ExpressionBAX, Fas, Bcl2PIG3 Binding to Transcription Replication-Repair FactorsTFIIH (XPB, XPD)

3、and p62 binds to p53PCNA (p21WAF1 and GADD45)Increased Expression p21WAF1, MDM2, cyclin G, and GADD45Modified from Harris, 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, hCHK2 Li-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

4、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DNA ligase (LIG3),单碱基损伤DNA glycosylase (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

5、/ERCC5, ERCC1, LIG1, 紫外线、吸烟 CSB/ERCC6,CSA/CKN1, XAB2, TFIIH (XPB/ERCC3 XPD/ERCC2, GTF2H1, GTF2H21, 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 .Wood et al, Science, 2001重组修

6、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, XRCC2 XRCC3, XRCC4, XRCC5, XRCC6 XRCC7, XRCC8, BRCA1, BRCA2 .XP病人和正常人中皮肤癌发生的年龄分布Kraemer, PNAS, 1997Skin cancers in normal populationSkin cancers in XP populationXP = xeroderma pigment

7、osum变量病例 对照OR (95% CI) P value No. (%) No. (%)总数 895 898年龄 0.493 55 439 (49.1) 455 (50.7) 55 456 (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 (1.091.78) 曾经239 (2

8、6.7)441 (49.1) 1.00 未延续 365 (40.8)147 (16.4) 3.97 (3.085.13)喝酒 0.0001 从无 235 (26.3)398 (44.3) 1.00 曾经206 (23.0)149 (16.6) 2.12 (1.602.81) 未延续 454 (50.7)351 (39.1) 2.10 (1.682.63)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-对照研讨中部分变量的频数分布2000-2006 *Differences

9、between cases and controls were analyzed by two-sided 2 and Students t tests 实验室血样处置流程PHAPHASpinWhole blood short-term culture 1ml 1ml 1mlMutagen sensitivityassayBPDE ControlBPDEGamma 1mlBPDE-Induce DNA adducts assayDNAextraction 1mlRT- PCR forgene expressionLong-termstoragecDNADNA 1mlRNAextractionG

10、enotypingLymphocyte isolation(frozen)CAT/LucassaysDNA repaircapacity2 mleachTransfectionHeparinized, 10-30 ml Sample collection(cases and controls)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Benzoapyrene吸烟修复复合体衔接POL/, ligasePCNA, RFC RPA NER Core ProteinsERCC1XPAXPB/ERCC3XPCXPD/ERCC2X

11、PE/DDB1/2XPE/ERCC4XPG/ERCC5Neumann et al., Mol Carcino, 2005Li et al., Int J Cancer. 2001BPDE-诱导DNA加成物的检测头颈部肿瘤研讨Cancer Res 2007; 67: (12). June 15, 2007变量 病例 (n = 803) 对照 (n = 839)P or OR (95% CI)* n (%) n (%)总数加成物中位数 29.22 / 107 508 (63.3) 419 (50.0) 1.71 (1.392.10)白人加成物中位数 29.22 / 107 431 (64.7) 3

12、38 (51.2) 1.81 (1.452.27)*Adjusted for the matching variables: age (in years), sex, ethnicity, smoking and alcohol use.染色体对DNA损伤剂的敏感性测定变量 病例 (n = 895) 对照 (n = 898)P or OR (95% CI)* n (%) n (%)总数中位数 0.43 548 (61.2) 457 (50.9) 1.57 (1.291.90)白人中位数 0.43 465 (60.6) 381 (49.9) 1.56 (1.271.91)*Adjusted fo

13、r the matching variables: age (in years), sex, ethnicity, smoking and alcohol use.Cancer Res 2021; 68: (11). June 1, 2021 宿主报告基因的DNA损伤修复测定4863 bp ApEnhPLucBgl II Xbal IPvu I BamH I Nar I Bgl II pCMVlucpCMVcat5000 bp ApEnhPcatHind IIIXbal IEcoR I EcoR I BPDEBPDEQiao et al., Mutat Res, 2002Cases = 803

14、Controls = 839 趋势检验 : P 0.001 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 High LowLi et al., Can Res, 2007病例 = 721对照 = 721 趋势检验: P 0.0001修复功能Cases = 767Controls = 763 趋势检验: P = 0.0001染色体敏感性DNA加成物Wang et al.,CCR, 2021Wang et al., Cancer Res, 2007反式蛋白质芯片测定修复蛋白程度Wei et al., CEBP, 200557病例，63对照0.0 1.0 2.0 3.0

15、 4.0 5.0 6.0 7.0 相对比数相对表达程度:ERCC1XPAXPCXPD/ERCC2XPF/ERCC4XPG/ERCC5(compared to beta-actin)NER修复蛋白程度与头颈部肿瘤发病风险病例 = 57对照 = 63Wei et al., CEBP, 2005Gene rs no.Allele 1Allele 2CodonWtVariantVar freERCC1 None ERCC2 1799793GA 312ASP(D) ASN(N) 0.24 1052559AC 751LYS(K) GLN(Q) 0.22ERCC3 NoneERCC4 2020955TC 6

16、62SER(S) PRO(P) 0.06ERCC5 17655GC1104ASP(D) HIS(H) 0.38ERCC6 2228528GA 399GLY(G) ASP(D) 0.22 2228526AG1097MET(M) VAL(V) 0.18 2228527AG1213ARG(R) GLY(G) 0.19 4253211GC1230ARG(R) PRO(P) 0.08 2228529AG1413GLN(Q) ARG(R) 0.19XPA NoneXPC 2228000CT 499ALA(A) VAL(V) 0.24 2228001AC 939LYS(K) GLN(Q) 0.34多碱基剪切

17、修复基因多态位点及对其功能的影响(nsSNPs in NER genes)多碱基剪切修复基因多态位点与头颈部肿瘤发病风险NER - SNPs 829 / 854 OR (95% CI)ERCC1C8092A AA vs CC+CA 0.9 (0.6-1.4)XPAG23A AA vs GG+AG 0.9 (0.7-1.2)XPCAla499Val Val/Val vs Ala/Ala+Ala/Val 1.7 (1.2-2.4)Lys939Gln Gln/Gln vs Lys/Lys+Lys/Gln 1.1 (0.8-1.4)ERCC2/XPD Asp312Asn Asn/Asn vs Asp/

18、Asp+Asp/Asn 1.2 (0.9-1.7)Lys751Gln Gln/Gln vs Lys/Lys+Gln/Gln 1.1 (0.8-1.4)XPG/ERCC5His1104Asp Asp/Asp vs His/His+His/Asp 0.8 (0.5-1.3)All SNPs combined asNo. of variant homozygotes no. (%) no. (%) OR (95% CI)0-2 114 (13.8) 140 (16.3) 1.03 453 (54.6) 477 (55.9) 1.2 (0.9-1.6)4 194 (23.4) 177 (20.7) 1.4 (1.0-1.9)5 53 (6.4) 55 (6.4) 1.2 (0.8-2.0)6 15 (1.8) 5 (0.6) 4.1 (1.4-12.0)P for trend 0.017Gene Comparison Cases/Controls Risk* Adjusted for age, sex, smoking and alcohol use (An et al., CEBP, 2007)结论需求有高