腫瘤遺傳標(biāo)記-DNA修復(fù)與腫瘤發(fā)病風(fēng)險(xiǎn)的研究ppt課件

1、腫瘤遺傳標(biāo)志-DNA修復(fù)與腫瘤發(fā)病風(fēng)險(xiǎn)的研討 魏慶義, M.D., Ph.D. 流行病系美國(guó)休斯頓德州醫(yī)學(xué)中心MD Anderson Cancer Center腫瘤死亡病因電離輻射 3%Tobacco 30%酗酒 5%成年人膳食和肥胖癥 30% 其他 7%免疫要素 5%缺乏運(yùn)動(dòng) 5%遺傳要素 5%病毒感染 5% 產(chǎn)前要素和生長(zhǎng)要素 5%Harvard School of Public Health, 1998 根本數(shù)據(jù): 百萬(wàn) 美國(guó)人口: 300.0 吸煙者: 46.5 每年癌癥患者 1.3 每年肺癌患者 0.5 每年頭頸部癌患者 0.04吸煙與腫瘤ACS, 2021遺傳要素一定在起作用!Pe

2、to J, Nature, 2001吸煙在不同人群中有不同危害中國(guó)美國(guó)Cell CycleArrestDNA RepairCancer ApoptosisTranscription Dependent ApoptosisTranscription IndependentApoptosis DNA損傷-修復(fù)與腫瘤發(fā)生的機(jī)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 修復(fù)基因和腫瘤發(fā)生 基因 腫瘤XP(A)皮膚癌 XP(B)皮膚癌XP(C)皮膚癌XP(D)皮膚癌XP(E-G)皮膚癌 hMLH1結(jié)腸癌hPSM1/2結(jié)腸癌hMSH2結(jié)腸癌hMSH3結(jié)腸癌hMSH6結(jié)腸癌 pRB 視網(wǎng)膜母細(xì)胞瘤P53, hCHK2 Li-Fraumeni綜合癥P16黑色素瘤ATM乳腺癌 BRCA1/2乳腺癌正常修復(fù)功能修

4、復(fù)功能缺陷DNA修復(fù)的特定通路DNA斷裂 重組HR和 非重組修復(fù) NHEJ 氧化損傷多堿基剪切修復(fù)(BER) DNA加成物多堿基剪切修復(fù)(NER)鉸鏈物NER, HR & NHEJ的綜合修復(fù) 人類 DNA修復(fù)通路 修復(fù)類型 基因種類 損傷種類 單堿基剪切修復(fù)DNA ligase (LIG3),單堿基損傷DNA glycosylase (MBD4, MPG, MYH, NTH1, OGG1, SMUG1,TDG, UNG), APE1, APE2, XRCC1,ADPRT, ADPRTL2, ADPRTL3 . 多堿基剪切修復(fù)XPA, XPC, XPE, XPF/ERCC4, 多堿基損傷 XPG

5、/ERCC5, ERCC1, LIG1, 紫外線、吸煙 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 . 堿基錯(cuò)配修復(fù)MSH2, MHS3, MSH6, MSH4,堿基錯(cuò)配MSH5, MLH1, MLH3, PMS1, PMS2, PMS2L3, PMS2L4 .Wood et al, Science, 2001重組修

6、復(fù) RAD50, RAD51, RAD51B, RAD51C, DNA雙鏈斷裂 RAD51D, RAD54L, RAD54B, V(D)J重組 RAD52, DMC1, MRE11A, NBS1, ERCC1, XPF/ERCC4, XRCC2 XRCC3, XRCC4, XRCC5, XRCC6 XRCC7, XRCC8, BRCA1, BRCA2 .XP病人和正常人中皮膚癌發(fā)生的年齡分布Kraemer, PNAS, 1997Skin cancers in normal populationSkin cancers in XP populationXP = xeroderma pigment

7、osum變量病例 對(duì)照OR (95% CI) P value No. (%) No. (%)總數(shù) 895 898年齡 0.493 55 439 (49.1) 455 (50.7) 55 456 (50.9) 443 (49.3)性別 0.231 男 674 (75.3) 654 (72.8) 女 221 (24.7)244 (27.2)種族 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)吸煙 0.0001 從 無(wú)291 (32.5)310 (34.5) 1.39 (1.091.78) 曾經(jīng)239 (2

8、6.7)441 (49.1) 1.00 未延續(xù) 365 (40.8)147 (16.4) 3.97 (3.085.13)喝酒 0.0001 從無(wú) 235 (26.3)398 (44.3) 1.00 曾經(jīng)206 (23.0)149 (16.6) 2.12 (1.602.81) 未延續(xù) 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 無(wú) 379 (42.3)387 (43.1) 1.01 (0.83-1.23)頭頸部腫瘤病例-對(duì)照研討中部分變量的頻數(shù)分布2000-2006 *Differences

9、between cases and controls were analyzed by two-sided 2 and Students t tests 實(shí)驗(yàn)室血樣處置流程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 煙草致癌物誘導(dǎo)的DNA損傷與修復(fù)機(jī)制OHBPDEOHO過(guò)氧化酶DNA加成物DNA恢復(fù)正常Benzoapyrene吸煙修復(fù)復(fù)合體銜接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-誘導(dǎo)DNA加成物的檢測(cè)頭頸部腫瘤研討Cancer Res 2007; 67: (12). June 15, 2007變量 病例 (n = 803) 對(duì)照 (n = 839)P or OR (95% CI)* n (%) n (%)總數(shù)加成物中位數(shù) 29.22 / 107 508 (63.3) 419 (50.0) 1.71 (1.392.10)白人加成物中位數(shù) 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.染色體對(duì)DNA損傷劑的敏感性測(cè)定變量 病例 (n = 895) 對(duì)照 (n = 898)P or OR (95% CI)* n (%) n (%)總數(shù)中位數(shù) 0.43 548 (61.2) 457 (50.9) 1.57 (1.291.90)白人中位數(shù) 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 宿主報(bào)告基因的DNA損傷修復(fù)測(cè)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 趨勢(shì)檢驗(yàn) : P 0.001 調(diào)整相對(duì)比數(shù)DRC (%) 四分位數(shù)個(gè)體DNA修復(fù)功能與頭頸部腫瘤發(fā)病分險(xiǎn) High LowLi et al., Can Res, 2007病例 = 721對(duì)照 = 721 趨勢(shì)檢驗(yàn): P 0.0001修復(fù)功能Cases = 767Controls = 763 趨勢(shì)檢驗(yàn): P = 0.0001染色體敏感性DNA加成物Wang et al.,CCR, 2021Wang et al., Cancer Res, 2007反式蛋白質(zhì)芯片測(cè)定修復(fù)蛋白程度Wei et al., CEBP, 200557病例，63對(duì)照0.0 1.0 2.0 3.0

15、 4.0 5.0 6.0 7.0 相對(duì)比數(shù)相對(duì)表達(dá)程度:ERCC1XPAXPCXPD/ERCC2XPF/ERCC4XPG/ERCC5(compared to beta-actin)NER修復(fù)蛋白程度與頭頸部腫瘤發(fā)病風(fēng)險(xiǎn)病例 = 57對(duì)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多堿基剪切

17、修復(fù)基因多態(tài)位點(diǎn)及對(duì)其功能的影響(nsSNPs in NER genes)多堿基剪切修復(fù)基因多態(tài)位點(diǎn)與頭頸部腫瘤發(fā)病風(fēng)險(xiǎn)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)結(jié)論需求有高