《药物的致癌性》PPT课件.ppt

药物致癌性的研究现状和动态 第二军医大学药物安全性评价中心第二军医大学卫生毒理学教研室张天宝 药物致癌性研究的必要性 肿瘤是一类严重影响人类健康和生命的疾病肿瘤已成为人类死亡的第1或2位原因 每年约有700万人死于癌症 据2003年WHO资料 目前每年新增肿瘤1000万人 其中男性530万人 与1990年相比 全球癌症患者发病率增长19 死亡率增长了18 我国恶性肿瘤 2000年 在各种死因中排在第二位 城市已排在首位 每年新增肿瘤200万人 据预测我国2010年年发病数220万 2020年为300万 Age adjustedCancerDeathRates bySite US 1930 2005 1957 1984 1999 2004年我国城市主要疾病死因构成比及死因顺位 特别严重的是 肿瘤是我国最佳劳动人口的首要死因 在35 59岁年龄人口中 所有年龄组的第一位死因都是肿瘤 只有到了60岁以后脑血管或心血管疾病才上升为第1位死因 Whatmaycausecancer HereditarydisordersChemicalsVirusesChronicinflammation WORLDHEALTHORGANIZATIONINTERNATIONALAGENCYFORRESEARCHONCANCERIARCMonographEvaluationsLYON FRANCE SlidecourtesyofV Cogliano IARC IARC 2009 monographs iarc frCarcinogenictohumans group1 108agentsProbablycarcinogenictohumans group2A 66Possiblycarcinogenictohumans group2B 248Notclassifiableastoitscarcinogenicitytohumans group3 515Probablynotcarcinogenictohumans group4 1 IARC IARCGroup1 Carcinogenictohumans Medicaldrugsandtreatments24Industrialprocesses13Infectiousagentsorprocesses10Physicalagents10Industrialchemicals7Inhaledparticulates5Metalsandinorganicsalts5Lifestylefactors incl herbalremedies 7Other8 Group2A 66Medicaldrugsandtreatments12 ChemicalCarcinogenesisinthe21stCentury Newperceptionsofpreviouslyknowncarcinogens CombinedeffectsofmultipleexposuresExamples AlcoholdrinkingandaflatoxinsAlcoholdrinkingandHBV HBCAlcoholdrinkingandtobaccosmokingTobaccosmokingandasbestos arsenic radon 在研究药物的潜在致癌作用中 致癌试验比现有遗传毒性试验和系统暴露评价技术更有意义 致癌试验仍是目前评价药物致癌作用最可靠和最有意义的方法 已评价的致癌物中有93 515 554 至少在三项标准遗传毒性试验中有一项呈阳性 表明在检测致癌物 敏感性 是成功的 然而鉴定非致癌物的能力 特异性 较差 183种在大 小鼠致癌试验中为阴性的物质80 以上有体外遗传毒性阳性的资料 TheEuropeanCentrefortheValidationofAlternativeMethods ECVAM Arecentanalysisofnearly1000chemicalsforwhichdatahavebeenpublishedhashighlightedthestrikinglyimprecisenatureofinvitrogenetictoxicologytestsindiscriminatingnon carcinogensfromcarcinogens Whenthestandardbatteryoftwoorthreeinvitrogenotoxicitytestswasperformed atleast80 ofthe177non carcinogeniccompoundstestedgaveafalsepositiveresultinatleastonetest ThefalsepositiveratewashighestinmammaliancelltestssuchasthosetodetectchromosomalAberrationsormicronucleusinChinesehamstercells orMutationsinthemouselymphomaassay AsimilaroutcomewasobtainedinanalysisbytheU S FDAofanevenlargerdatabaseofchemicals PerformanceofindividualgenotoxictestsindetectingrodentcarcinogensasanalyzedbyKirklandetal 2005 Performanceofsimultaneoustestingbatteriesofgenotoxictestsindetectingrodentcarcinogensasanalyzed Invitrogenotoxtesting theproblem poorspecificity 大多数致癌物在组合试验中呈阳性 Good 大多数非致癌物在组合试验中也呈阳性 Bad 特异性 敏感性 Ames MLAAmesMNMN Indomethacin 吲哚美锌 testednegativeforinvivocytogeneticassaysintheregulatorytests butwasreportedpositivefortheinductionofDNAadductsintheliterature Halothane 氟烷 andpyrazinamide 吡嗪酰胺 werealsoinvivopositiveforcomettestinhumanlymphocytesandinductionofspermheadabnormalitiesinmice respectively whichareconsiderednon regulatorytests 某些药物是非遗传毒性的致癌物 用遗传毒性试验无法检出 很多管理机构都提出了致癌试验的要求 日本 1990 如果临床预期连续用药6个月或更长时间 则需要进行致癌试验 尽管连续用药少于6个月 如果存在潜在致癌性因素 也可能需要进行致癌试验 美国 一般药物使用3个月或更长时间 需要进行致癌试验 欧洲 规定长期应用的药物 即至少6个月的连续用药 或频繁的间歇性用药以致总的暴露量与前者相似的药物需要进行致癌试验 2010年04月01日我国SFDA制定发布了 药物致癌试验必要性的技术指导原则 药物致癌试验 2010年04月01日SFDA制定发布了 药物致癌试验必要性的技术指导原则 预期临床用药期至少连续6个月的药物一般应进行 连续用药没有6个月 但以间歇的方式重复使用 如治疗慢性和复发性疾病 包括过敏性鼻炎 抑郁症和焦虑症 而需经常间歇使用的药物 一般也需进行 某些可能导致暴露时间延长的释药系统 也应考虑进行致癌试验 存在潜在致癌的担忧因素1 已有证据显示此类药物具有与人类相关的潜在致癌性 2 其构效关系提示致癌的风险 3 重复给药毒性试验中有癌前病变的证据 4 导致局部组织反应或其它病理生理变化的化合物或其代谢产物在组织内长期滞留内源性肽类 蛋白类物质及其类似物对于替代治疗的内源性物质 浓度在生理水平 尤其是当同类产品 如动物胰岛素 垂体来源的生长激素和降钙素 已有临床使用经验时 通常不需要进行致癌试验1 其生物活性与天然物质明显不同 2 与天然物质比较显示修饰后结构发生明显改变3 药物的暴露量超过了血液或组织中的正常水平 化学致癌的过程及其机制 化学致癌 chemicalcarcinogenesis 化学物质引起或增进正常细胞发生恶性转化并发展成为肿瘤的过程 具有这类作用的化学物质称为化学致癌物 chemicalcarcinogen 化学致癌研究的重要历史事件 1761 JHill提出使用鼻烟可能会诱发鼻咽癌1775 PPott提出扫烟囱男童阴囊癌与煤烟过度暴露有关1895 LRehn首次报道从事苯胺染料生产的工人发生膀胱癌1914 TBoveri提出恶性肿瘤起源于存在染色体异常的单个细胞 这就是著名的癌症体细胞突变理论和肿瘤单细胞克隆起源学说 1915 Yamagiwa KIchikawa通过长期给兔耳涂煤焦油成功地诱发皮肤癌 实验性化学致癌研究的开端 1932 1938 先后给雄性小鼠注射雌激素诱发乳腺癌 通过慢性饲喂偶氮染料O 氨基偶氮甲苯诱发出大鼠肝癌 使用 萘胺诱发狗膀胱癌成功1941 1944 首次提出启动 Initation 和促进 Promotion 的概念 根据多次使用巴豆油能促进苯并芘诱发小鼠皮肤癌提出小鼠皮肤癌两阶段致癌模型 1949Foulds提出肿瘤演进 Progression 概念1971 1981 CPeraino等 发现小鼠皮肤癌两阶段致癌理论同样适用于大鼠肝癌的发生情况 随后建立了适用于各种脏器肿瘤的多阶段癌变理论1984 ABalmain 首次报道在化学致癌物诱发的小鼠皮肤乳头状瘤中的c Ha ras基因被激活 随后发现多种致癌物可使不同的癌基因活化和抑癌基因失活 InitiatingEvent CellProliferation clonalexpansion Progression CellProliferation CellProliferation Malignancy SecondMutatingEvent N MutatingEvent Initiation Promotion StagesofCarcinogenesis CellularandMolecularMechanismsinMultistageCarcinogenesis INITIATION Initiatingeventinvolvescellulargenome MUTATIONSTargetgenes oncogenes tumorsuppressorgenes signaltransduction cellcycle apoptosisregulators Simple geneticchanges GenticandEpigeneticModelsofTheCancerInitiation Epigeneticallyreprogrammedcells Mutatorphenotypecells Endogenous Environmental ALTERATIONSINCELLULAREPIGENOME Normalcells Cancercells Clonalselectionandexpressionofinitiatedcells Mutatorphenotypecells Endogenous Environmental ACQUISITIONOFADDITIONALRANDOMMUTATIONS Normalcells Cancercells CellularandMolecularMechanismsinMultistageCarcinogenesis PROMOTION Reversibleenhancement repressionofgeneexpression increasedcellproliferation inhibitionofapoptosisNodirectstructuralalterationinDNAbyagentoritsmetabolites CellularandMolecularMechanismsinMultistageCarcinogenesis PROGRESSION Irreversibleenhancement repressionofgeneexpressionComplexgeneticalterations chromosomaltranslocations deletions geneamplifications recombinations etc Selectionofneoplasticcellsforoptimalgrowthgenotype phenotypeinresponsetothecellularenvironment Complex geneticchanges 致癌过程不同阶段的特征 InitiationDNAmodification Mutation GenotoxicOnecelldivisionnecessarytolockinmutationModificationisnotenoughtoproducecancerNonreversible SingletreatmentcaninducemutationPromotionNodirectDNAmodification Nongenotoxic NodirectmutationMultiplecelldivisionsnecessary ThresholdClonalexpansionoftheinitiatedcellpopulationIncreaseincellproliferationordecreaseincelldeath apoptosis Reversible Multipletreatments prolongedtreatment necessaryProgressionDNAmodification Genotoxicevent Mutation chromosomedisarrangement IrreversibleChangesfrompreneoplasiatoneoplasiabenign malignantNumberoftreatmentsneededwithcompoundunknown 遗传毒物对DNA和非DNA相互作用的机制 正常细胞 遗传改变 突变 遗传改变的细胞 癌细胞 异倍体细胞 表遗传干扰 遗传毒性致癌物 非遗传毒性致癌物 遗传毒性和非遗传毒性致癌物在多阶段致癌中的区别 遗传毒性和非遗传毒性致癌物作用机制的比较 化学致癌的生物学特征致癌物多数具有遗传毒性 遗传毒性致癌物尽管化学结构和性质不尽相同 但有一共同的特点 即皆为亲电子剂 致癌作用依赖于化学致癌物的剂量 大剂量的致癌物可增强肿瘤的发生 缩短潜伏期 肿瘤的产生取决于化学致癌物的总剂量 同时暴露于几种致癌物 可发生联合作用 致癌作用的充分表达需相当长的时间 无论致癌物的剂量和性质如何 在肿瘤形成前 总有一个潜伏期 在细胞恶变以前 细胞存在着多阶段的癌前病变 致癌作用所引起的细胞变化可传到子细胞致癌作用可被非致癌因子所修饰细胞增生是细胞癌变过程的重要阶段 Thedevelopmentofcancerisamulti stepprocess Initiation formsanearlyadenoma Promotion leadstoalateadenoma Progression leadsfirsttoacancerinsitu thenonto Malignantconversion whichleadstotrueacarcinomaThissetofprocessesoftentakesYEARS Genotoxiccarcinogens increasetumourfrequencyinanimalcancerbioassaypositiveresultsfrominvitroandinvivogenotoxicitytestseitherdirect actingorindirectlyactinggenotoxiccarcinogens Non genotoxiccarcinogens usuallyactastumorpromoterspositiveincancerbioassayinanimals butnegativeingenotoxicitytestsThemechanismofcarcinogenicitymayincludethechronicinjuryandregenerationhormonalmechanismsincreaseinthecellproliferationordecreaseinthecelldeathintargetorgan Theconceptofa complete vs an incomplete carcinogen Whenoneforeignchemicalissuffienttocausecancer eitherasadirectorindirectcarcinogen itissaidtobecompleteWhenitrequiresatumorpromotertocausecancer itisanincompletecarcinogenPromotorsarecompoundsthatinducecells likethemutatedcancerinitiatorcell togrowanddivide makingmore ICHGuidelineS1BonTestingforCarcinogenicityofPharmaceuticals choosingone2 yearrodentcarcinogenicitystudy rat plusoneotherstudythatsupplementsthe2 yearstudyandprovidingadditionalinformationthatisnotreadilyavailablefromthe2 yearstudy either 1 ashort ormedium terminvivorodenttestsystemor 2 a2 yearcarcinogenicitystudyinasecondrodentspecies mouse theshort ormedium termmodelswasintendedtofocusontheuseofinvivomodelsprovidinginsightintocarcinogenicendpointssuchasinitiation promotionrodentmodelsandmodelsofcarcinogenesisusingtransgenicorneonatalrodents 药物致癌性评价方法 StipulatedRationaleforChoosingaShort orMedium TermTestSystemasSupplementtoOne2 YearBioassay Themechanismofcarcinogenesisinthemodelshouldmostlikelyberelevanttohumans andthereforetheuseofthemodelshouldbeapplicabletohumanriskassessment Theuseofthemodelshouldsupplementthe2 yearcarcinogenicitystudyanditshouldprovideadditionalinformationthatisnotreadilyavailablefromthe2 yearstudy Animalwelfare animalnumbers andoveralleconomyofthecarcinogenicevaluationprocessshouldbeconsidered Two yearCarcinogenesis Bioassay Protocol CurrentGlobalCarcinogenicityStudyRequirements StandardTissueList KidneyUrinarybladderAortaHeartTracheaLungsLiverGallbladderPancreasFatSalivaryglandSpleenCervicallymphnodeMesentericlymphnodeThymusTongueEsophagusStomachDuodenumJejunumIleumCecumColonMammaryglandSkinSkeletalmuscleSciaticnerveParathyroidThyroidAdrenalglandPituitaryProstateSeminalvesiclesTestesEpididymidesOvariesOviductsUterinehornsUterinebodyCervixVaginaBrainSpinalcordSternumRib boneEyesHarderianglandsBMsmearNaresClitoral preputialglandZymbal sglandGrosslesions 美国毒性病理学会 STP 建议致癌试验进行组织病理学检查的最基本的受检内容目录 Tumor BearingAnimalsinControlGroupsfromRodentStudies Source J K Haseman unpublishedsummaryofU S NTPdata ComparativePercentIncidenceofPertinentNeoplasiainDifferentStrainsofRatsandMice 104WeeksOld Note F344 Fischer244rats S D Sprague Dawleyrats B6C3F1 mice C57BL 6N C3H HeN F1 CD 1 1CRCr CD 1mice NA nonapplicable theaveragenumberusedbyspecies strain genderwasinexcessof750animals Preclinicalapproachesforassessingcarcinogenicpotential Tumorigenicityinhumans nonhumanprimatesandrodents Spontaneoustumorratesinthebreederandcontrolanimals TheNeonatalMouse Pietraetal 1959 Theneonatalmouseisoneofthealternativeinvivomodels fordetectingthecarcinogenicpotentialofpharmaceuticals ThisisinagreementwiththesuggestionsofICH whichallowstheuseofonealternativestudyinplaceofoneofthe2 yearcarcinogenicitystudies Whentreatmentbeginswithinthefirst24hoursoflife thestudydesignisdescribedas newbornmouse neonatalmouse includestestitemadministrationatdifferenttimepointsfrombirthtothreeweeksofage Fujii 1991 reportedthattheneonatalmouseassayshowedasensitivityof85 andapositivepredictionrateof96 comparedtotheresultsoftheadultmouse2 yearcarcinogenicitystudy Flammangetal 1997 consideredthismodeltohavehighsensitivityandspecificitytodetectgenotoxiccarcinogensaswellaspresentingadvantagessuchasreducedtestarticlerequirements decreasedanimalnumbersandcostsandareducedcompletiontime Itdoesnotrespondtochemicalsactingviaepigeneticmechanisms McClainetal 2001 reportedthatneonatalmicehavebeenshowntohaveareducedtimefortumorinduction ahighermultiplicityofinducedtumors alowerspontaneoustumorrateandanequivalentorhighersensitivitytocarcinogenswhencomparedtoadultmice Thismodelalsorespondstoawiderangeofstructurallydissimilargenotoxiccompounds Additionally theneonatalmousepossessesthemajorityofthephaseIandIIbiotransformationliverenzymesinvolvedintheprocessesofactivationanddetoxificationofcarcinogensfromdifferentchemicalclasses CD 1mice10to12weeksofage Micewerecagedwith5femalespermaleandexaminedeachdayforthepresenceofavaginalcopulationplug Femaleswereisolateduntildelivery 6litterswith4neonates sex litterwereassignedtoeachgroupduringthefirstweekafterbirth Threeorfourdoselevels avehicleandapositivecontrolwereused Groupsconsistedof24animals sex group Theyweredosedonthebasisoftheiraveragebodyweight ondays8and15ofage usingdosevolumesofupto100and200 l respectively Doselevelswereselectedonthebasisoftheresultsobtainedindoserangefindingstudies inwhichtheMTDortheMFD MaximumFeasibleDose forneonatalmice weredetermined Thepupswereweanedaround22daysofage housed4 sex cageandthenmaintaineduntil1yearofage whentheyweresacrificed DEN diethylnitrosamine atadosageof2mg kgdissolvedinwaterwasusedasthepositivecontrol Tg AC v Ha ras TransgenicMouse 8 9weeksoldGroupsof15mice sex dosewererandomlyassignedtothestudygroups Thevehiclesusedfordrugsandpositivecontrolagentswereacetone ethanolorDMSO TPA 12 o tetradecanoylphorbol 13 acetate wasthepositivecontrolcompound26weeks Hemizygousp53 KnockoutMouse6to10weeksold GenotypeanalysiswasrecommendedpriortoassignmenttodosegroupsGroupsof15mice sex group Threedoselevels avehicleandapositivecontrolwereused Twoadditionalgroupsof15wild typemice sex groupreceivedthevehicleandthehighdosageofthetestcompound p Cresidineat400mg kg daybygavageincornoil 10ml kg orbenzeneat100mg kg daybygavageincornoil 5ml kg wererecommendedaspos