钙调素拮抗剂EBB抑制人乳腺癌细胞生长的实验研究

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1、北京协和医学院;中国医学科学院硕士学位论文钙调素拮抗剂EBB抑制人乳腺癌细胞生长的实验研究姓名：代晓华申请学位级别：硕士专业：生物化学与分子生物学指导教师：朱惠芳;熊冬生20090501北京协和医学院中国医学科学院硕士研究生学位论文白酶活性而抑制细胞凋亡。本实验采用PCR法扩增survivin序列，经酶切、连接，克隆至表达载体pAYZ，转化E．coli16C9，经测序鉴定阳性克隆菌落插入片段为正确编码的survivin基因序列。经低磷培养基AP5诱导表达，用His—tag亲和层析柱纯化，为进一步研究其生物学特性，靶向surviv

2、in治疗恶性肿瘤奠定基础。关键词：钙调素拮抗剂，小檗胺衍生物，MCF一7细胞，氧化应激反应，survivin基因。4北京协和医学院中国医学科学院硕士研究生学位论文AbstractMalignancyincidencewillincrease50％inthenexttwentyyearsintheworld．Atthesametime．ithasbeenthemostserioussicknessthataffectsournationality’Slives．Therefore，howcallwecuremalignancyis

3、thekeyprobleminbiomedicalresearch．Lookingforanti-tumoragents，specificallyagentsthatcaninhibittunlormetastasispresentsapromisingresearchindrugdiscovery．Traditionalplant-derivedmedicinesaresafeandhavebeenusedtotreatcancerforcenturies．Wehaveknownthatsomeplant．sourcemedi

4、cinejustlikePaclitaxelorcamptothecin，showedgoodanti—tumoreffects．Inordertodevelopmorepotentpharmacophorestructure，chargedgroupswereintroducedintotheplant．basedformulationsofchinesetraditionalmedicinethatledtodiscoveryofanewsyntheticderivativespossessinganti—tumormedi

5、cinewithlessside．effect．Inthisrespect，ChineseherbextractionBerbamineanditsderivativeO一4一Ethoxyl—Butyl—Berbarnine(EBB)reallyshowedanattractiveanti-tumoractivity．Berbamineisabisbenzylisoquinolinealkaloidandanaturalcalmodulinantagonist．StudiesshowedthatBerbaminehasanti．

6、tumoreffectsandincreasestheleukocytecount．EBBisaderivativeofBerbamine，whichisoneofthemosteffectivecalmodulinantagonists．EBBhassimilarpharmacologymechanismwithberbamine，buttheabilityofEBBtoantagonizecalmodulinis100timesmorethanberbamine．PreviousstudyshowedthatEBBinhib

7、itedthecellproliferation,reverseddrugresistanceandsuppressedtumorinvasioninseveralkindsoftumorcelllinesinvitro．EBBalsoshowedanti—tumoreffectsinvivo．Inthispaper,weinvestigatedtheeffectsofEBBonhumanbreastcancercelllineMCF-7andexploredthepossiblemechanism．MTTassayWasuse

8、dtoanalyzethegrowthinhibitioneffectsofEBBonseveraltumorcelllines．TheeffectsofEBBonreactiveoxygenspecies(ROS)productionandcellapopto