yCDglyTK融合自杀基因前药系统对鼻咽癌CNE22细

:863(2001AA217181):,,,:,Email:[e-mail]221,1,1,2,2,3(11,410008;21,410078;31,410078):yCDglyTKCNE22pcDNA3.1(-)CMVeEgr21yCDglyTKCNE22,,(MTT),CNE22,CNE22yCDglyTK;CNE22,yCDglyTKCNE22,CNE22:yCDglyTK;;/:R739.63;R730.231.3:A:1007-1520(2005)02-0072-06RadiosensitizationbyyCDglyTKfusionsuicidegene/prodrugsysteminhumannasopharyngealcarcinomaCNE22celllineZHANGJu2yi,ZHAOSu2ping,XIAOJian2yun,etal.(DepartmentofOtolaryngology,XiangyaHospital,CentralSouthUniversity,Changsha410078,China)Abstract:ObjectiveToinvestigatetheradiosensitizationbyyCDglyTKfusionsuicidegene/prodrugsysteminhumannasopharyngealcarcinomacells(CNE22).MethodsTheplasmidvectorofpcDNA3.1(-)CMVeEgr21yCDglyTKwasintroducedintoCNE22cellsbyelectroporation.Thenthecellswereexposedtovariousdosesofradiationandprodrug.ObservationwasmadeforthechangeofexpressionandfunctionoffusionsuicidegeneinyCDglyTK2expressingcellsthroughimmunohistochemistry,HPLC,MTTandclonogenicassay.TheinfluenceingrowthofCNE22cellswasobservedtoo.ResultsIonizationradiationcaninduceanincreasedexpressionofyCDglyTKgeneinsuicide2genepositivecells;CombinedwithionizationradiationandprodrugshowedgreatertoxicitytoyCDglyTK2expressingCNE22cellsthandisposedwithionizationradiationorprodrugalone.ConclusionTheyCDglyTKfusionsuicidegene/prodrugsystemcanenhancetheradiosensitivityofCNE22cells,andshowsynergistickillingeffectwhencombinedwithionizationradiation.Keywords:yCDglyTK;Radiosensitization;Nasopharyngealneoplasms/radiothor,,,,,,[1]pcDNA3.1(-)CMVeEgr21yCDglyTK,2,2711220054ChineseJournalofOtorhinolaryngology-SkullBaseSurgeryVol.11No.2Apr.2005©1995-2005TsinghuaTongfangOpticalDiscCo.,Ltd.Allrightsreserved.yCDglyTKCNE2211.1CNE22,10%RPMI21640,375%CO2RPMI21640GiBCOBRL1.2pcDNA3.1(-)CMVeEgr21yCDglyTK,pcDNA3.1(-)RT2PCR::52GGGA2GATTAGAGGGCAAAGTGGTC23:52ACGGCGTCGGTCACGGCATAA23BALB/C(46,20g,),1.3PE29600PCR(perking2Elmer),(BECKMAN),BioRad,Co60G418:Gibco,52FC52FU:Sigma,MTT:Sigma,AMV:Promega,Trizol:Gibco,TK:QED,:SP9002MouseSPKitZLI29032DABKit1.4pcDNA3.1(-)CMVeEgr21yCDglyTKpcDNA3.1(-)20g,CNE22:2KV25F600g/mlG41810d,,,300g/mlG418,yCDglyTK1.5RT2PCRyCDglyTKyCDglyTK1.0107,RNA,RT2PCR,,yCDglyTKmRNA1.6yCDglyTKT25pcDNA3.1(-)CMVeEgr21yCDglyTKpcDNA3.1(-)CNE220.25%,10ml,0.5ml,15cm4h,20ml24h,0Gy8Gy,12h-20,TritonX2100,yCDglyTK,1.752FCyCDglyTKCNE221105/212,90%,,200g/ml52FC2ml,30min2120Gy,8Gy5d,3,(HPLC)52FU,,1.8CNE22pcDNA3.1(-)yCDglyTK310396,3,16,,,24h,52FC050200400g/ml4,430min30Gy8Gy16Gy5d,9620lMTT(5mg/mL),4h,200l/DMSO,3710min,,96,490nm,,,=A/A100%1.9yCDglyTKCNE221.9.10.25%,,6cm(1)37,:yCDglyTKCNE222©1995-2005TsinghuaTongfangOpticalDiscCo.,Ltd.Allrightsreserved.CNE22yCDglyTK,31.9.212h,CNE22,0Gy1Gy2Gy4Gy6Gy8Gy10Gy,yCDglyTK52FC50g/ml,30min1Gy2Gy4Gy6Gy8Gy10Gy,14d,,30min,,50,CNE220GyPE=(/)100%,=/(PE),,D0(63%)DqSER=D0/D01(Gy)01246810CNE221001501504001000400020000yCDTK(52FC50g/ml)5006001200300012000600001.10SPSS10.0(onewayANOVA),SNKP0.0522.1RT2PCR(1)1yCDglyTKyCDglyTKcDNA707bp,CNE22,yCDglyTKmRNA1RT2PCR0.8%1DNA(2kb);23yCDglyTKyCDglyTKcDNA707bp;4CNE22;5DNA(2kb)2.2pcDNA3.1(-)CNE22yCDglyTK,yCDglyTKCNE22,8Gy(25)2pcDNA3.1(-)CNE22(SP400)3pcDNA3.1(-)CNE228Gy(SP400)4711©1995-2005TsinghuaTongfangOpticalDiscCo.,Ltd.Allrightsreserved.4yCDglyTKCNE22(SP400)5yCDglyTKCNE228Gy(SP400)2.3200g/ml52FC5d,CNE2252FU,52FC52FU0;yCDglyTKCNE2252FU168g/ml,52FC52FU84%;8Gy52FU198g/ml(6),52FC52FU99%,15%,Student2Neuman2Keuls(SNK),(P0.05)yCDglyTK/52FCEgr21,22.42.4.1CNE22pcD2NA3.1(-)yCDglyTK(2)2,,52FC(50200400g/ml),,yCDglyTK,Student2Neuman2Keuls(SNK),,yCDglyTK(P0.01),CNE22(P0.05),52FC,yCDglyTK68GyyCDglyTKCNE2252FC52FU252FC(g/ml)CNE22pcDNA3.1(-)yCDglyTK0100%100%100%5094.5%97.9%37.6%20086.3%86.8%14.0%40073.1%75.8%8.1%2.4.2(3)3,(8Gy16Gy),yCDglyTK,52FC,yCDglyTKStudent2Neuman2Keuls(SNK),(P0.05),(50200400ug/ml),(8Gy16Gy),yCDglyTK(P0.01)57,:yCDglyTKCNE222©1995-2005TsinghuaTongfangOpticalDiscCo.,Ltd.Allrightsreserved.352FC(g/ml)CNE228Gy16GpcDNA3.1(-)8Gy16GyCDglyTK8Gy16G067.3%53.5%64.2%55.4%62.1%54.9%5064.9%47.5%68.2%53.3%21.4%11.2%20053.3%38.8%57.7%43.8%10.3%6.7%40046.3%35.7%48.9%36.4%5.3%2.2%2.5,14d,CNE220Gy1,,(7),CNE22yCDglyTKD02.110.39,Dq1.490.14,SER5.41,yCDglyTKCNE227yCDglyTKCNE223,,60%20%30%,,,,550%60%,,/,,,(),,,CDglyTK,52FCGCVCD52FC(52)52Fu(52),52FuTP52FduTP(522522),RNA,RNA;,DNA,TKGCV(),GCV,DNA,DNA,,,,Niranjan[2]2,Rogulski[3],CDTK,9LGCV/52FC,SER2.44Ueno[4],,,,(MTT)yCDglyTK,yCDglyTKCNE22,,MTT,CNE22,52FC50g/ml,SER5.41,Rogulski[3]6711©1995-2005TsinghuaTongfangOpticalDiscCo.,Ltd.Allrightsreserved.2.44,CD(yCD)CD(bCD)[5],,CNE22,yCDglyTK2,,:,;Egr21,;[6],GCV;[7];,[8],,,,EB1[9,10],2,,:[1],,,1pcDNA3.1(-)CMVEgr21CDglyTK[J]1,2003,2(1):34-37.[2]NiranjanA,MoriuchiS,LunsfordLD,etal.EffectivetreatmentofexperimentalglioblastomabyHSVvector2mediatedTNFalphaandHSV2tkgenetransferincombinationwithradiosurgeryandgancicloviradministration[J].MolTher,2000,2(2):114-120.[3]RogulskiKR,ZhangK,KolozsvaryA,etal.Pronouncedantitu2moreffectsandtumorradiosensitizationofdoublesuicidegenetherapy[J].ClinCancerRes,1997,3(11):2081-2088.[4]UenoM,KoyamaF,YamadaY,etal.Tumor2specificchemo2radio2genetherapyfo