决策树是通过递回分割

AFive-GeneSignatureandClinicalOutcome

inNon–Small-CellLungCancer

From:nengljmed356;1january4,2007By:Hsuan-YuChen,M.Sc.,Sung-LiangYu,Ph.D.,etalReporter:R6謝廣宇

Background

Currentstagingmethodsareinadequateforpredictingoutcomedevelopeda5-genesignaturenon–small-celllungcancer(NSCLC)—mostcommoncauseofdeathfromcancerworldwiderelapseratewithearly-stageNSCLCis40%within5yearsafterpotentiallycurativetreatment決策樹是通過遞迴分割（recursivepartitioning）建立而成，遞迴分割是一種把資料分割成不同小的部分的疊代過程cDNAmicroarrayschemacDNA晶片製造原理MicroarrayanalysisOperationPrinciple:Samplesaretaggedwithflourescentmaterialtoshowpatternofsample-probeinteraction(hybridization)Microarraymayhave60KprobeMicroarrayProcessingsequenceFrom:Shin-MuTsengtsengsm@.twGene-expressionprofilingbymeansofmicroarraysandreverse-transcriptasepolymerasechainreaction(RT-PCR)examinedgeneexpressionin125surgicalspecimensofNSCLC,usingmicroarraysandreal-timeRT-PCRtoidentifyagenesignaturecorrelatedwithclinicaloutcomeMethodscomputer-generatedrandomnumberstoassignspecimensfrom185consecutivepatientsformicroarrayanalysisfrozenspecimensoflung-cancertissuefrom125randomlyselectedpatientswhounderwentsurgicalresectionofNSCLCatTaichungVeteransGeneralHospitalbetweenDecember1999andDecember2003------five-generisk-predictionmodelusinganindependentcohortof60randomlyselectedpatients125specimens--60wereadenocarcinomas,52weresquamous-cellcarcinomas,and13wereothertypesofcancernotreceivedadjuvantchemotherapy672genesassociatedwithinvasiveactivity,identifiedinapreviousstudy---rearrayedinduplicateonanylonmembrane4μgoftotalRNAfromeachspecimenvalidatelevelsofexpressionofgenesfoundonmicroarrayanalysisRT-PCRwasperformedon16genesandacontrolgeneforTATA-box–bindingprotein(TBP),withuseofspecificTaqManprobesandprimersets---transcriptswereamplifiedwithreagent(TaqManOne-StepRT-PCRMasterMixReagent,AppliedBiosystems)andasequencedetectionsystem(ABIPrism7900HT,AppliedBiosystems)Toreducebackgroundnoisebackgroundintensityvaluesoflessthan3000wereassignedvalueof3000log-transformedtoabase-2scaleGeneswithcoefficientsofvariationoflessthan3%wereexcludedfromfurtheranalysesthegene-expressionintensityvaluesweretransformedtoordinalcodingvaluesbyrankingoflevelofgeneexpressionamongthe485genesin125patients(60,625observations)intensityvaluewascodedas1-2-3-4accordingto0-25%-50%-75%-100%HazardratiosfromunivariateCoxregressionanalysistodeterminewhichgenesassociatedwithdeathfromanycauseorrecurrenceofcancerProtectivegenesweredefinedwithahazardratiofordeathof<1;riskgenesweredefinedwithahazardratiofordeathof>1univariateCoxproportional-hazardsregressionanalysisGenessignificantlycorrelatedwithsurvival---alinearcombinationofgene-expressioncodingvaluesweightedbyregressioncoefficientstocalculateariskscoreforeachpatientpatient’sriskscorewascalculatedassumoflevelsofexpressionofeachgene,asmeasuredbymicroarrayanalysis,multipliedbycorrespondingregressioncoefficientshigh-riskgenesignatureoralow-riskgenesignature,with50thpercentile(median)oftheriskscoreasthethresholdvalue(median,4.9;range,1.3to21.9)---thresholdvaluetoreflectalmosthalfofpatientswithearlystageNSCLCrelapsewithin5yearsafterpotentiallycurativesurgery,eliminateeffectofextremevalueslevelsofexpressionof16genesconfirmedbyRT-PCRandindexedbySpearman’srank-correlationtestfurtheridentified5genessignificantlyassociatedwithsurvivalrecursive-partitioningdecisiontree&Avadissoftware(StrandGenomic)ahigh-riskgenesignatureoralow-riskgenesignatureKaplan–Meiermethodtoestimateoverallsurvivalandrelapse-freesurvivalMultivariateCoxproportionalhazardsregressionanalysiswithstepwiseselectionevaluateindependentprognosticfactorswithsurvival,5-genesignature,age,sex,tumorstage,andhistologiccharacteristicsascovariatesTofurthervalidateourmodel,weappliedittomicroarraydatafrom86patientswithNSCLC,reportedbyBeeretalfivegenes(andtheircorrespondingAffymetrixprobesets)wereDUSP6(X93920_at),MMD(X85750_at),STAT1(M97936_at),ERBB3(S61953_at),andLCK(M26692_s_at);thecontrolgenewasTBP(X54993_s_at)maximumfollow-uptimeforsurvivalanalysisinourstudywas62months,weused5-yearsurvivaldatafor86patientsResults16genescorrelatedwithdeathfromanycause:4wereprotectivegenes(hazardratio<1)and12wereriskgenes(hazardratio>1)dual-specificityphosphatase6(DUSP6)monocyte-to-macrophagedifferentiationassociatedprotein(MMD)signaltransducerandactivatoroftranscription1(STAT1)v-erb-b2avianerythroblasticleukemiaviraloncogenehomolog3(ERBB3)lymphocyte-specificproteintyrosinekinase(LCK)5-genesignaturestronglyassociatedwithoverallsurvival(sensitivity,98%;specificity,93%;positivepredictivevalue,95%;negativepredictivevalue,98%;andoverallaccuracy,96%)AmongpatientswithstageIIdisease,overallsurvivaldidnotdiffersignificantlybetweenhigh-riskandlow-riskgenesignatures,probablyowingtosmallnumberofpatientsDiscussionNSCLCisaheterogeneousdiseasebasedonclinicalandpathologicalfindingsmayhavelimitofusefulnessforpredictingoutcomes,butmolecularmethodsaddvalueuseofmicroarraysinclinicalpracticelimitedbythelargenumberofgenesintheanalysis,complicatedmethods,lackofreproducibilityandindependentvalidationofresults,andneedforfresh-frozentissueRT-PCRinvolvingasmallnumberofgenesallowsforaccurateandreproduciblequantificationofresultsforRNAobtainedfromsmallamountsofparaffin-embeddedspecimens125frozentumorspecimensfrompatientswithNSCLCrandomlydividedintoatrainingset(63specimens)andatestingset(62specimens)selectionofgenesinmicroarraytrainingsetwasvalidatedinmicroarraytestingset,andpatternsofgeneexpressionfoundonmicroarrayanalysiswerevalidatedbyRT-PCRresultsinourChinesepatientswerealsovalidatedwithuseofasetofpublishedNSCLCmicroarraydatafrompatientsfromaWesternpopulationwithNSCLCproposewhohavetumorswithahigh-riskgenesignaturecouldbenefitfromCisplatin-basedadjuvantchemotherapy,thosewithalow-riskgenesignaturecouldbespared---large-scale,multicenterstudiesarenecessarytotestthisideaSTAT1arrestedgrowthandapoptosisinmanytypesofcancercellsbyinducingexpressionofp21WAF1andcaspase.MMDexpressedinmaturemacrophages,macrophageactivationpromotescancermetastasis,BUTfunctionofMMDproteinisunknownDUSP6inactivatesextracellularsignal-regul