CTTNconfersTrastuzumabresistancethroughDKK1/WNTsignalinginHER2-positiveBreastCancer

Abstract

Despitethetherapeuticsuccessoftrastuzumab,HER2-positive(HER2+)breastcancerpatientscontinuetofacesignificantdifficultiesduetoinnateoracquireddrugresistance.InthisstudyweexploredthepotentialroleofCTTNininducingtrastuzumabresistanceofHER2+breastcancer.ThegeneticaberrationofCTTNandsurvivalofHER2+breastcancerpatientswereanalyzedusingmultiplebreastcancerpatientcohorts.TheCTTNeffectoncancerstemcellactivitywasconfirmedbytumorsphereformationassay,ALDEFLUORassay,andinvivoxenograftexperiments.CTTN-inducedtrastuzumabresistancewasshownbysulforhodamineB(SRB)assay,andcolonyformationassays.RNA-seqanalysiswasconductedtoidentifythemolecularmechanismfortrastuzumabresistancebyCTTN.CTTNwasrelatedtopoorprognosisinHER2+breastcancer(OS,p=0.05inHYUcohort;OS,p=0.0014inKMplotter).Furthermore,overexpressionofCTTNconferredtrastuzumabresistanceandcancerstemcell-likepropertiesinvitroandinvivo.Mechanistically,themRNAandproteinlevelsofDKK-1weredownregulatedbyCTTN,whichactivatedtheWntsignalingpathway.Combinationaltreatmentwithtrastuzumabandꞵ-catenin/TCFinhibitor,FH535,couldovercomeCTTN-inducedtrastuzumabresistanceandreducecancerstemcell-likeproperties.CTTNactivatesDKK-1/Wnt/ꞵ-cateninsignalingtoinducetrastuzumabresistance,suggestingCTTNasanovelbiomarkerwithapoorprognosisandapossibletherapeutictargetinHER2+breastcancer.|HER2양성유방암의대표적인표적치료제인트라스투주맙(Trastuzumab)의임상적효능에도불구하고,이약물에대한선천적및후천적내성은여전히주요한문제이다.본연구에서는섬유형액틴(F-actin)결합단백질인CTTN이HER2양성유방암의새로운바이오마커로서암줄기세포의특성및trastuzumab에대한반응을조절하는기전을조사하였다.여러유방암환자코호트를이용하여CTTN발현에따른HER2양성유방암환자의생존및CTTN의유전적변형(geneticalteration)을분석한결과CTTN의높은발현이HER2양성유방암환자의좋지못한예후와관련이있음을밝혔다.Tumorsphereformationassay,ALDEFLUORassay,invivoxenograft실험을통해확인하였고CTTN이암줄기세포특성을조절함을확인하였고,sulforhodamineBassay(SRBassay)와colonyformationassay를통해CTTN의과발현이트라스투주맙에대한내성을유도함을확인하였다.CTTN을과발현시킨SKBR3세포주로진행한RNA-sequencing분석결과Wnt적대자(anatagonist)인DKK-1의발현이CTTN에의해감소되는것을확인하였다.그로인해GSK3-ꞵ의인산화와ꞵ-catenin의핵으로의이동이증가됨에따른c-Myc과CyclinD1의증가로CTTN이Wnt/ꞵ-catenin/TCF신호전달계를활성화시킴을증명하였다.CTTN을과발현시킨세포주에ꞵ-catenin/TCFinhibitor인FH535를처리하였을때Wnt신호전달계가억제되고암줄기세포성및trastuzumabresistance가완화되는것이확인되었다.결론적으로본연구결과는CTTN이DKK-1의발현을하향조절하여DKK-1이억제하는Wnt신호전달계를활성화시키며그에따라유방암줄기세포성및trastuzumab내성을유도하는것을밝혔으며,이는CTTN을HER2양성유방암의새로운예후바이오마커이자잠재적인치료적타겟(therapeutictarget)으로서의가능성을제안한다.