TargetingAminoAcidTransportinMetastaticCastration-ResistantProstateCancer:EffectsonCellCycle,CellGrowth,andTumorDevelopment
QianWang,JessamyTiffen,CharlesG.Bailey,MelanieL.Lehman,WilliamRitchie,LadanFazli,CynthiaMetierre,Yue(Julie)Feng,EstelleLi,MartinGleave,GrantBuchanan,ColleenC.Nelson,JohnE.J.Rasko,JeffHolst
Correspondenceto:JeffHolst,PhD,OriginsofCancerLaboratory,LockedBag6,Newtown,NSW2042Australia.(e-mail:
j.holst@centenary.org.au).
BackgroundL-typeaminoacidtransporters(LATs)uptakeneutralaminoacidsincludingL-leucineintocells,stimulatingmammaliantargetofrapamycincomplex1signalingandproteinsynthesis.LAT1andLAT3areoverexpressedatdifferentstagesofprostatecancer,andtheyarerespons
IBLeforincreasingnutrientsandstimulatingcellgrowth.
MethodsWeexaminedLAT3proteinexpressioninhumanprostatecancertissuemicroarrays.LATfunctionwasinhibitedusingaleucineanalog(BCH)inandrogen-dependentand-independentenvironments,withgeneexpressionanalyzedbymicroarray.APC-3xenograftmousemodelwasusedtostudytheeffectsofinhibitingLAT1andLAT3expression.ResultswereanalyzedwiththeMann-Whitney
UorFisherexacttests.Allstatisticaltestsweretwo-sided.
ResultsLAT3proteinwasexpressedatallstagesofprostatecancer,withastatisticallysignificantdecreaseinexpressionafter4–7monthsofneoadjuvanthormonetherapy(4–7monthmean=1.571;95%confidenceinterval=1.155to1.987vs0month=2.098;95%confidenceinterval=1.962to2.235;
P=.0187).InhibitionofLATfunctionledtoactivatingtranscriptionfactor4–mediatedupregulationofaminoacidtransportersincludingASCT1,ASCT2,and4F2hc,allofwhichwerealsoregulatedviatheandrogenreceptor.LATinhibitionsuppressedM-phasecellcyclegenesregulatedbyE2Ffamilytranscriptionfactorsincludingcriticalcastration-resistantprostatecancerregulatorygenes
UBE2C,CDC20,and
CDK1.InsilicoanalysisofBCH-downregulatedgenesshowedthat90.9%arestatisticallysignificantlyupregulatedinmetastaticcastration-resistantprostatecancer.Finally,LAT1orLAT3knockdowninxenograftsinhibitedtumorgrowth,cellcycleprogression,andspontaneousmetastasisinvivo.
ConclusionInhibitionofLATtransportersmayprovideanoveltherapeutictargetinmetastaticcastration-resistantprostatecancer,viasuppressionofmammaliantargetofrapamycincomplex1activityandM-phasecellcyclegenes.
L-typeaminoacidtransporters(LATs)supplycellswithlargeneutralaminoacids,whicharenotonlyrequiredforproteinsynthesisbutalsocontributetovarioussignalingpathways.Intracellularleucinelevelsaresensedbytheleucyl-transferRNAsynthetase,previouslyknowntocatalyzetheadenosinetriphosphate–dependentligationofL-leucinetotransferRNAduringproteinsynthesis(
1,
2).Leucyl-transferRNAsynthetaseactivatestheRagguanosinetriphosphatasecomplexandbindstoRaptortoactivatemammaliantargetofrapamycincomplex1(mTORC1)signalingonthesurfaceoflysosomes(
1–3).Inthiswayleucineisnotonlyanessentialaminoacidbutactsasarate-limitingsignalingmoleculeinthemTORC1pathway.
Incellsdeprivedofaminoacids,thereisanaccumulationofunchargedtransferRNA,whichbindstoandactivatesthegeneralcontrolnonrepressed2(GCN2)kinase.Inturn,GCN2phosphorylatesthetranslationinitiationfactor2α(eIF2α)onserine51,triggeringtranslationalupregulationofactivatingtranscriptionfactor(ATF)4(
4).ATF4itselfupregulatestheexpressionofaminoacidtransportersasameansofrestoringintracellularaminoacidlevels(
5).Therefore,understandinghowaminoacidtransportersregulateintracellularleucinelevels,andgeneratingnovelinhibitorsofthesetransporters,mayleadtopotentsuppressorsofmTORC1signaling.
ThetwodistinctfamiliesofLATsare1)solutecarrier7(SLC7)members(LAT1/SLC7A5andLAT2/SLC7A8),whichmediateNa+-independentneutralaminoacidexchangeasheterodimerswiththe4F2cell-surfaceantigenheavychain(4F2hc/SLC3A2/CD98)glycoprotein(
6,
7);and2)SLC43proteins(LAT3/SLC43A1andLAT4/SLC43A2)thatmediateNa+-independentuniportofneutralaminoacids(
8,
9).AlthoughtheexpressionofeachLATmembervariesdramaticallyindifferenttissues,thesetransportersarecommonlyupregulatedincancer.IncreasedLAT1expressionhasbeendetectedinlungcancer,coloncancer,breastcancer,headandneckcancer,genitalcancers,andsofttissuesarcomas(
10–12).WeandothershaveshownthatLAT1andLAT3areoverexpressedinprostatecancer(
11–14),withLAT1expressionincreasedinmetastasiscomparedwithprimarycancer(
10,
12).
WehypothesizedthatinhibitionofLAT1andLAT3mayofferaneffectivetherapeuticapproachforprostatecancer.
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