Antibody-Targeted Nanoparticles for Cancer Treatment · cytotoxic T cells [4]. However, the...
Transcript of Antibody-Targeted Nanoparticles for Cancer Treatment · cytotoxic T cells [4]. However, the...
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Antibody-TargetedNanoparticlesforCancerTreatmentThomasCarter1,PaulMulholland1,2,KerryChester1AbstractNanoparticlesarediverseandversatilewithphysicalpropertiesthatcanbe
employedforuseincancermedicine.Targetingnanoparticlesusingantibodies
andantibodyfragmentscouldovercomesomeofthelimitationsseenwith
currenttargetedtherapies.Thisreviewwilldiscusstheroleofantibody-targeted
nanoparticlesinthetreatmentofcancer:asdeliveryvehicles,targeted
theranosticagentsandintheevolvingfieldofcancerhyperthermia.
Keywords:Antibodies,nanoparticles,targeting,cancertreatment,clinicaltranslation,theranostics,hyperthermia1.0IntroductionAsourunderstandingofcancerbiologyhasevolved,sotoohastheappreciation
thatantibodytargetedcancertreatmentofferssignificantadvantagesover
conventionaltherapy.Anumberofstrategieshavebeensuccessfulintheclinic:
mostnotablytheuseofmonoclonalantibodies(mAbs)totargetcancer-specific
antigens[1],delivercytotoxicchemotherapyintheformofantibody-drug
conjugates(ADCs)[2],releasebrakesontheimmunesystem[3]andtorecruit
cytotoxicTcells[4].However,thesuccessesofthesetargetedtherapiesarenot
withoutdrawbacks,includingdose-limitingtoxicities.
Advancesinnanotechnologyhaveenabledthecreationofavastarrayofdevices,
collectivelyreferredtoasnanoparticles(NPs),whichpossessuniquephysical,
chemicalandbiologicalproperties.Theuseofantibodiestotargetthesenano-
sizeddrugdeliveryvehiclesoffersnewpotentialthatcanbeharnessedforusein
cancermedicine[5].Indeed,asmallnumberoftargetedNPshavebeenalready
beensuccessfullytranslatedintoearlyphaseclinicaltrialsusingtargeting
ligands[6-8].Antibodies(Abs)havebecomethemostwidelystudiedofthese
targetingligands[9],duetotheirhighspecificityandthevitalroletheyplayin
moderncancertherapeutics.
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ThemainchallengeforAb-targetedNPsisensuringthataswellas
demonstratingefficacytheNPhasanadequateplasmahalf-lifeandcan
specificallytargetcancercellswithatolerablesideeffectprofile.Thereare
controversiesoverwhichNPplatformscanbestoffertheseproperties,with
unansweredquestionsconcerningcellularinteractionanduptake,potential
toxicitiesandhowNPpropertiesaffectbehaviourbothin-vitroandin-vivo[10].A
robustunderstandingofthephysicalpropertiesofNPsisrequiredtoguide
researchintothecellularandphysiologicalinteractionsofNPs,generating
clinical-gradeproductsthatcouldbecomethenextgenerationofcancer
treatments.
ThisreviewaimstodefineandoutlinethedifferentNPplatformsthatare
currentlyusedinpre-clinicalandtranslationalcancermedicineandthe
challengesfacedintheAb-functionalisationofNPs.Researchregardingthe
variousapproachesandadvantagesofferedbyAb-targetedNPsincancer
therapywillalsobediscussed.
1.1.NanoparticlesNPsaredefinedbytheCommissionoftheEuropeanUnionasanynaturally
occurringormanufacturedparticulatematerialeitherunbound,asanaggregate
oragglomerate,whereatleasthalfoftheparticleshaveoneormoreexternal
dimensionsbetween1and100nminsize[11].However,thereremainsno
formalconsensusonthedefinition[12]andwhilstNPsareoftendefinedbytheir
sizeitisgenerallyconsideredimportanttoalsodefinethembypropertiesnot
sharedwithotherparticleswiththesamechemicalcomposition[13].For
examplehighmobilityinthefreestate,largesurfaceareatovolumeratioand,in
somecases,thenanosize-dependentexhibitionofquantumeffectswhichcontrol
physicalpropertiesandNPbehaviour[14].
Thiswideanddiverserangeoffunctionalpropertiesisfacilitatedbythevariety
ofmaterialsavailabletosynthesiseNPs.Theresultingversatilityoffersattractive
translationalpotentialforanumberofbiomedicalapplicationssuchastargeted
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deliveryoftreatment,innovativeimagingtechniquesandnoveltherapeuticsfor
hyperthermia.Table1showsthefavourablephysicalcharacteristicsoftheNP
treatmentplatformsdiscussedinthisreview.Giventhesecharacteristics,itisnot
surprisingthatoverthelastdecadenanoparticleshavereceivedincreasing
attentionfortheirexcitingpotentialinboththediagnosisandtreatmentof
cancer.
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Nanoparticle Size FavourablePhysicalCharacteristics
NATURALPOLYMERS*(Albumin)[15] 50-300nm
Biocompatible,biodegradable,non-toxicandnon-immunogenic.Straightforwardtocross-linkandchemicallymodify,drugsreadilyincorporatedintoalbuminpolymermatrix.
SYNTHETICPOLYMERS*(Poly-lacticoglycolicacid(PLGA)[16] 50-300nm
FDAapprovedsyntheticmaterialthatisnon-toxicandundergoeshydrolysisin-vivotoproducebiodegradablemetabolites.Drugseasilyincorporatedintothematrix.
LIPOSOMES*[17] 20–1000nm
Wellestablishedascancertherapeutics,canencapsulatehydrophilicandlipophillicdrugs,soluble,lowuptakebymacrophages,favourablestabilityin-vivo,andascolloids,protectdrugsfrombreakdown.
MICELLES*[18] 10-100nm
Uniquecore-shellarchitecture,hydrophobiccoreactsasnaturalcarrierenvironment,hydrophilicshellenablesstabilityinaqueoussolution,structuralmodificationscanfurtheraugmenttumourcelluptake
DENDRIMERS*[19,20] 5–20nm
Symmetricalbranchedpolymericmacromoleculeswithacentralcorealloweitherencapsulationorconjugationoftherapeuticagent.Self-assembling,polyvalent,chemicallystable,non-toxicandsoluble.
IRON-OXIDENANOPARTICLES*(Superparamagneticiron-oxidenanoparticles(SPIONs))[21] 10-100nm
Biocompatibleandbiodegradable,establishedclinicaluseasmagneticresonanceimaging(MRI)contrastagents,controllablebyanexternallyappliedmagneticfield,diverseformulationsallowfine-tuningofphysicochemicalproperties
SILICA(PorousSilicaNanoparticles(pSiNPs))[22] 50-1000nm
Goodchemicalandthermalstability,largesurfaceareaandporevolume.Canencapsulatelargeamountsofbioactivemoleculeandpromotecontrolleddrugrelease.Alsoofferssimplesurfacefunctionalisation
GOLDNANOPARTICLES*(Goldnanoparticles(AuNPs),Goldnanorods)[23] 1-100nm
Intenselightabsorptionandhighphotothermalconversionrate,andeaseofsynthesisinavarietyofshapesandsizes.Goodbiocompatibility,colloidalstabilityandsimpleligandconjugationchemistry.
CARBONNANOPARTICLES(Carbonnanotubes(CNTs))[24] 1-10nm
Highcarryingcapacity,andhighpropensitytotraversecellmembranesEasilychemicallymodifiedorfunctionalizedthroughformationofstablecovalentbonds.
QUANTUMDOTS(QDs)[25] 2-10nm
Broadabsorptionspectraandhighfluorescencequantumyield,highphotostability.Possibilityofmultiplexing(differentcoloursofQDsusedwithinoneassaywithasingleexcitationsource).CanbecombinedwithotherNPplatformsforcancertheranostics.
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Table1.PhysicalcharacteristicsofthecommonlyusedNPplatformsexploitedforclinicalapplications.NPslabelledwithanasterix(*)havebeenusedintheclinicforthediagnosisortreatmentofcancer[26].1.2.TargetingNanoparticlesforCancerTreatmentTheabnormalhyperpermeablevasculatureandimpairedlymphaticdrainage
foundwithintumours[27]canleadtopassiveaccumulationofNPswithin
malignanttissues.Thisprocessisknownasenhancedpermeabilityandretention
(EPR).EPRhasbeendemonstratedextensivelyinpre-clinicalresearchandinthe
clinicalsettingusingliposomaldrugssuchaspegylatedliposomaldoxorubicin
(PLD).ThesuccessofPLDliesinencapsulatingthetoxicanthracycline
doxorubicinwithin100nmliposomestolimitdruguptakeintosensitivehealthy
tissuessuchasthemyocardiumtherebyavoidingtheseveredose-limiting
cardiotoxicitycommonlyseenwithfreedoxorubicin[28].Oncelocalisedwithin
thetumour,liposomebreakdownreleasesthedoxorubicin,causinglocaltumour
celldeath.
WhilstEPRcanincreaseoveralltumouruptakeofNPs,specifictargetingofNPs
tocancercellscanbebestachievedthroughtheadditionofatargetingmoiety
[29],anumberofwhichhaveshownpromiseincludingproteins,nucleicacids
andsmallmolecules(Figure1).
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Figure1.SchematicshowingthemostcommonlyusedligandstoactivelytargetNPsincludingDNAaptamers(A),antibodies;asbothwholeimmunoglobulinGmolecules(IgGs)(B)andfragmentssuchassinglechainvariablefragments(scFv)(C),non-immunoglobulinproteinssuchastransferrin(D)andsmallmoleculessuchasfolicacid(E).Figureadaptedfrom[30].Antibodies,themostwidelyusedligandsforNPtargeting,recogniseavastarray
ofantigens,duetotheiruniquespecificityconferredbythecomplementarity
determiningregions(CDRs)(Figure2).Antibodiescanbereadilygeneratedas
intactIgGsorasfragmentssuchassinglechainvariablefragments(scFvs)
(Figure2)indefined,recombinantform[31,32].Antibodytargetingappearsto
becomplementarytopassiveintratumouralaccumulationmediatedbyEPR[33-
35].Ab-NPsalsoshowimprovedefficacyandcytotoxicitycomparedwithnon-
targetedNPs[36].
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Figure2.ImmunoglobulinG(IgG)Structure:Theheavychain(blue)ismadeupofonevariableregion(VH)andthreeconstantregions(CH1,CH2,CH3).Thelightchain(green)hasonevariable(VL)andoneconstant(CL)region.Oneachvariablechaintherearethreecomplementaritydeterminingregions(CDRs),whichbestowthevastvariabilityinspecificantigenbindingcapacity.VHandVLtogetherwithCH1andCLareknownastheFab(fragment,antigenbinding),andtheremainderofthemoleculeisknownastheFcregion(fragment,crystallisable),whichconfersbiologicalactivityandhalf-life.Recombinantantibodyfragmentsincludesinglechainvariablefragments(scFvs),whicharemadeupoftheVHandVLsegmentsjoinedusingapeptidelinker.2.0Antibody-Nanoparticlebio-conjugationThechallengesinestablishingconjugationstrategiesthatpreservethe
functionalityofboththeantibodyandNPduringtheconjugationprocesscanbe
broadlydividedintothreemaincategories;controllability,stabilityand
reliability.Controllabilityrelatestothenumberofantibodiesconjugatedtoeach
NPandtheorientationofthisinteraction,bothofwhichcangreatlyaffectthe
subsequentfunctionalabilityofanAb-NPconjugate[37].Whilstitremains
difficulttopre-determinethenumberofAbsconjugatedtoeachNP,itispossible
tofractionatefunctionalisedNPsbasedonthenumberoflinkermoleculesonthe
NPsurface[38].Sophisticatedtechniqueshavealsobeendevelopedtocontrol
antibodyorientation[39,40],resultinginpreservationofAbbioactivity[41].
ThesenewtechniqueshavebeenreportedtoachieveAbbioactivityashighas
88%ofthenakedantibody[41,42].
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ItiscruciallyimportanttomaintainstabilityoftheNPandAbthroughoutthe
conjugationprocessandtoensurethatthefinalconjugatehasacceptable
stabilityin-vitroandin-vivo.Harshreactionconditionsshouldbeavoided,as
thesecantriggerantibodyunfoldingwhichimpairsantigenbinding[43].Itis
alsonecessarythattheattachmentofAbtoNPsurfacedoesnotleadtounfolding
fromunfavourableinteractionsoftheproteinsidechainswiththeNPsurface
[44].Thechoiceofconjugationstrategyhasalsobeenshowntoaffectlong-term
stability.Forexample,whilstthehighlyspecificinteractionbetweenstreptavidin
andbiotinadaptormolecules(engineeredontothenanoparticlesurfaceand
antibodyrespectively)successfullygenerateAb-NPconjugatescapableof
targetingtumourcellsin-vitro[45],theseconjugatesdidnotdemonstrate
acceptablelong-termcolloidalstability.However,byaddingareactivethiol
grouptotheAbandcovalentlycross-linkingitviaabifunctionallinkermolecule
tothenanoparticlesurfacehigherbindingefficiencyandimprovedstabilitywas
achievedwithequalin-vitrotargetingefficacy[46].
Reliable,reproducibleandwellvalidatedconjugationproceduresareessentialto
enableupscaledevelopmentforuseinclinicalapplications[47]andtodate,this
hasbeenmostsuccessfullyachievedbydirectcovalentconjugationvialinker
molecules.Establishedcovalentconjugationmethodsincludecarbodiimide
coupling,maleimidecouplingandclickchemistry(coppercatalysedalkyne-azide
cycloadditionreactions)[48],asshowninfigure3.InthecaseofAuNPsgold-
sulphurlinkagecanbeexploited,utilisingthehighaffinitywithwhichthiol
modifiedligandsbindtogoldsurfaces[49].Theadvantagesanddisadvantagesof
conjugationstrategiesaresummarisedintable2,andhavebeenreviewedin
depthbyMontenegroet.al.[50].
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Figure3.Establishedmethodsforthecovalentconjugationofantibodiesto
nanoparticles:(A)carbodiimidecoupling,(B)maleimidecouplingand(C)
Coppercatalysed‘click’cycloadditionreaction[48].
ConjugationStrategy
Advantages Disadvantages
PhysicalAdsorption(ionic,electrostatic,van-der-Waals)
• Simpleandstraightforward• UnnecessarytomodifyeithertheAbortheNP
• Electrostaticattractioncanorientatetheantibodies‘endon’preservingbindingability
• Reversible• HydrophobicinteractioncancausedenaturationofAb
• ElectrostaticattractionisweakandpHdependent
• Competitivedisplacementbyserumproteinscanoccur
CovalentConjugation(includingvialinkermolecules)
• Higherstabilityandimprovedreproducibility
• ModificationstoAbusuallynotrequired
• Orientedbindingpossible• Useoflinker-moleculecanavoidhostilereactionconditions
• Possibletocontrolvalency
• Reactionconditionsmayleadtoproteinunfolding/reduction
• Conditionscanaffectantigenbindingcapacity
• Choiceofbindingmoietiesorlinkercansignificantlyaffectfunction
Useofadaptormolecules(biotin/streptavidin)
• Usuallyorientatedbinding• Canresistharshreactionconditions
• Difficulttocontrolvalency• Expensivetechnique• Reversibleattachment
BispecificAntibodyConjugation[51]
• Noneedfordisruptivechemicalconjugation
• MinimalimpactuponNPstability
• Longtermstabilityunknown• Onlyoneantigenbindingsiteavailable
Table2.SomeadvantagesanddisadvantagesofAb-NPbio-conjugationstrategies.Tableadaptedfrom[50,51].
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3.0ClinicalApplicationsofNanoparticlesforCancerTherapyAnumberofuntargetedNPshaveenteredinearlyphaseclinicalcancertrials
[26].Theseincludecyclodextrin-polymercamptothecin-containingNPsfor
advancedovariancancer[52],micellesdeliveringpaclitaxeltopatientswith
advancedgastriccancer[53]andAuNPsdeliveringrecombinanthumanTNF-
alphatopatientswithadvancedsolidtumours[54].SuperparamagneticIron-
oxideNPs(SPIONs)havefoundclinicalapprovalforuseascontrastagentsin
cancerimaging[55,56]andhavealsobeensuccessfulinearlyphasetrialsin
cancerhyperthermia[57,58].SomeuntargetedNPshavebeenfullyapprovedfor
clinicaluseascancertherapeutics[26]includingtheliposomaldrugPLD[59],
andnanoparticlealbumin-boundpaclitaxel(nab-paclitaxel)[60].
Ab-targetedNPsintheclinicareshowingearlypromisebuthavebeensofar
beenrestrictedtoliposomes.Inonetrial,ascFvagainsttransferrinreceptorwas
usedtotargetliposomesbearingp53DNAinpatientswithavarietyofadvanced
solidtumours[61].Doserelatedaccumulationofthetransgenewasobservedin
tumoursbutnotinnormalskintissue.Furthermore,7outof11patientshad
stablediseaseafter6weeksoftreatmentwithtolerablesideeffects[61].PLDhas
alsobeentargetedtotheFDA-approvedchimericanti-epidermalgrowthfactor
receptor(EGFR)mAbcetuximabinaphaseIclinicaltrialwithsomeevidenceof
clinicalefficacy.Furthermore,thesideeffectsseenwerelessseverethanwould
beexpectedwitheitherfreedoxorubicinorcetuximab[6].Themostcommon
dose-limitingtoxicitywasmyelosuppression,whichcouldbemanaged
prophylacticallywithgranulocyte-colonystimulatingfactor(G-CSF)[6].
TargetedPLDhasalsobeeninvestigatedintheclinicusingananti-human
epidermalgrowthfactorreceptor2(HER-2)scFv[7].Theseearlyclinicaltrials
areencouragingandfurthertranslationofAb-targetedNPsisunderpinnedbya
numberofexcitingpreclinicalstudies.Theongoingpre-clinicaldevelopmentof
Ab-targetedNPsfordeliveryofcytotoxicdrugs,radiotherapyandnucleicacids
andtheirroleincancertheranosticsandcancerhyperthermiaisdiscussed
below.
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4.0PreclinicalDevelopmentofantibody-targetednanoparticlesforcancertherapy4.1DeliveryofcytotoxicagentsAbtargetedNPdrugdeliveryoffersanumberofbenefitsoversystemic
administrationoffreedrugs.Theseincludeimprovedintratumouraldrug
distribution,controlledreleaseofdrugswithinthetumourmicroenvironment,
superiorefficacyandmoretolerablesideeffects.Althoughantibody-targeted
liposomes(immunoliposomes)remainintheleadfordeliveryofcytotoxicagents
[62],otherinnovativeapproachesareclosebehindusingwholeAbsaswellasAb
fragments.Thisisillustratedbythevarietyofdrug-loadedNPsinpreclinical
development(Table3).
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Nanoparticle Antibody/Fragment
MolecularTarget
CytotoxicAgent
Stage Cancer Reference
PEGylatedPLGAPolymer
mAb DR-5 Camptothecin In-vivo Colorectal [63]
Liposome mAb GD2 Etoposide In-vitro Mixed(GD2expressing)
[64]
Lipid-PolymerHybrid
mAb EGFR Adriamycin In-vivo HepatocellularCarcinoma(HCC)
[65,66]
PLGAPolymer mAb EGFR Paclitaxel In-vivo Lung [36,67]PorousSilicaNPs(pSiNPs)
mAb p75(NTR),EGFR,CD20
Camptothecin In-vitro Glioblastoma,Neuroblastoma,B-celllymphoma
[68]
Polymericnano-micelles
mAb HIF-1α Paclitaxel In-vitro Gastric [69]
CarbonNanotube(CNT)
mAb CEA Doxorubicin In-vitro Colon [70]
AuNPs mAb EGFR Gemcitabine In-vitro Pancreatic [71]PLGA/MMTPolymer
mAb HER2 Paclitaxel In-vitro Breast [72]
SPIONs mAb HER2 Paclitaxel/Rapamicin
In-vitro Breast [73]
Liposome mAb 2C5 Doxorubicin In-vivo Glioma,Lung [74,75]PLAPolymer mAb SM5-1
bindingprotein
5-FU In-vivo HCC [76]
Lipid-PolymerHybrid(PLGA)
Half-Ab(hAb)
CEA Paclitaxel In-vitro Pancreatic [77]
Iron-Oxide(SPION) ScFv Endoglin Docetaxel In-vitro Ovarian [66]PEGylatedPolymer ScFv CD44v6 Arsenic
trioxideIn-vivo Pancreatic [67]
PLGAPolymer ScFv SM5-1bindingprotein
Paclitaxel In-vitro HCC [78]
Liposome ScFv HER-2 Doxorubicin PhaseIClinicalTrial
Breast [7]
Liposome ScFv c-Met Doxorubicin In-vivo Lung [79]PLGA Fab HER2 Pseudomonas
ExotoxinA(PE38KDEL)
In-vivo Breast [80]
Liposome Fab EGFR Doxorubicin PhaseIClinicalTrial
Mixed(EGFRexpressing)TumourTypes
[6,71]
Table3.Drug-loadedNPstargetedusingantibodiesorantibodyfragments.WholeAbs:ThemajorityofwholeAbtargetedNPshavebeendirectedtoEGFRor
HER2;bothofwhichhaveillustratedtheprospectofNP-mediatedintracellular
drugdeliverythroughreceptormediatedinternalisation.Oneexampleused
cetuximabtotargetgemcitabine-loadedAuNPs[71].Intheseexperiments,the
authorsdemonstratedspecifictargetingoftheNPstoEGFRexpressing
pancreaticcelllinesin-vitroandadditionalantibody-mediatedcytotoxicity
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comparedtonon-targetedcounterparts[71].TargetingEGFRhasalsoshownin-
vivoefficacyinarangeofmodels.Forexample,indeliveryofpaclitaxel-loaded
polymericNPstolungtumours[36],camptothecinloadedporoussilicaNPstoa
rangeofEGFRover-expressingtumoursincludingglioblastoma[68]and
adriamycinencapsulatedpolymer-lipidNPstohepatocellularcarcinoma[65].In
thisfinalexample,therewasalsoevidencesuggestingthatboththeantibodyand
adriamycinwerecontributingtotheobservedcytotoxicity[65].HER2targeting
usingtrastuzumabhasbeenshowntobeeffectiveusingpaclitaxelandrapamicin
partitionedwithinpolymercoatedSPIONs;In-vitroefficacywasover7times
higherthanthenon-targetedcounterparts[73].Inanotherstudy,HER2-targeted
polymericNPsdeliveringpaclitaxelshowedsustaineddrugreleasein-vitro[72].
PolymericNPshavealsobeensuccessfullytargetedtothepro-apoptoticcell
surfacedeath-receptor,DR5[63].Intheseexperiments,theDR5-targeted
polymericNPswereshowntoinitiateapoptosisin-vitroand,whencaptothecin
wasencapsulated,theNPwasabletoovercometheresistancecommonlyseen
withsingleagentanti-DR5therapyin-vivo.Inanotherexample,hypoxia
induciblefactor1α(HIF-1α),anextracellularproteinoverexpressedina
numberofhumancancersinresponsetolocalhypoxia,wassuccessfullytargeted
withanti-HIF-1αAbfunctionalisedpaclitaxelloadedpolymericNPs[69].The
NPswereshowntoselectivelyinternaliseincellsoverexpressingHIF-1αand
paclitaxelmediatedcytotoxicitywasshowntobespecifictowardsHIF-1α
expressingcells.Othertargetsinvestigatedforcontrolleddeliveryofdrug-loaded
NPsincludethecellsurfaceglycoproteinandtumourmarkercarcinoembryonic
antigen(CEA)andthep75neurotrophinreceptor(NTR)[68].NTRisamember
ofthetumournecrosisfactor(TNF)superfamily,whichhasaroleincelldeath
andisoverexpressedinanumberofmalignanciesincludingsarcomaand
malignantmelanoma[81].Drug-loadedNPshavealsobeentargetedusingSM5-
1,amouse-humanchimericantibodywhichishighlyspecifictotheSM5-1
bindingprotein,atargetknowntobeoverexpressedinanumberofcancers
includinghepatocellularcarcinoma(HCC),melanomaandbreastcancer.Binding
oftheSM5-1Abtoitstargetproteininhibitscellgrowthandinducesapoptosisin
cancercellsinacaspasedependentmanner[82].PolymericNPsloadedwith5-
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Flurouracil(5-FU)havebeensuccessfullytargetedusinghumanisedSM5-1inits
wholeAbform,demonstratingsustaineddrugreleaseandfavourableanti-
tumouractivityagainstsubcutaneousandorthotopicHCCxenografts[76].
PLDhasbeenshowntoexhibitenhancedtumourretention,Abdependent
endocytosisandincreasedcytotoxicitywhentargetedusingtheanti-nucleosome
mAb2C5inorthotopicgliomatumoursin-vivo[74].The2C5Abspecifically
recognisesextracellularandtumour-cellboundnucleosomesthatarisefrom
apoptotictumourcellsin-vivo[75].Evenwhenusedatsubtherapeuticquantities,
2C5isaneffectivetumourtargetingmoietyforPLD,alsoshowingefficacy
againstprimaryandmetastaticlungtumoursinmice[75].Centralnervous
system(CNS)tumourspresentauniquechallengetotreatmentduetothe
presenceofthebloodbrainbarrier(BBB);auniquephysiologicalbarrierwhich
functionstoprotectthebrainbutalsolimitsdrugdeliverytotheCNS[83].To
facilitatemovementpasttheBBB,polymeric,2C5-targetedNPshavealsobeen
co-targetedusingananti-transferrinreceptor(TfR)Ab.In-vivointratumoural
localisationwasfoundtobesignificantlyhigherforthedualtargetedNPthan
thatseenforNPstargetedwitheachantibodyalone[84].Morerecently,
transferrinreceptorbispecificantibodyplatformshavebeenshownto
successfullycrossandtargetpasttheBBB[85],andthesebispecificAbscould
equipdrugloadedNPstomoreeffectivelytargetCNStumours.These
experimentssuggestthatAbtargetedNPscouldofferanovelsolutiontodeliver
treatmentpasttheBBB.
AbFragments:Abfragmentsareconsideredtomakeattractivetargetingagents
thankstoasmallersizeandpotentiallyreducedimmunogenicitycomparedwith
wholeAbs[86].Themostcommonlyusedfragments,scFvs,arereadily
generatedusingrecombinantantibodytechnologyandhavebeensuccessfully
appliedinthefollowingexamples.ScFvshavebeensuccessfullyexploitedto
targetarsenicloadedpolymericnanoparticlestargetedtoCD44v6,a
transmembraneglycoproteinoverexpressedonpancreaticadenocarcinoma
cells.Accumulationandretentionwithinthetumourwassignificantlyhigherfor
CD44v6-targetedNPsthanforthenon-targetedcontrols.Furthermore,the
targetedNPsinhibitedtumourgrowthsignificantlymorethanthenon-targeted
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NPsorfreearsenic[67].Inanotherexample,phagedisplaytechnologywasused
toobtainacell-internalisingscFvtotargetc-Met;atyrosinekinasereceptor
expressedonbothtumourandendothelialcells[79].Whenthisanti-c-MetscFv
wasusedtotargetPLDtolungtumours,significantdisruptionofthetumour
vasculaturewasobservedinadditiontodirecttumourcellcytotoxicity[79].
Duetotheirmagneticproperties,ithaslongbeenproposedthatSPIONscouldbe
targetedusinganexternallyappliedmagneticfield[87].However,following
successfulinitialin-vivostudies,problemswithbloodvesselembolisationledto
thedevelopmentofalternativemethodsoftargeting,includingAbs[88].More
recently,thepossibilityofmagnetictargetinghasbeenrevisitedincombination
withanscFvtargetedtoendoglin,acellsurfaceglycoproteinoverexpressedon
ovariancancerendothelialcells[66].ThescFvswereconjugatedtothesurfaceof
ironoxidenanoparticlespre-loadedwithβ-cyclodextrinencapsulateddocetaxel
tocreatedrug-loaded,endoglintargetedSPIONs.In-vitrostudiesshowedthatthe
scFvdirectedspecificbindingontoendoglinpositivecellsandalsothatthe
majorityoftheSPIONswerelocalisedwithinthemagneticfield[66].
ProgresshasalsobeenmadeusingFab-targetedNPs.Forexample,Fab
fragmentsderivedfromahumanisedanti-HER2mAbhavebeensuccessfully
usedtotargetpseudomonasexotoxinA(PE)encapsulatedwithinPLGANPs[80].
PEisapotentimmunotoxinwithsignificanttoxicity,whichhassofarlimitedits
useasananti-cancerdrugdespiteevidenceofimpressiveanti-tumouractivity
[89].Theresultsfromtheseexperimentsdemonstratedimprovedanti-tumour
activityandreducedsystemictoxicitywhencomparedtothenon-encapsulated
anti-HER2-PEAb-drugconjugate(ADC).
4.2DeliveryofradiationtreatmentRadiationistheprimarytreatmentmodalityinthemanagementofmany
cancers.Themainchallengeinradiotherapyishowtoachieveadequate
radiationexposuretothetumourwhilstavoidingdamagetosurrounding
healthyareas,especiallyinsensitivetissuessuchasthebrain.Ab-targetedNPs
haveshownpromiseinaddressingthischallengeforexamplebyfacilitating
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neutroncapturetherapy(NCT);atechniquewhichreliesontheintratumoural
injectionofanon-radioactiveisotope,mostcommonlyBoron-10(10B),which
actsasacaptureagentandreleaseslocalisedionisingradiation[90].In-vitro
studieshavetargeted10B-loadedliposomestoEGFR-expressinggliomacellsin-
vitro[91],raisingthepossibilitythatAb-targetedNPscouldovercomethe
barrierofdirectintratumouralinjectionofcaptureagents.Inasecondexample,
cetuximab-conjugatedSPIONswerefoundtosensitiseradioresistantgliomasto
theeffectsofradiation,possiblythroughproductionofreactiveoxygenspecies
andtheinhibitionofDNAdamagerepair[92].
4.3.Deliveryofnucleicacids
Ab-targetedNPdeliveryvectorssuchaspolymericNPs[93],AuNPs[94]and
liposomesarealsobeginningtofindaroleindeliveryofnucleicacidsforcancer
therapy.ThisprovidesopportunityforthedeliveryofDNAtoinduceexpression
oftherapeutictumoursuppressorgenes,ordeliveryofsmallinterferingRNA
(siRNA)oranti-senseDNAsequenceswhichcandisruptthetranslationof
oncogenes[95].EncapsulationofnucleicacidinNPsaimstoovercomebarriers
suchasphysiologicalinstability,poorintracellulardeliveryandoff-targeteffects
[96].Forexample,inonestudy,polyethyleniminewasusedtocondenseand
compactDNAencodingthep53gene[97]generatingpolymer-DNANPswhich
werethentargetedusingJ591,ananti-PSMAmAb.TheresultantJ591targeted
NPsshowedefficienttransfectioninvariousprostatecancercelllinesin-vitro
andeffectivePSMAspecifictargetingwhentestedin-vivoinprostatecancer
xenografts[97].J591isparticularlyfavourableforclinicaltranslationduetoits
previoususeinaphaseIIclinicaltrial[98].
Fordeliveryofanti-senseDNA,AuNPshavebeencoatedwithahighlyorganised
anti-sensenucleicacidlayerdesignedtobindHER2mRNA.Correspondingsense
DNAsequenceswerefunctionalisedviaclickchemistrytoanti-HER2Absand
hybridisedtotheanti-senseDNAontheNPsurface.TheseNPshadsuperior
uptakeintoHER2positivecellsin-vitrocomparedtonon-targetedcounterparts.
Inaddition,aconcentrationof1nManti-HER2NPswassufficienttocompletely
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blockexpressionofHER2intargetcellswhilstcontrolNPseithercontaining
scrambledanti-sensenucleicacidsorfunctionalisedusinganoff-targetAb
controlwereunabletoinhibitanyexpression[99].Asimilarapproachwas
undertakenusingclickchemistrytoconjugatechemicallystabilisedantisense
nucleicacidsandtrastuzumabtothesurfaceofpolymericmicelles.Thesedual
functionalisedpolymericNPsdemonstratedHER2targeteddeliveryandefficient
geneknockdowninHER2positivecellsin-vitrowhencomparedwithnon-
targeted,non-stabilisedcontrols[100].
Anotherexampleofefficientnucleicaciddeliveryutilisedaliposomal
formulationofantisensenucleicacidsagainstBCL2,anantiapoptoticgene
overexpressedinthemajorityofacuteleukaemias.Inthisstudy,theliposome
wastargetedusingtheanti-CD20mAbrituximab[101].Thetargetedliposomes
demonstratedeffectivereductionsofBcl-2proteinwithincancercells.Further
in-vivostudiesusinghumanBurkitt’slymphomaxenograftsshowedsignificant
reductionsintumourgrowth.Furthertothis,theliposomeswerefoundtobe
stableforover1yearinstoragewhetherasasuspensionorlyophilisedpowder,
highlightingfeasibilityforclinicaltranslation[101].
4.4.Antibody-conjugatednanoparticlesastheranosticagents:
NPsofferanattractiveplatformfordevelopmentofso-calledtheranosticagents,
whichcombinetherapyanddiagnosiswiththeaimofreducingtheneedfor
multi-stepproceduresandavoidingtreatmentdelays[102].Antibodytargeting
hasamajorroletoplayinthisfield(Figure4),andSPIONshavebecomethe
mostadvancedtheranosticNPsusingarangeofinnovativeapproaches.One
successfulexampleofAb-SPIONtheranosisusedthecommercialanti-EGFRmAb
cetuximab,conjugatedtoPEG-polymercoatedSPIONsthroughcarbodiimide
chemistry,totargetEGFRandthemutantvariantEGFRvIIIexpressingorthotopic
gliomatumours.ThemAb-functionalisedSPIONstriggeredapoptosisinhuman
gliomacellsin-vitroandconferredasignificantsurvivaladvantageoverboth
cetuximabandSPIONsalonewheninfusedintoorthotopichumanGBMmodels
[103].Intheseinnovativestudies,thechallengeofcrossingtheBBBtoachieve
18
adequateintratumouraldistributionwasaddressedusingconvection-enhanced
delivery(CED).PotentinhibitionofEGFRphosphorylationandincreasedlevels
ofcaspasecleavagewereobservedalongwithconsistentcontrastenhancement
oftheSPIONsonT2-weightedMR.Inadditiontothispromisingexperimental
data,toxicityexperimentsinhealthymiceshowednoevidenceofdamageto
normalbraintissues[103].
Targetingchemotherapyincombinationwithimagingagentsdemonstrates
anothertheranosticapproachthathasbeenexemplifiedusinghybrid
superparamagneticiron-platinumNPs(SIPPs)targetedtotheprostatecancer
targetPMSA.TheSIPPswereencapsulatedwithpaclitaxelinsidephospholipid
micellesandthenconjugatedusingstreptavidin/biotinadaptormoleculesto
anti-PMSAAbs.Theresultinghybrid-NPswereshowntotargetPMSApositive
prostatecells,inducecytotoxicitysimilartofreepaclitaxelin-vitroandfunction
asMRIcontrastagentsin-vivo[104].
Iron-oxidemagneticnano-crystals(MNCs)havealsobeeninvestigatedfor
theranosticpotential.TheseparticlesretaintheimagingabilitiesofSPIONsbut
aresmalleranddonothavesurfacecoatings.Assuch,theyhavetheadvantageof
beingreadilyincorporatedintothestructureoforganicpolymericNPsto
generatemultifunctionalNPs.Inoneexample,doxorubicinandMNCloaded
PLGApolymernanoparticleswereconjugatedviacarbodiimidecouplingtothe
anti-HER2mAbtrastuzumabformingNPswiththepotentialtobothdetectand
treatmalignantbreastcancercells.In-vitroresultswiththeseNPsdemonstrated
excellentefficacyasMRprobeswithspecificityforHER2positivecells.Aswellas
this,theNPsshowedsustainedreleaseofdoxorubicin[105].IronoxideMNCs
havealsobeensuccessfullyemployedastheimagingcomponentofdocetaxel
loadedPLGANPstargetedusingascFvagainstprostatestemcellantigen(PSCA).
PSCApositivecellsincubatedwiththetargetedNPsappeareddarkeronT2-
weightedMRimagingcomparedwithnon-targetedequivalents[106].Unusually
inthisstudy,drugreleaseshowedinitialrapiddesorptionofdrugfromthe
particlesurface,followedbymoresustainedreleaseduringdegradationofthe
polymermatrix.In-vitroefficacystudiesindicatedthatthedocetaxel-loadedNPs
19
wereeffectiveatalowerdosethanfreedocetaxel.PolymericPLGAnanoparticles
havealsobeenusedtoprovideavehiclefortheranosisbycombining
doxorubicinwithindocyaninegreen(ICG);aheat-generatingnear-infrared(NIR)
dye[107].Whenconjugatedtoanti-HER-2Absusingcarbodiimidechemistry,
theNPsdemonstratedtargetedcelluptake.Furthermore,NIRlaserexcitationof
theICGreleasedcytotoxicheatin-vitro.Theseresultsshowcaseasingle
theranosticNPwithtriplefunctionsin(i)imaging,(ii)deliveryofchemotherapy,
and(iii)inductionoflocalhyperthermia[107].
TheranosticAb-targetedlipidbasedNPshavealsobeencreatedusingquantum
dots(QDs)astheimagingcomponent,rapamycinfortherapyandtrastuzumab
fortargeting.TheresultingNPstargetedandimagedHER2positivecellsin-vitro,
showninboth2Dand3Dmodelsusingconfocalmicroscopy.Inthisstudy,lipid-
encapsulationaloneappearedtoimparta2foldincreaseineffectivenessover
freerapamycin,whilsttheadditionofaHER2targetingAbimpartedafurther5
foldincreaseinefficacyinHER2positivecells[108].
Multi-walledcarbonnanotubes(CNTs)offernewpotentialascancer
theranostics,duetotheirhighcarryingcapacityandhyperechogenicity
(increasedultrasoundcontrast).Preliminaryresultsinvestigatingthe
theranosticapplicationofCNTsconjugatedwithAbagainstPSCAhavebeen
encouraging.WhenfunctionalisedwithAbs,theCNTsshowedefficacyas
contrastagentsandenhanceduptakeintoPSCA-expressingcells.Moreover,after
loadingwithdoxorubicin,theAb-functionalisedCNTsaccumulatedwithin
tumourtissuesandinhibitedtumourgrowthin-vivo[109].
20
Figure4.Schematicillustratingthe4majorcomponentsofanAb-targetedtheranosticNP:(1)Deliveryvehicle,(2)Imagingagent,(3)Therapeuticcomponentand(4)TargetingAb.Figureadaptedfrom[110].4.5.CancerhyperthermiaAlmost50yearsagoitwasshownthattemperaturesofjust42°Ccouldinduce
cancercelldamagewithinarelativelyshorttimeframewhilstnon-malignant
cellswereabletowithstandthisriseintemperature[111].Morerecentwork
hasshownthatthehypoxictumourmicroenvironmentandtheincreasedmitotic
rateofmalignantcellspredisposemalignantcellstotemperaturesensitivity
[112],andthathyperthermiatreatmentshowssynergismwithconventional
therapies[113].Whilstpotentiallyaneffectivetreatment,themajorchallenge
fortherapeuticapplicationofhyperthermiaisrestrictingtreatmenttodiseased
areasandavoidingdamagetohealthytissues.Heat-generatingNPsofferan
elegantsolutiontothisproblem,particularlyiftheNPscanbelocalisedwithin
tumourspriortoheatinduction.Localisationhasbeenachievedbydirect
injectionintotumourtissue,butincreasinglyantibodyfunctionalisationisbeing
utilisedtotargetNPstothetissueofinterest(Table5),asillustratedwiththe
examplesbelow.
21
Nanoparticle Externalactivatingfield
Antibodytarget
Setting CancerTreated
Reference
SuperparamagneticIron-oxide
nanoparticles(SPIONs)
Alternatingmagneticfield(AMF)
MembraneProtein
In-vivo Breast [114,115]
GoldNanoparticles(AuNPs)
Laserlight EGFR In-vitro OralSquamousCellcarcinoma
[116]
Mucin-7 In-vitro UrothelialCells [117]Near-Infrared(NIR)light
TROP-2 In-vitro CervicalCancer [118]EGFR In-vitro OralSquamous
Cellcarcinoma[119]
Short-waveradiofrequency(RF)energy
EGFR In-vivo Pancreatic [120,121]
CarbonNanotubes(CNTs)
NIRlight CD133 In-vitro Glioblastoma [122]HER-2&IGF1R
In-vitro Breast [123]
CD22/CD25 In-vitro BurkittsLymphoma
[124]
Table4.Ab-targetedNPsemployedforcancerhyperthermia.
SPIONs:ArethemostclinicallyadvancedNPsforheattreatmentduetotheir
historyofuseasapprovedMRIcontrastagents[55]andinpioneeringclinical
trials(untargeted)forheattreatmentofglioblastoma[57].Whenexposedtoan
alternatingmagneticfield(AMF)SPIONsconvertmagneticenergytothermal
energythroughBrownianandNeelrelaxation.SuperparamagneticNPs,unlike
largerferri-orferromagneticNPs,donotretainmagnetismwhenthefieldis
removedandcangenerateheatatlowermagneticfieldamplitudes,makingthem
moreattractiveforbiomedicalapplications[125,126].
AntibodytargetingofSPIONsforhyperthermiawasfirstachievedbyconjugating
an111InradiolabelledmAbagainstanintegralmembraneglycoprotein(highly
expressedonanumberofhumancancers)to20nmdextran-coatediron-oxide
nanoparticles[115].FollowingintravenousinjectionoftheNPsintoinathymic
micebearingHBT3477(breast)xenografttumours,micewereexposedto
variouslevelsofexternalAMF.Electronmicrographstakentwodaysfollowing
AMFtherapyshowedtumourcellnecrosisatallstrengthsofAMFapplied.
22
TumoursdidnotrespondtoeitherSPIONsorAMFalone,andtoxicitywasonly
seenatthehigheststrengthofAMFapplied.Subsequentstatisticallyvalidated
resultsshowedthattherapeuticresponsesweregeneratedwithoutnormal
tissuetoxicityandthattheheatingdosecorrelatedwellwithresponse[114].
Despitein-vivosuccess,subsequentstudieshavemetwithchallengesduetothe
rapidclearanceofSPIONsfromcirculationviathereticuloendothelialsystem
(RES)wheninjectedintravenously[127].Whilstthischaracteristiclendsitself
welltoSPIONuseascontrastagents,itisdetrimentaltoachievingsufficient
intratumouralSPIONconcentrationstogeneratetherapeuticheating.Anumber
ofsolutionshavebeenproposedtoovercomethis,includingtheuseofblocking
agentstoprolongSPIONcirculatorytime[128],tailoringtheNPsurface
chemistriesorthetargetingmoietiestooptimisecellularinteractionsofSPIONs
[41,129]ordirectintratumouralinjectionwithtargetingantibodiesfunctioning
toretainSPIONsattheinjectionsite.
AuNPs:Ab-targetedAuNPshavealsobeenexploitedasvectorsforphotothermal
therapy(PTT).Thisprocessreliesuponthegenerationofvibrationalheatenergy
followingtheexcitationofphotosensitisersthroughtheabsorptionofspecific
wavelengthsoflight;mostcommonlyinthenear-infrared(NIR)range.AuNPs
stronglyabsorbandscatterlight;thisabsorptioncanbetunedbymodifications
inthesizeandshapeoftheNPsinadditiontotheincorporationofother
materialssuchassilica[116,119].ThelightabsorbedbyAuNPsconvertsreadily
andrapidlyintoheatandthis,alongwiththeestablishedbiocompatibilitymake
AuNPsattractivephotothermalagents[116].ToavoidaccumulationofAuNPsin
healthytissues,anumberofpassiveandactivetargetingstrategieshavebeen
testedtotargetthemtotumourcells.ThesestrategiesincludePEGylation,
liposomeencapsulationandantibodyconjugation[130].Thiol-terminatedPEG
derivativesarecommonlyusedtocoatthesurfaceofAuNPsactingtoimprove
colloidalstability,avoidtheRESuptakeofAuNPsandprovidea‘linker’to
conjugateAuNPstobiomoleculessuchasantibodies[131].RecentAb-targets
investigatedincludetrophoblastcellsurfaceantigen2(TROP2),a
transmembraneglycoproteinoverexpressedinanumberofepithelialcancers
andassociatedwithpoorprognosisincervicalcancer[118].Inthisstudy,hollow
23
goldnanosphereswereconjugatedtoanti-TROP2mAbsusingathiol-terminated
PEGlinker,generatingfunctionalisedNPsabletoreducecellviability
significantlymorethannon-targetedcontrolsfollowingexposuretoNIRlaser
[118].Inanotherstudy,goldnanospheresweretargetedtourothelialcancer
cellsusingantibodiesagainstMucin-7;acommonlyusedtargetinbladdercancer
[117],andactivatedusinggreenlaserlight.
AswellasabsorbingNIRorvisiblelight,Ab-targetedAuNPsalsoreleaseheat
followingabsorptionofshort-waveradiofrequency(RF)energy;behavingas
targetedradiofrequencyablation(RFA)agents.Short-waveRFfieldshavethe
advantageofpenetratingdeeperintotissuesthanNIRlight.Whentheanti-EGFR
antibodycetuximabwasconjugatedto20nmgoldnanorods,selectivetargeting
wasseenin-vitroand,followingexposuretoshort-waveRFfields,necrotic
cellularinjurywasachieved;similartotheeffectsofinvasiveRFAintheclinic
[120].WhenthesameNPsweretestedin-vivo,significantcelldeathwas
observed36hoursfollowingtreatmentofEGFRamplifiedpancreaticxenograft
tumours,withnodamageinselectedhealthytissuesincludingtheliver[121].
CNTs:NIRactivatedAb-targetedCNTshavealsobeenemployedtoinducePTT
in-vitro[132].ExamplesoftargetsinvestigatedincludeHER2andIGF1R,to
targetCNTstobreastcancercells[123],CD22andCD25totargetBurkitts
lymphoma[124]andCD133totargetglioma-likestemcellsinthetreatmentof
glioblastoma[122].Whilsttheseearlystudiesshowpromise,CNTsaccumulate
intheliver,arenon-biodegradable,andshowpoorsolubilitywithatendencyto
aggregate.Thesechallengesoftoxicitylimitthecurrentclinicaltranslationof
CNTs,andarebeingaddressed[133].
5.0Conclusions
ThisreviewhasillustratedthewaysinwhichthediversityandversatilityofNPs
canbefurtherexploitedbyAbtargeting.Awiderangeofpre-clinicalresearch
hasdemonstratedthatAb-targetedNPscanbeusedtodeliverycytotoxic
chemotherapydirectlytocancercells,resultinginefficacioustreatmentwith
24
reducedsideeffectsprofilescomparedtofreedrugs.Breakthroughshavealso
beenmadeinutilisingAb-NPstoaugmentradiotherapyandaidincancergene
therapy.CancertheranosticshasevolvedinparalleltoNPs,andanumberof
innovativeapproacheshaveshowcasedsingleagents,whichcancombinecancer
imagingandtherapy.Finally,theuseofhyperthermiainthetreatmentofcancers
hasbeenmadepossiblethankstotheinherentphysicalpropertiesofSPIONs,
AuNPsandCNTs.
Targetedtherapyremainsattheforefrontoftranslationalcancerresearchand
significantprogresshasbeenmadeintheclinicaltranslationofantibody-
targetedNPbasedcancertreatment,includingphaseIclinicaltrialsusing
antibodytargeteddrug-loadedliposomes.Thesepioneeringstudieshavebuilt
uponprevioussuccessesdevelopingNPs,refiningbio-conjugationstrategiesand
optimisingthebiocompatibilityandbioavailabilityoftheseproducts.
6.0FuturePerspectivesFutureresearchonAb-NPswilldependuponreliableandreproduciblebio-
conjugationstrategiesthatcanbescaleduptogoodmanufacturingpractice
(GMP)standardforclinicaltranslation[129].OtherNPssuchasmesoporous
silicaNPs(MSNs)(Table1)couldbeutilisedwhichofferimpressivedrug
carryingcapacity,andcanbeequippedwithtargetingligands[134].Inaddition,
multifunctionalhybridNPscanbegeneratedforuseincombinationtherapy
and/ortheranostics.
Theroleofimmunotherapyinthetreatmentofcancercontinuestoevolve.
WhilsttheuseofAb-targetedNPsincancerimmunotherapyisearlyin
development[135],recentbreakthroughsincludetheuseofPLGANPs
conjugatedtobothprotein-MHCcomplexesandanti-CD28mAbstoactas
syntheticantigenpresentingvehicleswhenadministeredin-vitro[136].Inthe
future,Ab-targetedNPscouldbeusedtoprimetheimmunesystemnotonlyto
recognisetumourantigensbutalsotostimulateanti-tumourimmunity.Whilst
Ab-targetedNPsareonlyjustreachingtheclinicalsetting,theirdiversepotential
25
foruseincancerdiagnosticsandtherapeuticspredictsanimportantroleinthe
futureofcancertreatment.
6.0ExecutiveSummaryIntroduction
• Targetedcancertreatmentofferssignificantadvantagesoverconventionaltherapybutcurrenttargetedtreatmentsareoftenlimitedbysystemictoxicity
• Nanoparticles(NPs)possessuniquephysical,chemicalandbiologicalpropertiesthatcouldovercometheselimitationsandbeharnessedforuseincancermedicine
Nanoparticles• NPsarenaturallyoccurringormanufacturedparticulatematerialeitherunbound,asan
aggregateoragglomeratewithonedimensionbetween1and100nminsize.• NPspossessinterrelatedproperties;highmobilityinthefreestate,largesurfaceareato
volumeratios,andsometimestheexhibitionofquantumeffects.• ThewideanddiverserangeoffunctionalpropertiesexhibitedbyNPsispossiblethanks
totheutilityofavarietyofmaterialstosynthesisethem.Targetingnanoparticlesforcancertreatment
• PassiveaccumulationofNPsoccurswithinmalignanttissuesinaprocessknownasenhancedpermeabilityandretention(EPR).
• SpecifictargetingofNPstocancercellscanbeachievedthroughtheadditionofatargetingmoiety,includingproteins,smallmoleculesandaptamers.
• Antibodies(Abs)arethemostpromisingtargetingligandsandcanbereadilygeneratedasintactIgGmoleculesorasfragments(ScFvs)indefined,recombinantform.
• AntibodytargetingiscomplementarytoEPRandenablesspecificreceptormediatedinternalizationoftheNP.
Antibody-Nanoparticlebio-conjugationstrategies• Challengesinbio-conjugationcanbebroadlydividedintothreemaincategories;(i)
controllability,(ii)stability,and(iii)reliability.• Sophisticatedconjugationtechniquesenablethecontrolofantibodyorientationsto
ensuretargetbindingandpreserveAbbioactivity• Whilstbothbifunctionallinkermoleculesandadaptormoleculescangeneratefunctional
conjugates,covalentlinkermoleculesdemonstratesuperiorlongtermstability.• Establishedcovalentconjugationstrategiesincludecarbodiimidecoupling,maleimide
couplingandclickchemistry(coppercatalyzedalkyne-azidecycloadditionreactions).ClinicalApplicationsofNanoparticlesforCancerTherapy
• NPbaseddrug-deliverysystemssuccessfullytranslatedintoearlyphaseclinicaltrialsincludeliposomes,polymericNPs,dendrimers,micellesandgoldNPs(AuNPs).
• SuperparamagneticIron-oxideNPs(SPIONs)havebeenclinicallyapprovedforuseascontrastagentsincancerimagingandhavebeensuccessfulinearlyphasetrialsforcancerhyperthermia
• Antibody-targetedliposomes(immunoliposomes)havebeenusedinthreeearlyphaseclinicaltrials.
PreclinicalDevelopmentofantibody-targetednanoparticlesforcancertherapy• ChemotherapyloadedAb-NPscanofferimprovedintratumouraldrugdelivery,superior
efficacycomparedwithfreedrugs,andcontrolledreleaseofdrugswithintumourcells.• Targetingpastthebloodbrainbarrier(BBB)totargetcentralnervoussystem(CNS)
tumoursmaybepossiblethroughdualtargetingoftransferrinreceptorandcancerspecifictargets.
• Ab-targetedpolymericNPs,AuNPsandliposomescanbeusedtostabilisenucleicacidsanddeliverthemdirectlytocancercellsforgenetherapy.
• SPIONsbecombinedwithotherNPsincludingpolymersandquantumdotstoproducemulti-functionaltheranosticNPsthatcanimagetumoursanddeliverycytotoxicdrugsincombination
• SPIONs,AuNPsandCNTsareabletogenerateheatfromwithinthetissueofinterestwhenactivatedbyexternallyappliedalternatingmagneticfields(SPIONs)ornear-infraredfields(AuNPs/CNTs).
26
• SPIONshavebeentargetedusingAbsinthepre-clinicalsettingwithsuccesslimitedbyrapiduptakebythereticuloendothelialsystemandclearancein-vivo.Strategiesarebeinginvestigatedtoreducethisuptake.
Conclusions• Ab-targetedNPsofferaversatileplatformforthedevelopmentofthenextgenerationof
cancertherapeutics• Progresshasbeenmadeintheclinicaltranslationofantibody-targetedNPbasedcancer
treatmentsFuturePerspectives
• Reliableandreproduciblebio-conjugationstrategiesthatcanbescaleduptogoodmanufacturingpractice(GMP)standardwillfacilitateclinicaltranslationofAb-targetedNPs.
• Ab-targetedNPsmayhaveafutureroleincancerimmunotherapy,includinginthedevelopmentofsyntheticantigen-presentingvehicles.
Financial&competinginterestsdisclosureTheauthorsaregratefulforthesupportreceivedfromtheSeventhFrameworkProgramme(FP7)forDARPintargetedmagnetichyperthermictherapyforglioblastoma(DartrixECGRANT:278580),andfromtheDepartmentofHealthandCancerResearchUKExperimentalCancerMedicineCentre(ECMC).Theauthorshavenootherrelevantaffiliationsoffinancialinvolvementwithanyorganizationorentitywithfinancialinterestinorfinancialconflictwiththesubjectmatterormaterialsdiscussedinthemanuscriptapartfromthosedisclosed.Thefundershadnoroleinstudydesign,datacollectionandanalysis,decisiontopublish,orpreparationofthemanuscriptNowritingassistancewasutilisedintheproductionofthismanuscript.Bibliography1. JarboeJ,GuptaA,SaifW.Therapeutichumanmonoclonalantibodies
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