Clinical Proteomics Technologies for Cancer

What is Cancer Proteomics?

Theterm"proteome"referstotheentirecomplementofproteins,includingthemodificationsmadeto
aparticularsetofproteins,producedbyanorganismoracellularsystem.Thiswillvarywithtimeand
distinctrequirements,suchasstresses,thatacellororganismundergoes.Theterm"proteomics"is
alargescalecomprehensivestudyofaspecificproteome,includinginformationonprotein
abundances,theirvariationsandmodifications,alongwiththeirinteractingpartnersandnetworks,in
ordertounderstandcellularprocesses.Clinical proteomicsisasubdisciplineofproteomicsthat
involvestheapplicationofproteomictechnologiesonclinicalspecimenssuchasblood.Cancer,in
particular,isamodeldiseaseforapplyingsuchtechnologiestoidentifyuniquebiosignaturesand
biomarkersresponsibleforthediagnosis,prognosisandtherapeuticpredictionofsuchdisease.
Biomarkersarebiologicalmoleculesfoundinblood,otherbodyfluids,ortissuesthatareasignofanormalorabnormalprocess,
orofaconditionordisease.Theymayalsobeusedtoseehowwellthebodyrespondstoatreatmentforadiseaseorcondition.

Why Proteomics?
Thegreatestpromiseforthedetectionandtreatmentofcancerliesinthedeepunderstandingofmolecularbasisfordisease
initiation,progressionandefficacioustreatmentbasedonthediscoveryofuniquebiomarkers.Althoughprogressincancer
genomicshasbeenrapidduringthepastfewyears,itonlyprovidesuswithaglimpseofwhatmayoccurasdictatedbythe
geneticcode.Inreality,westillneedtomeasurewhatishappeninginapatientinrealtime,whichmeansfindingtelltaleproteins
thatprovideinsightintothebiologicalprocessesofcancerdevelopment.Thisisbecausegenesareonlythe"recipes"ofthecell,
whiletheproteinsencodedbythegenesareultimatelythefunctionalplayersthatdrivebothnormalanddiseasephysiology.

Theaccessibilityofcancerrelatedproteinsintissuesandbodilyfluidshastriggeredextensiveproteinfocusedresearchforthe
huntofbiomarkers.Proteomicshastheabilitytointerrogateavarietyofbiospecimensfortheirproteincontentsandaccurately
measuretheconcentrationsoftheseproteins.Thiscanprovidescientistsandclinicianswithapowerfultooltounderstandthe
differentprocessesinvolvedincancerdevelopmentandprogressioninhopetoidentifybiomarkersspecificforthesecellular
processesalongwiththoseindicatingefficacioustherapeuticintervention.

What are the unique challenges for proteomics compared to genomics?

What are the unique challenges for proteomics compared to genomics?

Thebiggestconceptualchallengeinherentinproteomicsliesintheproteome'sincreaseddegreeofcomplexitycomparedtothe
genome.Forexample:

One gene can encode more than one protein(evenupto1,000).Thehumangenomecontainsabout

21,000proteinencodinggenes,butthetotalnumberofproteinsinhumancellsisestimatedtobebetween250,000
toonemillion.
Proteins are dynamic.Proteinsarecontinuallyundergoingchanges,e.g.,bindingtothecellmembrane,
partneringwithotherproteinstoformcomplexes,orundergoingsynthesisanddegradation.Thegenome,onthe
otherhand,isrelativelystatic.
Proteins are co and posttranslationally modified.Asaresult,thetypesofproteinsmeasuredcanvary
considerablyfromonepersontoanotherunderdifferentenvironmentalconditions,orevenwithinthesameperson
atdifferentagesorstatesofhealth.Additionally,certainmodificationscanregulatethedynamicsofproteins.
Proteins exist in a wide range of concentrations in the body.Forexample,theconcentrationofthe
proteinalbumininbloodismorethanabilliontimesgreaterthanthatofinterleukin6,makingitextremelydifficultto
detectthelowabundanceproteinsinacomplexbiologicalmatrixsuchasblood.Scientistsbelievethatthemost
importantproteinsforcancermaybethosefoundinthelowestconcentrations.

Proteomics as the Bridge between Genomics and Biology

Inordertobetterunderstandcancerbiologyandtoacceleratethedevelopmentofbothcancerdiagnosticsandtherapeutics,
biologicalinsightfromgenomicanalysisisbeingintegratedwiththeanalysisofproteincontentintumorcells.Byunderstanding
theproteincomponentsresultingfromgeneticaberrationsincancer,scientistscanbegintopiecetogetherwhatchangesare
occurringinacancerproteome.Basedupontheprogressmadeinthereproducibilityandtransferabilityofproteomicworkflows
andmethodologyintheClinicalProteomicTechnologiesforCancerinitiativelaunchedin2006,ClinicalProteomicTumor
AnalysisConsortium(CPTAC),istolinkcancergenometoproteomebysystematicallyanalyzingtheproteincontentoftumorsof
whichthereiscomprehensivegenomiccharacterizationfrominitiativessuchasTheCancerGenomeAtlas.Thisintegrative
approachwillproduceadeeperunderstandingofcancerbiology,withhighqualitydatasets,reagents,andanalyticallyvalidated
quantitativeassaystobemadepubliclyavailable.

What Tools are Utilized in Proteomics?

Describedbelowaresomeofthemaintechnologiesbeingusedtoadvanceourunderstandingofproteinbiochemistry.

Mass Spectrometry
Massspectrometry(MS)isanevolvingtechnologythatallowsscientiststodetectandquantifyproteinsina
complexbiologicalmatrix.Suchmethodsareveryprecise,distinguishingproteinsthatdifferincomposition
byasinglehydrogenatom,thesmallestatom.Despiteitspotential,MStechnologiesarenotyetcapableof
separatingthecomplexproteinmixturesfromunprocessedhumanbiospecimens.Additionaltechnologies
suchasorganelleorproteinfractionationoraffinitycapturehavebeendevelopedtoreducethecomplexity
ofproteinsinbiospecimensbyenrichingforasubsetofproteinsofinterest,inadditiontoimprovingthe
sensitivityofinstrumentationfordetectionandquantificationofproteins.

Protein Microarrays
Proteinmicroarraysarepowerfultoolsforcapturingandmeasuringproteinsfrombiospecimeninahigh
throughputfashion.Aproteinmicroarraytypicallyconsistsofasmallpieceofglassorplasticcoatedwith
thousandsof"capturereagents"(moleculesthatcan"grab"specificproteins).Thistechnologyallows
scientiststoisolateandstudymanypotentialbiomarkerproteins.Proteinmicroarrayscanbeminiaturized
tocontaintensofthousandsofcapturefeaturesarrangedinagrid,eachspecificforagivenprotein,
therefore,theyareconsideredamultiplexeddeviceforexample,theycantestformultiplebiomarkers
simultaneously,whichisessentialforclinicaluse.

Nanotechnologies

Nanotechnologies
Nanotechnologyisthecreationofmanufacturingdevicesandcomponentsthatrangefrom1to100
nanometers.Ananometerisonebillionthofameter,or1/80,000thewidthofahumanhair.
Nanotechnologydeviceshavethepotentialtogreatlyexpandthecapabilitiesofproteomics,addressing
currentlimitationsinselectivelyreachingatargetproteininvivothroughphysicalandbiologicalbarriers,
detectinglowabundancetargets,andprovidinga"toolbox"totranslatethediscoveryofproteinbiomarkers
tonoveltherapeuticanddiagnostictests.Typicalnanodevicesincludenanoparticlesusedforthetargeted
deliveryofanticancerdrugs,energybasedtherapeutics(includingheatandradiation)andimaging
contrastreagents.Nanowiresandnanocantileverarrayscanbeusedinbiosensorsthatmeasureminute
quantitiesofbiomarkersinbiologicalfluids.

Formoreinformation,seetheNCIAllianceforNanotechnologyinCancer

Bioinformatics
Majorareasoffocusinbioinformaticsresearchincludedatamodelinganddatabasedesign,datainteroperabilityand
comparison,geneandproteinexpressionanalysis,structuralpredictions,vocabulariesandontologies,aswellasmodelingfor
systemsbiology.InthesecondphaseoftheCPTCprogram,thedevelopmentofnewbioinformatictoolsforintegrativeanalysisof
genomicandproteomicdataisnecessarytodrivethecollaborative,multidisciplinaryeffortrequiredtodrivediscoveryfromthe
laboratorytoclinicalpractice.

Biospecimens
Cancerresearchhascometorelyheavilyonthequalityofbiospecimensforthemeasurementofgeneticandproteinexpression,
andthelinkageofthatinformationwithclinicalstatusanddiseasepathwayssuchastumorgrowth,migration,metastasis,
angiogenesis,andapoptosis(celldeath).Sincecancerdiagnosisandtreatmentoftenbeginwithdiagnosticbiopsiesfollowedby
surgicalresectionofthetumor,therearemanyopportunitiestocollectvaluablebiospecimensforresearch.TheNCIhas
recognizedthecriticalneedforresearchaccesstolargenumbersofhighqualitybiospecimensannotatedwithclinicaldata.NCI
isaddressingthiscriticalneedthroughitsOfficeofBiorepositoriesandBiospecimenResearch.

Reagents
Thereisagrowingneedinthefieldofproteomicsforhighquality,wellcharacterizedstandardreagentsthatcanimprovethe
specificityandreproducibilityofproteomictechnologies.Onewidelyusedreagentinproteomicresearchisanantibody,a
naturallyoccurringserumproteinwhosebiologicalrolerequireshighantigenspecificity.Theyhavelongbeenusefulasthe
captureanddetectionreagentsinproteomics.

Alternativeaffinityreagentssuchasaptamershaverecentlyshowngreatpromiseasanadjuncttoantibodies.Thesenucleicacid
basedmoleculespossessproteinbindingspecificity,similartoantibodiesthatmakethemusefulasproteincaptureanddetection
reagents.

Standardproteomicreagentswillbeusefulformanyapplicationsincancerresearch,including:

Reportermoleculesthatdetectthepresenceofatarget(ormodificationstoit)inaparticularbiologicalsample
Capturemoleculesforpurifyingthetargetfromacomplexbiologicalsamplepriortoidentificationandquantification
using,forexample,massspectrometry
Functionalstudiestovalidatetheroleofapotentialtherapeutictargetpriortolaunchingdrugdiscoveryor
developmentefforts
Referencematerialsforcalibratinginstrumentsorcomparingdifferentproteomicplatformtechnologies.