Neoplasia2020/21lecture4

Dr Heyam AwadMD,FRCPath

ILOS

• Tounderstandtheconceptofthehallmarksofcancerandthattheyarephenotypicchangesneededinallcancercells.• Tolistthetumorenablers• Tounderstandtheroleofhostcellsincarcinogenesis• Indepthunderstandingofthefirsthallmark:selfsufficiencyingrowthsignals.

Intro

• Canceroccursduetosuccessfulbreachofanticancermechanisms,i:eofregulatorymechanismsinourcellsandtissuesthatpreventuncontrolledproliferation.• Eachcancercellneedstoacquirecertainphenotypiccharacteristicstobecomemalignant.• Ofcoursethesephenotypiccharacteristicsreflectgeneticmutations(oralterations)• Thephenotypiccharacteristicsneededforcancercellstosurvivearecalled:thehallmarksofcancer.

HallmarksofCancer

• Hallmarksofcancer:changesincellphysiologyresultingfromgeneticchangesthatresultincancer.

HALLMARKSOFCANCER:theoriginalscientificwork:• DouglasHanahan andRobertA.Weinbergpublishedin2000areviewarticleabouttheexpandingknowledgeoftumorigenesis.TheysuggestedthatALLtumorcellsneedtoacquireSIXtraitsinorderforthecellstobetransformedtomalignantcells.• Thesesixtraitswerecalled:hallmarksofcancer• Theoriginalworkwasrevisitedandanotherarticlewaspublishedin2013..Seenext

• The2013articledescribedthe6characteristics(hallmarks)andemphasizedtheimportanceofgenomicinstabilityandtumorpromotinginflammationastwoenablersofmalignancy(theyenableandhelpthedevelopmentofthe6hallmarks.• Alsotwonew,emerginghallmarkswerediscussed:evasionoftheimmunesystemandreprogrammingofmetabolism• Moreover,thestromal,hostderivedcellsarethoughttoplayamajorroleincarcinogenesisastheyprovideasuitablemicroenvironment ofthetumortoevolve.

ThehallmarksofcancerYoucanreadthisarticlefrom:

https://doi.org/10.1016/j.cell.2011.02.013

Iencourageyoutoatleasttakealook.Thisisafantasticpaper.

Someofyourcolleagues(schoolofmedicine,UniversityofJordan)havetranslatedthepapertoArabic:

https://drive.google.com/file/d/1kFQruzNpAtUw_DCDPpWEzi9BQ3px1iOK/view?usp=drivesdk

Leading Edge

Review

Hallmarks of Cancer: The Next GenerationDouglas Hanahan1,2,* and Robert A. Weinberg3,*1The Swiss Institute for Experimental Cancer Research (ISREC), School of Life Sciences, EPFL, Lausanne CH-1015, Switzerland2The Department of Biochemistry & Biophysics, UCSF, San Francisco, CA 94158, USA3Whitehead Institute for Biomedical Research, Ludwig/MIT Center for Molecular Oncology, and MIT Department of Biology, Cambridge,MA 02142, USA*Correspondence: dh@epfl.ch (D.H.), weinberg@wi.mit.edu (R.A.W.)DOI 10.1016/j.cell.2011.02.013

The hallmarks of cancer comprise six biological capabilities acquired during themultistep develop-ment of human tumors. The hallmarks constitute an organizing principle for rationalizing thecomplexities of neoplastic disease. They include sustaining proliferative signaling, evading growthsuppressors, resisting cell death, enabling replicative immortality, inducing angiogenesis, and acti-vating invasion andmetastasis. Underlying these hallmarks are genome instability, which generatesthe genetic diversity that expedites their acquisition, and inflammation, which fosters multiple hall-mark functions. Conceptual progress in the last decade has added two emerging hallmarks ofpotential generality to this list—reprogramming of energy metabolism and evading immunedestruction. In addition to cancer cells, tumors exhibit another dimension of complexity: theycontain a repertoire of recruited, ostensibly normal cells that contribute to the acquisition of hall-mark traits by creating the ‘‘tumor microenvironment.’’ Recognition of the widespread applicabilityof these concepts will increasingly affect the development of new means to treat human cancer.

INTRODUCTION

We have proposed that six hallmarks of cancer together consti-tute an organizing principle that provides a logical framework forunderstanding the remarkable diversity of neoplastic diseases(Hanahan and Weinberg, 2000). Implicit in our discussion wasthe notion that as normal cells evolve progressively toa neoplastic state, they acquire a succession of these hallmarkcapabilities, and that the multistep process of human tumorpathogenesis could be rationalized by the need of incipientcancer cells to acquire the traits that enable them to becometumorigenic and ultimately malignant.

We noted as an ancillary proposition that tumors aremore thaninsular masses of proliferating cancer cells. Instead, they arecomplex tissues composed of multiple distinct cell types thatparticipate in heterotypic interactions with one another. We de-picted the recruited normal cells, which form tumor-associatedstroma, as active participants in tumorigenesis rather thanpassive bystanders; as such, these stromal cells contribute tothe development and expression of certain hallmark capabilities.During the ensuing decade this notion has been solidified andextended, revealing that the biology of tumors can no longerbe understood simply by enumerating the traits of the cancercells but instead must encompass the contributions of the‘‘tumor microenvironment’’ to tumorigenesis.

In the course of remarkable progress in cancer researchsubsequent to this publication, new observations have servedboth to clarify and to modify the original formulation of the hall-mark capabilities. In addition, yet other observations have raisedquestions and highlighted mechanistic concepts that were notintegral to our original elaboration of the hallmark traits. Moti-

vated by these developments, we now revisit the original hall-marks, consider new ones that might be included in this roster,and expand upon the functional roles and contributions madeby recruited stromal cells to tumor biology.

HALLMARK CAPABILITIES—CONCEPTUAL PROGRESS

The six hallmarks of cancer—distinctive and complementarycapabilities that enable tumor growth and metastatic dissemina-tion—continue to provide a solid foundation for understandingthe biology of cancer (Figure 1; see the Supplemental Informa-tion for downloadable versions of the figures for presentations).In the first section of this Review, we summarize the essenceof each hallmark as described in the original presentation in2000, followed by selected illustrations (demarcated by sub-headings in italics) of the conceptual progress made over thepast decade in understanding their mechanistic underpinnings.In subsequent sections we address new developments thatbroaden the scope of the conceptualization, describing in turntwo enabling characteristics crucial to the acquisition of the sixhallmark capabilities, two new emerging hallmark capabilities,the constitution and signaling interactions of the tumor microen-vironment crucial to cancer phenotypes, and we finally discussthe new frontier of therapeutic application of these concepts.

Sustaining Proliferative SignalingArguably the most fundamental trait of cancer cells involves theirability to sustain chronic proliferation. Normal tissues carefullycontrol the production and release of growth-promoting signalsthat instruct entry into and progression through the cell growth-and-division cycle, thereby ensuring a homeostasis of cell

646 Cell 144, March 4, 2011 ª2011 Elsevier Inc.

SO:

• FORCANCERTODEVELOP:weneedthe8characteristics(hallmarks)plusthe2enablingcharacteristicsandthecorrectmicroenvironment.

• Alltheaboveareacquiredasmutationsorepigeneticalterationsaccumulatingovertime• So:no singlemutationisenoughtocausecancer.

• Carcinogenesisisamultistep process.• Eachstepresultsfromageneticchange• Eachgeneticchangeresultsinacquiringacertainphenotype(oneofthehallmarks)• Thereisnospecificsequenceofacquiringthesephenotypes…themainissueistoacquirethemall

Hallmarksofcancer

• Eighthallmarks• 1.selfsufficiencyingrowthsignals• 2.insensitivitytogrowthinhibitorysignals• 3.evasionofcelldeath• 4.limitlessreplicativepotential• 5.sustainedangiogenesis• 6.abilitytoinvadeandmetastasize• 7.reprogrammingofmetabolism• 8.evasionoftheimmunesystem

Twoenablers

• Enablers (helpinthetumorigenesisprocessandsettherightconditionsforgeneticchangestodevelop)• 1.genomicinstability• 2.inflammation

• Tumormicroenvironment• Tumorsneedasuitableenvironmenttoflourish.• Thisisobtainedfromthehostcells:Stromalcells,mainlyfibroblastsandpericytes

1.Selfsufficiencyingrowthsignals

• Tumorsproliferateregardlessofthenormalregulatorymechanisms.• Thisisachievedbymutationsinproto-oncogenes• Oncogenescauseincreasedgrowthbyincreasingoncoproteins whichactasgrowthfactors,growthfactorreceptors,transcriptionfactors..etc

Physiologiccellproliferation

• 1.growthfactorbindstoitsreceptor• 2.thiscausestransientlimitedactivationofthereceptor• 3.theactivatedreceptoractivatessignaltransducingproteinsontheinnerleafletoftheplasmamembraneandthetransducedsignalistransmittedthroughthecytosoltothenucleus• 4.activationofnuclearregulatoryfactorsthatregulateDNAtranscription• 5.entryintocellcycle

Increasedgrowthcanhappenifthereisinterferencewithanyofthe5stepsinthepreviousslide.

1.Increasedgrowthfactors

2.Increasedreceptorsorincreaseintheirfunction

3.Increasedsignaltransduction

4.Increasedtranscriptionfactors5.Cellcycleentry

Growthfactors

• Sometumorsproducetheirowngrowthfactors,ie factorstheyrespondto.• Thiscreatesanautocrine loop• Eg glioblastomasproducePDGFandexpressitsreceptor

Growthfactors

• Tumorcellscaninteractwiththeirstromatoproducegrowthfactors.• Thisisanexampleofhowmicroenvironment(interactionwithhostcellsincludingstromalcells)canhelptumorstogrow.

GFreceptors

• GFreceptorscanincreasecellproliferationintwosituations:• 1.Overexpression ofthereceptor..So,ReceptorishyperresponsivetoGFeveninlevelsthatdon’tnormallytriggerproliferation.• 2.Mutantreceptorproteins.Sothereceptoritselfismutatedandacquiresaconfigurationthatisalwaysstimulated.ThiscausescontinuousmitogenicsignalevenintheabsenceofGF.

RememberthatwhenaGF(orindeedanyligand)bindstoitsreceptor,thiscausesconformationalchangesinthereceptorproteins,soitsshapechangesandthischangeiswhatactivatesthereceptor.Ifthereisamutationthatcausesconformationalchangeresultinginthereceptor’sshapebeingintheactivestate,thenwedon’tneedanystimulatorforthereceptortofunction!It’sinthealert,functioningstatecontinuously.

Noticetheshapechangeanddimerizationofthisreceptorwhenattachedtoitsligand.

AmplificationofGFreceptor/example

• Geneencodingher2/neu isamplifiedin25-30%ofbreastcancersandadenocarcinomaoflung,ovaryandsalivarygland• HighlevelofHER2/Neu proteininbreastcancerworsensprognosis(resultsinmoreproliferation)• Ifher2receptorblocked..Byantiher2antibodies..Treatmentused(refertodetailsinpreviouslecture:thecasestudy)

Signaltransducingproteins

• Mainsignaltransducingproteinsinvolved:RASandABL

RAS

• RAS isthemostcommonlymutatedoncogeneinhumans• 30%ofhumancancerscontainRASmutation• Evenhigherincidenceincolonandpancreaticcarcinomas

HowdoesRASfunctionnormally?

• RASisasmallproteinthatbindsGDPandGTP(it’saGprotein)• ItisinactivewithGDPbinding.• Ifagrowthfactorbindstoareceptor..GDPisphosphorylatedtoGTP..• ThisActivesRAS• ActivationisshortlivedasGTPase hydrolysesGTPtoGDP• GTPaseactivatingproteins(GAPs)preventoveractivationofRAS,sotheyactasbrakesofRASactivation.

Gcoupledproteins,whichRASisoneofthem.• Seventransmembranealphahelices,coupledwithGprotein(GTPbindingprotein).• Ligandbinding:GDPintheGproteinchangestoGTP…receptoractivated.

• ActivatedRAS..Sendsmessengerstothenucleusbytwopathways(seenextslide)• 1.RAF..MAPK• 2.PI3K..AKT..mTOR

• If1or2mutated..CanmimicRASeffect• Eg RAFmutatedin60%ofmelanomas

RASRASisstimulatedwhenagrowthfactorbindstoitsreceptor.

ThisbindingcausesphosphorylationofGDPtoGTP.

GTPactivatesRAS

ActivatedRASactsviatwopathwaystosendsignalstothenucleus

1. mTORpathway.

2. RAF- MAPKpathway

Thesetwopathwayssendmessagestothenucleustoexpresstranscriptionfactorsthatallowcellstoentercellcycleandproliferate.

HowRASisactivated(mutated)

• PointmutationinanaminoacidresidueswithintheGTPbindingpocketor intheenzymaticregionofGTPhydrolysis• Bothresultindefectivehydrolysis..LeadingtotrappedRASinactivephosphorylatedGTPboundRAS

ABL

• Proto-oncogenewithtyrosinekinaseactivityregulatedbyinternalnegativeregulatorydomain• ABL-BCRtranslocation..Resultsinahybridgene• Thehybridgeneproducesnoveltyrosinekinasewhichisalwaysactiveandstimulatingproliferation…ThisisthePhiladelphiachromosomewetalkedaboutpreviously.• ABL-BCRpathwayactivatesallthedownstreamsignalsofras

Philadelphiachromosome

• CML=chronicmyelogenousleukemia:t(9;22)=Philadelphiachromosome.ABL-BCRfusiongene….tyrosinekinasewhichisalwaysturnedon.• KinaseinhibitorscanbeusedtotreatCML• Imatinab/gleevec ..Inhibitsthekinaseandtreatstheleukemia• =targetedtherapy

Nucleartranscriptionfactors

• Mostcommonlyinvolvedinhumancancers:MYC

myc

• Canactivateorrepressexpressionofothergenes• Activatescyclins andcyclin dependentkinasesCDK(socellsentercellcycleanddivide)• Myc alsoinhibitscyclindependentkinaseinhibitors(CDKI)

• Thesecyclins,CDK,CDKIplayaroleinentrytocellcycle,whichwewilldiscussindetailinthislecture.

Entryintocellcycle

• Allthestimulimentionedtillnowaimforquiescentcellstoenterthecellcycle.• Eachphaseinthecellcycledependsonsuccessfulcompletionofthepreviousone• Cyclestopswhenessentialgenefunctionislost

cyclins

• Progressionthroughcellcycle,especiallyG1-Sisregulatedbyproteinscalledcyclins• Cyclins activatekinasesCDK(cyclin dependentkinases)• Cyclin andCDKformcomplexesthatphosphorylatetargetproteinsthatdrivethecellthroughthecellcycle.• Cyclins:D,E,A,B(theyappearinthissequence)

TheDEABsequenceofcyclins !

• SO:cyclin/CDKcomplexescauseproliferation.• Thesesareinhibitedbycyclin dependentkinaseinhibitors(CDKI)• CDKIimportantforenforcingthecheckpointsanddelayingcellcycle.

• Mutationscausingincreasedcyclins orCDKcauseselfsufficiencyingrowthsignals.• MutationsinhibitingCDKIwillcauseincreasedgrowth.

• Examples:cyclin Disactivatedinseveraltumorsmainlylymphomas.

Summary1/3

• Transformedcellsacquireseveralmutationsresultinginseveralphenotypes.• Thereare8phenotypesneededineachtransformedcellandthesearecalledthehallmarksofcancer.• Allthesehallmarksmustbegainedthroughmutations,butthereisnospecificsequenceforacquiringthem.• Wedon’tneed8mutationsforthe8phenotypesasonemutationmightcausemorethanphenotype.• these8are:selfsufficiencyofgrowthfactors,insensitivitytogrowthinhibitorysignals,evasionofcelldeathandtheimmunesystem,changesinmetabolism,immortality,sustainedangiogenesisandabilitytoinvadeandmetastasize.• Genomicinstabilityandinflammationactasenablersofmalignancy;theysettheproperenvironmentformutationstooccur.• Tumorsalsoneedapropermicroenvironmentprovidedbyhoststromalcells.

Summary2/3

• Selfsufficiencyingrowthsignalscanoccurthroughincreasedgrowthfactors,growthfactorreceptors,signaltransductionproteins,transcriptionfactorsorcellcyclestimulators.

• GFcanbesynthesizedbytumorcellsorhoststromalcells.• GFreceptorscanbeactivatedviaoverexpressionorchangesintheirarchitecturethatmakesthemactiveevenwithoutbindingtoGF.

• Signaltransductioncanincreaseviaincreaseinanyproteininvolvedinsecondmessengersthatconveygrowthsignalstothenucleus.TheseincludeRASandABLplustheirdownstreamproteinpathways(BRAF- MAPkinaseandMTORpathways)

• RASisthemostcommonlymutatedoncogeneinhumans.• RASisaGproteinthatisstimulateduponphosphorylationofGDPtoGTP.• PointmutationsthatcauseentrapmentofRASintheactivatedstatecausecelltransformation.

Summary3/3

• ABLisaprotooncogenethatcanbestimulatedviatranslocation(9;22)withformationofABL-BCRfusiongenethatencodesakinasewhichisactiveandcausescellproliferation.Thiscausesaleukemiathatcanbetreatedbyblockingthekinase.• Cellcycleisstimulatedbycyclins/Cyclindependentkinase(CDK)complexes.IncreaseinCyclinsorCDKcantransformcells.• Cyclin/CDKcomplexesarenormallyregulatedbyCDKinhibitors..Adecreaseintheinhibitorscanalsotransformcells.

Testyourself

• Thequestionstofollowarefromwebpath website,it’saverygoodsourceofpathologyMCQquestions:https://library.med.utah.edu/WebPath/EXAM/EXAMIDX.html

- Myaimhereistoshowyouhowlong,seeminglydifficultquestionsareactuallyveryeasy.Justknowyourconceptsandpracticehowtodealwiththesequestions:let’stry..

• A73-year-oldmanhasanepisodeofhematemesis.UpperGIendoscopyrevealsanirregular4cmgastricantral ulceration.Biopsiesareperformedandmicroscopicallyrevealadenocarcinoma.MolecularanalysisshowsDNAhypermethylation oftheCDKN2complex.Throughwhichofthefollowingmechanismshasthisabnormalgeneexpressionmostlikelyoccurred?• AAmplification• BEpigeneticalteration• CGrowthfactorbinding• DPointmutation• EReducedmiRNA expression

answer• A73-year-oldmanhasanepisodeofhematemesis.UpperGIendoscopyrevealsanirregular4cmgastricantral ulceration.Biopsiesareperformedandmicroscopicallyrevealadenocarcinoma.MolecularanalysisshowsDNAhypermethylationoftheCDKN2complex(thisistheonlyrelevantpieceofinformation.Ifyouknowthathypermethylationisanepigeneticchangethenyousolvedthewholeissue..Throughwhichofthefollowingmechanismshasthisabnormalgeneexpressionmostlikelyoccurred?• AAmplification• BEpigeneticalteration• CGrowthfactorbinding• DPointmutation• EReducedmiRNA expression

• Columnarepithelialcellsfromthecolonicmucosaarestudiedtoidentifyabnormalitiesincellsignalingpathways.AbnormalepithelialcellsfromcolonicadenocarcinomaareshowntohaveamutationthatblockshydrolysisofGTP-boundactiveRAS.NormalcolumnarcellshaveactiveRASproteinthatundergoeshydrolysistotheinactiveGDP-boundform.Whichofthefollowingsignalingpathwaysismostlikelyabnormallystimulatedinthecarcinomacells?• AADP• BBCR-ABL• CCyclicAMP• DCyclin D1• EMAPkinase• Fp53

• Columnarepithelialcellsfromthecolonicmucosaarestudiedtoidentifyabnormalitiesincellsignalingpathways.AbnormalepithelialcellsfromcolonicadenocarcinomaareshowntohaveamutationthatblockshydrolysisofGTP-boundactiveRAS.NormalcolumnarcellshaveactiveRASproteinthatundergoeshydrolysistotheinactiveGDP-boundform.Whichofthefollowingsignalingpathways ismostlikelyabnormallystimulatedinthecarcinomacells?• AADP• BBCR-ABL• CCyclicAMP• DCyclin D1• EMAPkinase.Allwhatyouneedtoknowhereisthesignalingpathwaysofras:RAF- MAPKandMTOR• Fp53

• A50-year-oldwomannotesalumpinherbreast.Herphysician'sassistantpalpatesa2cmfirmmassinherleftbreast.Afineneedleaspirationbiopsyisperformed,andonmicroscopicexaminationacarcinomaispresent.MolecularanalysisshowsHER2positivitybutestrogenreceptornegativityinthesemalignantcells.Throughwhichofthefollowingmechanismshasthisabnormalexpressionmostlikelyoccurred?• AAmplification• BEpigeneticalteration• CGrowthfactorbinding• DPointmutation• EReducedmiRNA expression

• A50-year-oldwomannotesalumpinherbreast.Herphysician'sassistantpalpatesa2cmfirmmassinherleftbreast.Afineneedleaspirationbiopsyisperformed,andonmicroscopicexaminationacarcinomaispresent.MolecularanalysisshowsHER2positivitybutestrogenreceptornegativityinthesemalignantcells.Throughwhichofthefollowingmechanismshasthisabnormalexpressionmostlikelyoccurred?• AAmplification ..Alltheinformationisirrelevant..TheyarejustaskingabouthowHER2isactivated,andyoushouldknowbynowit’sthroughamplification.HER2=EGFR• BEpigeneticalteration• CGrowthfactorbinding• DPointmutation• EReducedmiRNA expression

• A66-year-oldmanhasnoteddarkerurineforthepast2weeks.Aurinealysis showshematuria.Cystoscopyisperformedandthereisa3cmmassinthedomeofthebladder.Biopsiesofthemassaretakenandonmicroscopicexaminationshowaurothelial carcinoma.CellsofthisneoplasmdemonstrateasinglemutationcausingcellularinabilitytohydrolyzeGTP,thusresultingincellulartransformation.Whichofthefollowingoncogenesismostlikelyimplicatedinthiscase?• AABL• BERBB2• CSIS• DRAS• EN-MYC

• A66-year-oldmanhasnoteddarkerurineforthepast2weeks.Aurinealysisshowshematuria.Cystoscopyisperformedandthereisa3cmmassinthedomeofthebladder.Biopsiesofthemassaretakenandonmicroscopicexaminationshowaurothelial carcinoma.CellsofthisneoplasmdemonstrateasinglemutationcausingcellularinabilitytohydrolyzeGTP,thusresultingincellulartransformation.Whichofthefollowingoncogenesismostlikelyimplicatedinthiscase?• AABL• BERBB2• CSIS• DRAS….YouonlyneedtoknowthatRASisaGproteinanditsfunctionisrelatedtoGDP/GTP.Youshouldbeabletoanswerthequestionevenifyouhavenoideawhattherestofthechoicesare.• EN-MYC

• A52-year-oldmanhashadincreasingfatigueforthepast6months.Onphysicalexaminationhehasapalpablespleentip.LaboratorystudiesshowaWBCcountof189,000/microliter.Theperipheralbloodsmearshowsmanymatureandimmaturemyeloidcellspresent.Cytogeneticanalysisofcellsobtainedviabonemarrowaspirationrevealsat(9:22)translocation.Thistranslocationleadstoformationofahybridgenethatgreatlyincreasestyrosinekinaseactivity.Whichofthefollowinggenesismostlikelytranslocated tocausethesefindings?• Ap53• BRB• CABL• DNF-1• ERAS

• A52-year-oldmanhashadincreasingfatigueforthepast6months.Onphysicalexaminationhehasapalpablespleentip.LaboratorystudiesshowaWBCcountof189,000/microliter.Theperipheralbloodsmearshowsmanymatureandimmaturemyeloidcellspresent.Cytogeneticanalysisofcellsobtainedviabonemarrowaspirationrevealsat(9:22)translocation.Thistranslocationleadstoformationofahybridgenethatgreatlyincreasestyrosinekinaseactivity.Whichofthefollowinggenesismostlikelytranslocated tocausethesefindings?• Ap53• BRB• CABL..Thisisveryeasy,youonlyneedtopracticetopicktherelevantinformationinthesequestions• DNF-1• ERAS