Cancer stem cells modulate patterns and processes of...

Accepted for publication in Philosophy and Biology 25.04.2018

1

Cancerstemcellsmodulatepatternsandprocessesofevolutionincancers

LucieLaplane1,2

1.UMR8590Institutd’HistoireetPhilosophiedesSciencesetdesTechniques(IHPST),CNRS,UniversitéParisIPanthéon-Sorbonne,ENS,13rueduFour,75006Paris,France;2.UMR1170,Inserm,GustaveRoussy,114rueEdouardVaillant,Villejuif,France

Correspondenceto

LucieLaplaneIHPST,UniversitéParisIPanthéon-Sorbonne.13rueduFour75006Paris,Francee-mail:[email protected]:+33(0)143546036fax:+33(0)143252948

Abstract

Theclonalevolution(CE)modelandthecancerstemcell(CSC)modelaretwoindependentmodelsofcancers,

yetrecentdatashowsintersectionsbetweenthetwomodels.ThisarticleexplorestheimpactsoftheCSCmodel

ontheCEmodel.IshowthatCSCrestriction,whichdependsonCSCfrequencyincancercellpopulationsandon

theprobabilityofdedifferentiationof cancernon-stemcells (non-CSC) intoCSCs, can favoror impede some

patternsofevolution(linearorbranchedevolution)andsomeprocessesofevolution(drift,evolutionbynatural

selection,complexadaptations).TakingCSCrestrictionintoaccountfortheCEmodelthushasimplicationsfor

thewayinwhichweunderstandthepatternsandprocessesofevolution,andcanalsoprovidenewleadsfor

therapeuticinterventions.

Keywords

Cancerstemcell;Clonalevolution;Cancerprogression;Drift;Evolutionbynaturalselection

Acknowledgments

Thisworkwasfinancedbytheprogram“StratifiedOncologyCellDNARepairAndTumorImmuneElimination”

(SOCRATE)oftheSiteofIntegratedResearchInCancerology(SIRIC)inGustaveRoussy(grantINCa-DGOS-INSERM

6043)andtheCancéropôleÎle-de-France(grant2016-1-EMERG-50-CNRSDR1-1).

Iamdeeplygratefultothereviewers. Ihavebenefitedfromthebestreviewspossibleandtheirremarksand

critiqueswereparticularlyhelpfultoreviseandstrengthenmyargument.KateMacCorddidanamazingjobnot

onlywitheditingtheEnglishbutalsowithhelpingmetoclarifymyarguments.Thispaperalsogreatlybenefited

frommanycomments fromphilosophers,biologists, andoncologists,on itsearlyand/or later versions. I am

particularlygrateful toMelindaFagan, JeanGayon,Pierre-LucGermain,PeterGodfrey-Smith,KateMacCord,

FrancescaMerlin,ThomasPradeu,MurielleGaudry,StéphaneGiraudier,Anne-MarieLyne,MrinalPatnaik,Leïla

Perié,andespeciallytoEricSolaryandallofmyteamatGustaveRoussy.


2

Theclonalevolution(CE)modelandthecancerstemcell(CSC)modelaretwomodelsofcancerthathavean

ambiguous theoretical relationship. The CEmodel describes cancer as an evolutionary process inwhich the

accumulationofgeneticandepigeneticalterationsresultsinthediversificationofcancercellsthroughspaceand

time(Figure1a).TheCSCmodelstatesthatcancersmimictissueorganizationandemergeanddeveloponlyfrom

arestrictedfractionofcancercellswithstem-likepropertiescalledCSCs(Figure1b).Historically,theCSCmodel

wasframedinoppositiontotheCEmodel(Reyaetal.2001;Wichaetal.2006;Shipitsinetal.2007;Pantic2011).

Someinvestigatorshavesincesuggestedthatthetwocompetingmodelsare“notnecessarilymutuallyexclusive”

(Polyak2007,3160;CampbellandPolyak2007;Lagasse2008), thatoneor theothercouldmoreadequately

explaindifferenttypesofcancer(AdamsandStrasser2008;Shackletonetal.2009),orthatthetwomodelsare

complementaryandshouldbecombinedbecause theyeachprovideexplanationsofdifferentaspectsof the

samecancer(Fabianetal.2009;Greaves2010;Langetal.2015).

Several linesofexperimentationhaveshownthatCEoccurs inCSCs(forthefirstevidenceseePiccirilloetal.

2009;Andersonetal.2011;Nottaetal.2011).TakingintoaccounttheimpactofCEonCSCsintheCSCmodel

appearstobeimportantforatleasttworeasons.First,forthetherapeuticstrategythatemergedfromtheCSC

model and that consists of specifically targeting the CSCs during cancer treatments, itmeans that CSCs can

represent a heterogeneous target. CSCs can differ both synchronically (at the diagnosis for example) and

diachronically (from diagnosis to relapse for example). This heterogeneity between CSCs can result in the

coexistenceofCSCswithvarioussensitivitiestotherapies(e.g.Meyeretal.2015).Second,theaccumulationof

new mutations in CSCs can modify their properties. For example, additional mutations could lead to the

acquisition of migratory ability, distinguishing “stationary CSCs” from “migrating CSCs” able to generate

metastases(Brabletzetal.2005;Bapat2007;Odouxetal.2008;TakebeandIvy2010).Bynottakingintoaccount

CE,theCSCmodelmissestheheterogeneityofthisCSCpopulation.TheincorporationofCEintotheCSCmodel

isnowfairlyaccepted(e.g.KresoandDick2014).

Incontrast,theCEmodelstilllargelyignorestheCSCmodel.Throughoutthispaper,IarguethatCSCsimpactCE,

particularly through a phenomenon that I call “CSC restriction” (referring to the fact that CSCs are only a

restrictedfractionofagivencancer),andthusthattheCEmodelneedstoincorporatesomefeaturesoftheCSC

modelinordertoaccountforalloftheconsequencesthatCSCrestrictionhasforCE.

Thisresearchtookrootfrompreviousphilosophicalworks.InhisbookonDarwinianpopulations,Godfrey-Smith

(2009) introduced a framework that allows one to distinguish different types of Darwinian populations, for

example paradigmatic Darwinian populations in which evolution by natural selection can produce complex

adaptations,i.e.noveltraits,versusmarginalDarwinianpopulationsinwhichevolutionbynaturalselectioncan

only produce simple adaptations, i.e. change in trait frequency. Distinguishing between types of Darwinian

populationsdependsonvariousparametersofevolutionthatcanberoughlymeasured(amongwhicharethe

classicalrequirementsofevolutionbynaturalselectionsuchasvariation,heritability,anddifferentialfitness,as

wellasadditionalonesthatcanalsomodifyhowDarwinianapopulationis).Applyingthisframeworktocancers,

Germain(2012)highlightedthatcancercellpopulationsaremuchlesslikeparadigmaticDarwinianpopulations

thanusuallyassumedandthisisinlargepartduetotheCSCmodel,whichappearsasamajorfactorthatlimits

theabilityofcancercellstoevolvebynaturalselection(seesection1.1).Germain’sconclusionwasthepointof


3

departureformywork.IfcancercellsaremuchlesslikeparadigmaticDarwinianpopulationsthanexpected,then

whatkindofpopulationsaretheyexactly?Inthispaper,IbuildaframeworksimilartoGodfrey-Smith’sinwhich

themeasureofsomefactorsassociatedwithCSCschangestheCEfeaturesofcancercellpopulations.Thetwo

factorsofinterestareCSCfrequency(howmanycancercellsareCSCs?)andnon-CSCdedifferentiation(whatis

theprobabilityofnon-CSCsbecomingCSCs?),whichcapturewhatIcall“CSCrestriction”.TheCEfeaturesunder

the influence of CSC restriction are processes of evolution (evolution by natural selection versus neutral

evolution,complexversussimpleadaptations)andpatternsofevolution(linearversusbranchedevolution).

ThefirstsectionshowsthatCSCrestrictionlimitsevolutionbynaturalselection(andmoregenerallyCE),prevents

complex adaptations, and favors drift and linear evolution. Scientists and philosophers have compared CSC

restriction to other phenomena well known in evolutionary biology such as reproductive specialization or

effectivepopulationsize,raisingthequestionofwhetherwecoulddescribetheimpactofCSCrestrictiononCE

throughclassicalparametersofevolution.Thesecondsectionshowsthatnoneoftheseparametersaccurately

accountforCSCrestriction,inparticularbecauseitreliesontwodifferentfactors(CSCfrequencyandprobability

ofdedifferentiation).Thelastsectionexploresthepotentialbiomedicalinterestsofinvestigatingtheimpactof

CSCrestrictiononpatternsofCE.

Figure1TheCEmodelandtheCSCmodel.TocomparetheCEmodelandtheCSCmodel,Iproposetwoclassicalrepresentationsofthemodels(aandb,toppanels)andtwomoreabstractedrepresentationsofthesemodels(a and b, bottom panels). The more abstracted representations are useful to represent graphically theconsequenceofCSCrestrictiononCEinthenextfigure


4

1.CSCrestrictionimpactsCE

BeforeexploringtheimpactofCSCsonCEitisnecessarytohaveaclearunderstandingofCSCs,andinparticular

ofwhatIcallCSCrestriction.TheCSCmodelpicturescancersasacaricatureofthenormaltissuesfromwhich

theyemerge(forarecentdiscussion,seeBatlleandClevers2017).Inmostnormaltissuesinhomeostasis,there

is a hierarchical organizationof cells inwhich a small fractionof stem cells sit at the apexof a hierarchyof

differentiatingcells.Whenstemcellsdivide,theycangiverisetonewstemcells,thusensuringthemaintenance

ofthestemcellpool,and/ortheycandifferentiateintomorematurecells(Fig1b).Incancers,thisorganization

isaltered(differentiationcanbeblocked,biased,oramplified),yetinmanycases,ahierarchyismaintainedand

onlyafractionofthecancercells(theCSCs)isabletodevelopandmaintainthetumor.1

TheexplanatoryvalueoftheCSCmodelreliesontwopremises:firstthatnotallcancercellsareCSCs,andsecond,

thatCSCscanbedistinguishedfromnon-CSCs(i.e.theobservedfunctionaldifferencebetweenCSCsandnon-

CSCsisnotstochastic).ThemassivesearchtoidentifyCSCsinallcancershasproducedconflictingdataforboth

premises.First,estimatedCSCfrequencycanvarybyordersofmagnitude,fromveryrareinsomecancers(e.g.

50in106inacutemyeloidleukemia;BonnetandDick1997)toverycommoninothers(e.g.1outof4cellsin

advancedmelanoma;Quintanaetal.2008).SomemutationscanresultinanincreaseinCSCfrequency,indicating

thatCSCfrequencycanalsochangeduringcancerprogression(Nottaetal.2011;Clappieretal.2011).Thus,the

CSCfrequencyvariesdependingoncancertypesandstages,anditcanbemeasured,atleastroughly.2Second,

in some cancers, non-CSCs can dedifferentiate into CSCs (e.g. Chaffer et al. 2013; Koren et al. 2015; Van

Keymeulen et al. 2015;Medema 2017). Dedifferentiation affects the quantification of the cancer cells that

contribute tocancermaintenanceandevolution through timebecausewhendedifferentiation ispossible, in

additiontotheactualCSCs,thenon-CSCsrepresentapoolofpotentialCSCs.Howmanyofthesenon-CSCswill

becomeCSCsisavariablethatcanbemeasuredastheprobabilityofdedifferentiation.Dedifferentiationofnon-

CSCsintoCSCsisaprocessspecifictosomecancers(mostlyobservedinepithelialcancersandneverobservedin

bloodcancerssofar)andtheprobabilityofdedifferentiationofanon-CSCintoaCSCcanvarybetweencancers

(e.g.betweenbreastcancersfromdifferentcelloforigin,Latiletal.2017).

Iusetheterm‘CSCrestriction’torefertothesetwodynamicaspectsofCSCs(CSCfrequency,e.g.theiractual

number,andnon-CSCdedifferentiation,e.g.theprobabilitythatnon-CSCsdedifferentiateintoCSCs).Thelevel

of CSC restriction thus varies in different cancer types and/or stages. CSC restriction is high when the CSC

frequencyandtheprobabilityofdedifferentiationarelowandvice-versa.Thissectiondiscussestheimpactof

thelevelofCSCrestrictiononCE,basedonphilosophical,modeling,andbiologicalliterature.

1.1.CSCrestrictionlimitsevolutionbynaturalselectionandpreventscomplexadaptations

1ThecelloforigincaneitherbeastemcelltransformedintoaCSCoranon-stemcellthatreacquiredstemnessattransformationandbecameaCSC.Foradetailedaccountofthehistory,structureandcontentoftheCSCmodel,seeLaplane(2014,2016,chap2-4).2CSCquantificationreliesonmanyexperimentalparametersthatcanallintroducebiases(reviewedinKresoandDick2014;NassarandBlanpain2016).


5

In theCEmodel, all the cancer cells can, inprinciple, evolve.Cancers arepopulationsof cells that asexually

reproducethroughcelldivision.Duringdivisiontheycanacquiremutationsthattheytransmittotheirdaughter

cells.Theimpactofmutationsoncellfitnessisvariablesothattwotypesofmutationsareusuallydistinguished

incancers:passengermutations,i.e.thosethatdonotchangethefitnessofthecells,anddrivermutations,i.e.

thosethatincreasetheirfitness(amutationcanbepassengerinonecontextanddriverinanother).Cancercells

thus fulfill the minimal requirements for evolution by natural selection; they are populations of cells that

reproducewithsomeheritablevariationinfitness(Lewontin1970;1978).TakingtheCSCmodel intoaccount

makes some difference in understanding evolution in cancer cells because it indicates an inequality in the

contributionofcancercellstoclonalevolution.TheCSCmodeldistinguishestwofunctionallydistinctpopulations

ofcells,onewithavirtuallyunlimitedproliferativeability(theCSCs),andtheotherwithalimitedproliferative

ability (the non-CSCs) (Fig. 1b). On short time scales (days to weeks) this functional distinction makes no

difference.Buton the timescaleof thedisease (months toyears), itmakesamajordifferenceasmutations

occurringinnon-CSCsarewashedawayfromthepopulationthroughexhaustionoftheproliferativeabilityof

thesecellsandultimatelycelldeath.Non-CSCsareevolutionarydead-endsandonlythemutationsoccurringin

CSCscanpersistovertimeandgeneratelong-standingsubclones(i.e.thevariouspopulationsofcellswiththe

samepoolofmutationsthatconstituteagivencancer)(Fig.2a-b).Theproportionofmutationsthatwillbeable

toparticipateinCEoverthecourseofthediseasedependsbothonthefrequencyofCSCsinthepopulation(Fig.

2c)andtheprobabilityofdedifferentiationofnon-CSCs(Fig.2d).ThisledGreavestoarguethatCSCsarethemain

unitsofselectionincancers(GreavesandMaley2012;Greaves2013,2015).Thequestionthenis:whatdifference

doesitmakeforCE?Germain(2012)offeredananswer:CSCrestrictionlimitsevolutionbynaturalselectionand

makescomplexadaptationsunlikely.

ThemainaimofGermain(2012)wastoassesstheroleofevolutionbynaturalselectionincancers.Toachieve

thisgoal,heusedGodfrey-Smith’s(2009)famousaccountofDarwinianpopulationsandappliedittocancers.

The importance ofGodfrey-Smith’s framework forGermainwas that it allows amore precise description of

differenttypesofDarwinianpopulationsasfunctionsofroughlymeasurableparameters;thetwoextremetypes

of Darwinian populations being described as paradigmatic Darwinian populations andmarginal Darwinian

populations.Evolutionbynaturalselectioncanoccurinbothbutplaysamuchgreaterroleintheformer.The

generalconclusionofGermainisthatalthoughcancercellsareDarwinianpopulations,theyarenotparadigmatic

populationsbecausethereareanumberofparametersforwhichtheyscoreratherlow(seeTable1inGermain

2012).My interesthere is in the impactofCSC restrictiononevolutionbynatural selection,aquestion that

GermainaddressedbyscoringcancercellpopulationswithandwithouttakingintoaccounttheCSCmodel.For

oneparameter,reproductivespecialization,takingtheCSCmodelintoaccountmakesalargedifferenceinhow

thecancercellsscore,andthereforeonhowDarwiniantheyare.


6

Figure2CEwithandwithouttakingintoaccountCSCrestriction.(a)IntheclassicalCEmodel,whichdoesnottakeCSCsintoaccount,allcancercellsareconsideredtobeunitsofselection.(b)WhenCSCrestrictionistakenintoaccount,onlytheCSCsareconsideredasmeaningfulunitsofselectionthroughoutthecourseofthedisease,becausethemutationsthatoccurinnon-CSCsarewashedawayfromthepopulation.(c)CSCrestrictiondependson the frequency of CSCs and can vary from very low to very high, which differentially affects CE. (d) CSCrestrictionalsodependsontheprobabilityofdedifferentiationofnon-CSCsintoCSCs,asmutationsoccurringinnon-CSCs are not washed away if the mutated cells dedifferentiate into CSCs. High or low probability ofdedifferentiationwilldifferentiallyaffectCE

Reproductive specialization refers to the germ/soma distinction and related phenomena in which only a

sequesteredpartoftheindividualcangiverisetoanewindividualthroughsexualorasexualreproduction(the

germ cells), while the rest of the individual cannot (the somatic cells) (see Godfrey-Smith 2009, p. 92). In

multicellularorganisms,reproductivespecializationpartiallysuppressesevolutionbynaturalselectionatthecell

levelbymakingthesomaticcellsevolutionarydead-ends(seeGodfrey-Smith2009,p.101).Germainarguedthat

the coexistence of CSCs and non-CSCs in cancers produces a reproductive specialization comparable to the

germ/somadistinction:non-CSCsareunabletogrowatumor likesomaticcellsareunabletoproduceanew


7

organism,whereasCSCsareable togrowa tumor likegermcells are capableofproducinganeworganism.

Greavesmadeasimilarcomparison:

Darwinianmodelsofcancernowneedtoadopttheconceptofcancerstemcells;thatsmallpopulationof

self-renewingcellsthatmaintainmostofthecancercellpopulationand,inself-replicating,providethe

essentialreservoirforfurthergeneticvariabilityandselection,equivalentinevolutionarytermstogerm

cells(Greaves2007,218).

GreavesandGermainagreeonthephenomenon:CSCrestrictionisamechanismbywhichanumberofmutations

arewashedawayfromthepopulation(allthosethatoccurinnon-CSCs,dedifferentiationeventsapart,seeFig.

2).Thisphenomenonalsoappliesinsomaticcellsofnormaltissueswheremutationsoccurringinnon-stemcells

areremovedfromthepopulationduetoreplicationlimitsofcells(againwiththeexceptionofdedifferentiation),

and it isnowaclassicargument thatstemcells limitcancer risk (Cairns1975;Pepperetal.2007).3Whether

reproductivespecializationaccuratelycapturesthisphenomenonisaquestionthatIwilladdresslater(section

2.2).TheimportantconclusionhereisthatthehighertheCSCrestriction (i.e.thelowertheCSCfrequencyandtheprobabilityofnon-CSCdedifferentiation),thelesscancercellsareparadigmaticDarwinianpopulations.

From this first conclusion, a second conclusion follows in Godfrey-Smith’s framework: the less paradigmatic

cancercellpopulationsare, the less likelycomplexadaptationsare tooccurbecauseparadigmaticDarwinian

populationsarecharacterizedasthoseinwhichnaturalselectioncanexplaintheoriginofnoveltraits(complex

adaptations) whereas natural selection only changes the distribution of already existing traits (simple

adaptations)inmarginalDarwinianpopulations(seeGodfrey-Smith2009,chapter3).Complexadaptationsrely

oncumulativeevolution:

Itcanseemoddtosaythatselection,whichhastodowithsortingthingsthatalreadyexist,cansomehow

bringnewthings intoexistence.Butnaturalselectioncanreshapeapopulation inawaythatmakesa

givenvariantmorelikelytobeproducedviatheimmediatesourcesofvariationthanitotherwisewould

be.Selectiondoesthisbymakingintermediatestagesontheroadtosomenewcharacteristiccommon

ratherthanrare,thusincreasingthenumberofwaysinwhichagivenmutationalevent(orsimilar)will

suffice toproduce the characteristic inquestion. Somekindsofnovelty canbeproducedeasilybyan

evolutionaryprocesswithoutthisroleforselection,butotherkinds—complexandadaptedstructures—

cannot(Godfrey-Smith2009,43).

CSCrestrictionimpedescomplexadaptationsbylimitingcumulativeevolutionintwoways(Germain2012).First,

CSCrestrictionlimitsthesizeofthecancercellpopulationinwhichevolutionarychangescanaccumulate(Fig.

2).Anycomplexadaptationsinnon-CSCsareveryunlikelytooccurgiventhatnon-CSCscanonlyproliferatefor

shortperiodsoftime,andthusmutationswouldbequicklylost.Second,CSCscanbequiescent(i.e.nondividing

forlongperiodsoftime),whichlimitsthepossibilityofcumulativeevolutionintheCSCpopulationitself.

3TomasettiandVogestein(2015)arguedthat“variationincancerriskamongtissuescanbeexplainedbythenumberofstemcelldivision,”whichstartedamassivedebateoncanceretiologyandonthecausalcontributionattributabletoenvironment,heredity,andstemcellreplication(seethelatestresponseofTomasettietal.2017).


8

GermainconvincinglyarguedthattheCSCmodel,andmorepreciselywhatIcallthelevelofCSCrestriction,plays

animportantroleinwhatwecanexpectfromevolutionbynaturalselectionincancers.Inparticular,thehigher

theCSCrestriction,thelesslikelycomplexadaptationsaretoappear(Figure3).4Moregenerally,CSCrestriction

isawaytowashoutagreatamountofthemutationsthatoccurinagivencellpopulation.Thisnotonlylimits

evolutionbynaturalselectionbutlimitsCEingeneral.

Figure3AdaptationsdependonCSCrestriction.ThelowertheCSCfrequencyandthelowertheprobabilityofnon-CSCdedifferentiation,themoreunlikelycomplexadaptationsaretooccur.Theblackandwhitegradientindicatesdirections:thelowertheCSCfrequencyandtheprobabilityofdedifferentiationofnon-CSCsintoCSCs,the less likelycomplexadaptationsare.Theexactshapeofthegradientremainstobeempiricallyevaluated.Noticealsothatsomevaluesarebiologicallyabsurdorequivalent.Forexample,havingaCSCfrequencyof100%andaprobabilityofdedifferentiationof1isabitabsurd,becauseifthereareonlyCSCsthentherearenonon-CSCstodedifferentiate.Similarly,iftheprobabilityofdedifferentiationis1thenCSCfrequencycannotbe0%

1.2.CSCrestrictionfavorsdrift

Germain’sinvestigationfocusedonevolutionbynaturalselection.AsCEisnotrestrictedtoevolutionbynatural

selection,Iwouldliketocontinuehisinvestigationatalargerscale.Thissectionfocusesondrift.TheCEmodel

wasoriginallyframedwiththeideathatcancerisaDarwinianprocessofgradualevolutionbynaturalselection.5

Yet,sinceGermain’sarticlewaspublished,evidenceforneutralevolutionhasbeenreportedinhepatocellular

carcinoma(Lingetal.2015)andcolorectalcancers(Sottorivaetal.2015;Sottorivaetal.2017).Applyinganeutral

evolutionmathematicalmodeltothesequencingdataof904cancersfrom14types,Williamsetal.(2016)found

that the neutral model fitted with high precision 323 of them (more than a third), suggesting that neutral

evolutioncouldaffectmanymorecancersthananticipated.Inthesecancers,theyargue,mostofthemutations

responsibleforthecancerexpansionwerealreadypresentinthefirstmalignantcellandthefollowingevolution

wasneutral,aphenomenonsometimereferredtoasa“BigBang”modelofcancerprogression(Sottorivaetal.

4Ifocusonmutationsbecausetheyaretheusualtraitsthatareusedtoreconstructandstudytheevolutionofcancercellpopulations,buttheargumentwouldapplyforanyheritabletrait.5HereIfocusontheassumptionthatCEisaprocessofevolutionbynaturalselectionbecause,tothebestofmyknowledge,noevidencesuggeststhatCSCrestrictioncanfavororlimitpunctuatedorgradualevolution.ButtheassumptionthatCEisagradualprocesshasbeenchallengedbytheobservationofseveralcomplexmutationaleventsthatinducemultiplemutationsinaburst(e.g.Stephensetal.2011;Rauschetal.2012;Nik-Zainaletal.2012a;Bacaetal.2013).TheextentandlimitsoftheanalogywithEldredgeandGould’spunctuatedevolutionisdiscussed in Gao et al. (2016). Markowetz (2016) suggested that saltationist theories such as the hopefulmonsterstheoryofRichardGoldschmidtmightbemorerelevantincancers.


9

2015;Amaroetal.2016).6Anotherpan-analysisalsohighlightedthatnegativeselection,whichispredominant

inthegermline,isnearlyabsentincancerandsomaticevolution(Martincorenaetal.2017).DoesCSCrestriction

impactwhether cancers followapatternofneutral evolution?AmodeldevelopedbySottorivaet al. (2010)

suggestsasmuch.Inthismodel,SottorivaandcolleaguesobservedthatCEwasimpactedintwowayswhenthey

factoredintheCSCmodel(i.e.bydistinguishingtwofunctionallydistinctpopulationsofcancercells,CSCsand

non-CSCs). First, as alreadydiscussed in section1.1,CSCs repressCEbymaking theacquisitionofmutations

slower. Second, they observed a different clonal expansion process.When the CSCmodel is not taken into

account,asmallnumberoftheoccurringmutationswereselectedforandinvadedlargeportionsofthecancer

cellpopulation.Incontrast,theimplementationoftheCSCmodelresultedinthepersistenceofalargeproportion

of mutations that remained at a low frequency in the population, consistent with neutral evolution. They

concluded“thattheintrinsicpropertiesoftheCSCmodelmightpropelanalternativeprocesstonaturalselection,

referredtoasgeneticdrift”(Sottorivaetal.2010,52),andattributedtheoccurrenceofdrifttosamplingerrors

thataremuchmorefrequent insmallpopulations—theCSCmodelstimulatessamplingerrormechanismsby

limitingtheproportionofcellsthatcanacquireandmaintainmutations.

TheconclusionofSottorivaetal.(2010)thatCSCrestrictionfavorsdriftraisesthequestionofwhetherthereis

higher CSC restriction in the cancers that fitted the neutralmodel inWilliams et al. (2016). To answer this

question,onewouldhavetoaligntheanalysisofneutralevolutionfromwholegenomesequencingdatawith

scoringsof CSC frequency andprobability of dedifferentiation in various cancer types andalso in cancers at

variousstagesofprogression.ThiswouldshowhowachangeinCSCrestrictioncanimpacttheprocessesofCE

(Fig.4).

Figure4CSCrestrictionanddrift.Theaxesinthisschematicdiagramarethesameasthepreviousdiagram(Fig.3).LowerCSCfrequencyandprobabilityofdedifferentiationnotonlymakecomplexadaptationsveryunlikely,they also favor drift. Empirical comparison between processes of evolution and level of CSC restriction arerequiredtocharacterizetheCEprocessesassociatedwitheachofthepointsofthistwo-dimensionalspace

1.3.CSCrestrictionfavorslinearevolution

6 Whether a first stage of evolution by natural selection precedes neutral evolution is a matter of debate(Martincorenaetal.2015;Simons2016).


10

TheCEmodelwasfirstrepresentedasalinearmodelofevolutionwhereinmutationsaccumulatesequentially,

witheachnewcloneoutcompetingthepreviousone,sothatsubclonesaccumulatingnewmutationsproceed

one after another (Fig. 5a). In this pattern of CE, intratumoral heterogeneity (ITH) is diachronic rather than

synchronic, such that cells accumulate mutations through time but the mutations appear homogenous

throughoutthecellpopulationatanygiventime(i.e.atT1allcellshavemutation1,atT2allcellshavemutations

1and2,etc.)(e.g.Houetal.2012).Inmostcasesoflinearevolutionthough,selectivesweepsappearincomplete

andformersubclonespersist,butdonotgiverisetonewsubclones(Fig.5b)(e.g.Merlevedeetal.2016).When

theydogiverisetonewsubclones,theresultisabranchedpatternofevolution:mutationsrandomlyaccumulate

incellsofvarioussubclonescontinuouslyleadingtotheappearanceofnewsubclones(Fig.5c)(e.g.Yachidaetal.

2016),which is themost commoncase (Andoretal.2016). In this section Idevelop thehypothesis thatCSC

restrictionmightalsoimpactpatternsofevolution,andmorepreciselythatahigherlevelofCSCrestrictionmight

favorlinearevolution.

Figure5ThreepatternsofCE. (a)LinearhomogeneousCE ischaracterizedbythesequentialaccumulationofmutations with each new clone outcompeting the previous so that subclones accumulating newmutationssucceedoneanother.(b)Whenselectivesweepsareincompleteandformersubclonespersistwithoutgivingrisetonewsubclones,theCSislinearheterogeneous.(c)Whenformersubclonesaccumulatemutationsandgiverisetonewsubclones,CE isbranched:mutations randomlyaccumulate incellsofvarioussubclonescontinuouslyleadingtotheappearanceofnewsubclones

Withsimilarcelldivisionandmutationratesinafixedperiodoftime,asmallerpopulationwillaccumulatefewer

mutationsthanabiggerpopulation.Thisistruebydefinitionifmutationsoccurduringdivision.Whenamutation

doesoccur,allthingsbeingequal,itshouldalsohaveahigherprobabilityofinvadingthepopulation,asthereare

fewercellstooutcompete.Thereisthusaninversecorrelationbetweentheprobabilityofoccurrenceofnew

mutationsand theprobabilityofpropagationof themutations. Ifmutationsare less frequentandpropagate

moreeasilyinasmallerpopulation,thenthereisahigherchancethatthesecondmutationoccursontopofthe


11

firstmutation(becausethefirstmutationispresentinalargerfractionofthepopulationatthetimeofoccurrence

ofthesecondmutation).Ifthesecondmutationdoesoccurontopofthefirst,andthethirdontopofthesecond,

thenwehavelinearevolution.Incontrast,inaverylargepopulation,mutationspropagatelesseasilyandoccur

morefrequently,leavinglesstimeforthepreviousmutationtopropagate,andreducingthechancethatanew

mutationoccursinacellbearingthepreviousone.Ifthenewmutationoccursinacellthatdoesnothavethe

previousmutation,thenwehavebranchingevolution.Thus,ceterisparibus,smallerpopulationsaremorelikely

toundergolinearevolutionandlargerpopulationsaremorelikelytoundergobranchingevolution.AsonlyCSCs

canparticipateinCEoverthecourseofthedisease,itisactuallytheCSCrestrictionthatmattersforpatternsof

CE.HigherCSC restriction should thus favor linearCE,whereas a loss of CSC restrictionby an increasedCSC

frequency and/or an increased probability of dedifferentiation should increase the probability of branching

evolution(Fig6a).

TherelationshipbetweenCSCrestrictionandpatternsofCEhasnotyetbeenstudied,butthereissomeindirect

evidenceinfavorofthishypothesis.First,tothebestofmyknowledge,allcasesoflinearevolutionthathave

beenreportedare inbloodcancers(supplementarytable1,grey lines),whicharecharacterizedbya lowCSC

frequencyandnodedifferentiation,thusahighCSCrestriction.TheonlyexceptioncomesfromShainetal.(2015)

who reported linear evolution in thepremalignant stages ofmelanocytic neoplasms, suggesting that in solid

tumors,theinitialpremalignantstagecouldfollowalinearpatternofevolutionbeforeexplodingintoabranched

patternofevolutionatinitiationofthemalignantstage(whichwouldbeinlinewiththefactthatcancersstart

fromthemutationofasinglecellandthuswithasmallpopulationofCSCs)(Fig6b).Cancersinwhichnon-CSCs

candedifferentiateandCSC frequency ishigher, suchasbreast cancers, coloncancers, advancedmelanoma,

brain cancers, osteosarcoma, and pancreatic cancers, follow branched patterns of evolution (supplementary

table1).Thesecondkindofevidencecomesfrompatternsofaccumulationofmutationsinnormaltissues(i.e.

withoutanyovertcancerbeingdiagnosed).Presenceofmutationsusuallyassociatedwithcancerswithoutovert

malignancieshavebeenwelldocumented in twotissues:skinof theeyelidandblood.Stemcell restriction is

muchlowerintheformertissuethaninthelatter(higherstemcellfrequencyanddedifferentiationhavebeen

reportedinskinepithelium).Interestingly,thepatternsofmutationsinthesetissuesareverydifferent.Inthe

eyelid,Martincorenaetal.(2015)observedahighrateofmutationsgivingrisetonumerousbutverysmallclones

(i.e.cloneswithsmallnumbersofcells).Incontrast,inthebloodveryfewmutationshavebeenobserved,but

thesegiverisetobiggerclones(Genoveseetal.2014;Jaiswaletal.2014;Xieetal.2014).

Toconcludethisfirstsection,CSCrestrictionlimitsevolutionbynaturalselection,andmoregenerallyCE,prevents

complexadaptations,andfavorsdriftandlinearevolutioninamannerthatdependsonCSCfrequencyandthe

probabilityofdedifferentiation(Fig3,4and6).ByignoringtheCSCmodelorbyconsideringitasanindependent

modelofcancer,theCEmodelmissestheseimpactsofCSCrestrictiononCEcharacteristics.


12

Figure 6 Patterns of evolution and CSC restriction. (a) Lower CSC frequency and lower probability ofdedifferentiationfavorslinearevolution.(b)Duringdiseaseprogression,cellpopulationsmightmoveinthistwo-dimensionalspace.AlossofCSCrestrictionthroughanincreaseinCSCfrequencyand/orincreaseinprobabilityofdedifferentiationmightfavoratransitionfromalinearCEtoabranchedCE

2.TraditionalparametersofevolutiondonotaccuratelydescribetheconsequencesofCSCrestrictionforCE

ThefirstsectionshowedthattheCEmodelshouldtake intoaccounttheCSCmodel,andmorepreciselyCSC

restriction,becauseitcanaffectboththepatternsandtheprocessesofCE.ThisdistinctionbetweenCSCsand

non-CSCsandits impactonCEresemblesomephenomenaalreadywellknownanddescribedinevolutionary

biology.Scientistsandphilosophershavediscussedtheresemblancewithvariousparametersofevolutionsuch

asheredity(Greaves2013),reproductivespecialization(Germain2012;Greaves2013),andeffectivepopulation

size(e.g.Merloetal.2006;Pepperetal.2009;Sottorivaetal.2010;Lipinskietal.2016;LeanandPlutynski2016).

Thissectionpresentsanddiscussesthesecomparisonsaswellasadditionalones(bottleneckanddependenceof

fitnessonintrinsicproperties).Iarguethatnoneoftheseparametersareentirelysatisfactorytodescribethe

impactofCSCrestrictiononCEandshowthatthedifficultiescomefromthefactthatCSCrestrictiondependson

twofactors:theCSCfrequencyandtheprobabilityofdedifferentiationofnon-CSCsintoCSCs.

2.1.CSCrestrictionandheredity

ReferringtoLewontin’sclassicalminimalrequirementforevolutionbynaturalselection,Greaveshasarguedthat

itisthelackofhereditythatsuppressesCEinnon-CSCs,sothatonlyCSCsaregenuineunitsofselectionincancers

(e.g.Greaves2013,104).Thisclaimreliesonamisunderstandingoftheparameterofheredity.Heredity,asa

parameterofevolution,referstothefidelityofheredityateachgeneration.Sowhatisscoredinheredityisthe

“similaritybetweenparentandoffspring,duetocausalroleoftheparents”(Godfrey-Smith2009,39).Incancers,

the fidelity of heredity largely depends on the quantity ofmutations,which is highly variable. For example,

comparing30cancertypes,Alexandrovetal.(2013)reportedamutationalloadrangingfromabout0.001/Mbto

more than400/Mb.Whereasmelanomacanpresent thousandsofmutations, and thusavery low fidelityof

heredityduringcelldivision,veryfewmutations,ifany,werereportedinependymomas(apediatrictumorofthe

centralnervoussystem) indicatingahighfidelityofheredity (Macketal.2014).Thesedifferencesdependon

severalfactorssuchasgeneticinstability,orexposuretomutagenicagents.Forexample,thefidelityofheredity


13

isfarlowerinlungcancersofsmokersthanofnon-smokers(Alexandrovetal.2016).CSCrestrictionplayslittle

role infidelityofheredity.WhatGreavesmeantby lackofhereditywasthatnon-CSCscanonlygiverisetoa

limitednumberofgenerationsofcellsandthatonlyCSCscanreproduceinthelong-term,afactweallagreeon

butstruggletodepictappropriately.

2.2.CSCrestriction,reproductivespecializationandbottleneck

ReferringtoGodfrey-Smith’snowfamousparametersofevolution,Germain(2012)claimedthatCSCrestriction

changesthedegreeofreproductivespecialization(seesection1.1).Iagreethatthereisananalogybetweengerm

linesequestrationinmulticellularorganismsandCSCrestrictionincancers.Thereis,inbothcases,adivisionof

labor thathasmassive impactson the long-term fitnessof the cells.But this analogyhas several limitations.

Reproductive specialization refers to thegerm/somadistinction,or equivalentphenomena. It relieson three

conditions. There is a reproductive specializationwhen: (1) there is reproduction; (2) only a part of the cell

populationisinvolvedinthisreproduction;and(3)thesecellsarethegermcells,whicharedistinctfromsomatic

cells,thatis,theybelongtoaseparatedcellline.

Whetherthereisreproductionwithincancersisadebatablematter.Metastasescouldbeacandidate(seeLean

andPlutynski2016;GermainandLaplane2017),butmyargumentisthatCSCrestrictionimpactspatternsand

processesofCEatthecellularlevel,regardlessoftheformationofmetastases.Onecouldthenarguethatcell

lineagesthatmakeupeachtumorare individualsthatreproduce. Inasense,eachCSCgivesrisetoa lineage

containingabodyofnon-CSCs.WhenaCSCself-renewsandproducesanewCSCthatgivesrisetoanewlineage,

thendowehaveacaseofreproduction?Whethereachlineagecancountasanindividualseemsdebatableto

megiventheabsenceofintegration.7But,ifweagreethatthereisagenuinecaseofreproductionhere,thenwe

alsomeetthesecondcriteriaforreproductivespecialization,namelythatonlyapartofthecellsconstitutingthe

individualareinvolvedinthereproduction,heretheCSCs.However,wethenfacealimitationintheanalogywith

regardtothethirdcriterion:CSCsandnon-CSCsarenottwoseparatedcelllineslikesomaticandgermcellsare.

Whilegermlinecellsaresequestered,CSCs(aswellasnormalstemcells)arenot,astheCSCsgiverisetothe

non-CSCs.ForCE,thismightbeconsideredasanegligibleproblemwhentheprobabilityofdedifferentiationof

non-CSCsintoCSCsisnull.Butitbecomesamajordifferenceassoonasnon-CSCscanbecomeCSCs.Inthiscase

thedistinctionbetweengermlineandsomacanno longerapply. If there isreproduction,then it isofavery

differentkind,onethatwouldinvolveabottleneck.

Abottleneckiswhenreproductionoccursfromonlyasmallpartoftheparentalorganism(seeGodfrey-Smith

2009,chapter5).Atfirstsight,bottleneckandreproductivespecializationmightseemverysimilar,asbothare

processesthatwashawaysomemutations,whichisexactlywhattheCSCvs.non-CSCdistinctiondoes.However,

theydifferintheirconsequencesforevolution:reproductivespecializationpreventsanymutationoccurringin

thesomaticcells/non-CSCsfrombeingtransmittedtothenextgeneration,whereasabottleneckdoesnotas

virtuallyanycellcanbetransmittedtothenextgeneration.Thus,incaseswherenon-CSCscandedifferentiate

7TherearesomefeedbackloopsbetweenmaturecellsandCSCs(e.g.Reynaudetal.2011),butthosearenotrestrictedtotheirownlineagesandinsomecases,likebloodcancers,thecellsfromdifferentlineagesareallmixedandmovingaround.


14

intoCSCs,thequestionbecomeswhetherCSCrestriction(CSCfrequencyandprobabilityofdedifferentiation)

equates to the degree of bottleneckishness. If each CSC lineage counts as an individual, then the degree of

bottleneckishness isalwaysthesame:eachindividualstartswithonlyoneCSC,whatevertheprobability isof

dedifferentiation.IfthedegreeofbottleneckishnessisalwaysthesamewhiletheCSCrestrictioncanvary,then

itcannotdescribethedegreeofCSCrestriction.

Hence,thereproductivespecializationparametercouldonlycapturetheimpactofCSCrestrictiononCEincases

wheretheprobabilityofdedifferentiationisnull,withthelimitationsintheanalogythatCSCsandnon-CSCsare

notseparatelineagesandprovidedthatlineagesdoreproduce.OnecouldthenarguethatthefrequencyofCSCs

equates to the degree of reproductive specialization. But CSC restriction also depends on the probability of

dedifferentiation,andwhennon-CSCscandedifferentiateintoCSCs,thenthereisnoreproductivespecialization,

butratherabottleneck.However,asthedegreeofbottleneckishnessisstable(onecell,theCSC),itisunlikelyto

do a better job of capturing the impact of CSC restriction on CE, which changes with the probability of

dedifferentiationandCSCfrequency.8

2.3.CSCrestrictionanddependenceoffitnessonintrinsicproperties

Besidesbeingacaseofreproductivespecialization,Godfrey-Smith(2009)alsodescribesgermlinesequestration

asacaseoflowdependenceoffitnessonintrinsicproperties,whichquantifieshowmuchreproductivesuccess

dependsonintrinsicproperties(e.g.thegeneticcontentofthecells)versusextrinsicproperties(e.g.thelocation

inaparticularenvironment).Lowdependenceoffitnessonintrinsicpropertiesrepressesevolutionbynatural

selection,asinthecaseofgermlinesequestration,becausethedifferenceinthelong-termfitnessofthegerm

linecellscomparedtosomaticcellsisnotduetointrinsicproperties(thatcanbepositivelyselectedfor)butto

thefactthatthecellsendedupbeingpartofoneoranothergroupofcellsduringdevelopment.Ifreproductive

specializationdoesnotexactlycaptureCSCrestriction,candependenceof fitnesson intrinsicpropertiesdoa

betterjob?ThequestionhereiswhetherthedifferenceinfitnessbetweenCSCsandnon-CSCsisduetocontext

orintrinsicproperties.Inotherwords,isCSCidentityintrinsicorextrinsic?

Ifnon-CSCscandedifferentiateintoCSCs,itmeansthatundercertaincircumstancesinsomecancers,stemness

can be acquired. But the ability or not to dedifferentiate is not sufficient to answer our question, as the

microenvironmentcanplayvaryingrolesinbothcases.Inpreviouswork,Ihaveshownthattwoquestionsmust

bedistinguishedtounderstandtheidentityofstemcells,includingCSCs:(1)cannon-stemcells/non-CSCsacquire

stemnessanddedifferentiateintostemcells/CSCs?(2)Isstemnessacquisitionand/ormaintenancedependent

uponthemicroenvironment?Dependingontheanswerstothesequestions,whicharespecifictotissuesand

cancers,fourdifferentidentitiescanbedistinguishedforthepropertyofstemness(Laplane2016):

1. Categorical:stemnessispurelyintrinsic.

8Iamverygratefultobothanonymousreviewersforthethoroughdiscussionthathasleadtothisversionoftheargument.


15

2. Dispositional:stemnessisanintrinsicpropertywhoseexpressiondependsonextrinsicstimulifromthe

microenvironment.

3. Relational:stemnessisanextrinsicpropertyinducedandcontrolledbythemicroenvironment.

4. Systemic:stemnessisanextrinsicpropertycontrolledatthecellpopulationlevel.

Inthelastthreecases,thefitnessoftheCSCsdepends,tovariousdegrees,oncontext(whichcanincludethe

tumor-microenvironment,clone-cloneinteractions,andfeedbackloopsbetweenmaturecellsandCSCs).Incases

wherestemnessisarelationalproperty,beingaCSCdependsonthelocationofthecancercells.Dependenceof

fitnessonintrinsicproperties,inthisinstance,isverylow.Itishigherincaseswherestemnessisadispositional

propertybecausebeing aCSCdependson intrinsicproperties, although it is still limitedby the fact that the

reproductivesuccessofeachCSCisregulatedbythemicroenvironment.Incaseswherestemnessisasystemic

property,beingaCSCdependsonvariousfactorsincludingintrinsicpropertiessuchasstochasticgeneexpression

andextrinsicpropertiessuchasfeedbackloopsandothercollectiveprocesses;here,dependenceoffitnesson

intrinsicpropertieshangsupon the relativecontributionof intrinsicandextrinsic factors in stemness.Finally,

dependenceoffitnessonintrinsicpropertiesishighincaseswherestemnessisacategoricalpropertybecause

thedifference in fitness in the long term ismainlydue to intrinsicproperties. Thus, the identityofCSCs can

provideinterestinginsightsforCE,forexample,cancersinwhichstemnessisacategoricalpropertyshouldbe

morelikelytoevolvebynaturalselectionthancancersinwhichstemnessisarelationalproperty.But,thedegree

ofdependenceoffitnessonintrinsicpropertiesdoesnotcorrelatewiththedegreeofCSCrestriction.

2.4.CSCrestrictionandeffectivepopulationsize

SomebiologistsandphilosophershavesuggestedthatCSCrestrictionsimplylimitstheeffectivepopulationsize,

asopposedtothecensuspopulationsize(e.g.Merloetal.2006;Pepperetal.2009;Sottorivaetal.2010;Lipinski

etal.2016;LeanandPlutynski2016).

Infact,thequestionofwhethertheentireneoplasmoraminorityofneoplasticcellsiscapableof

self-renewalis,atleastinpart,aquestionmerelyoftheeffectivepopulationsizeoftheevolving

cellsinaneoplasm(Pepperetal.2009,66).

TheclaimhereisthattheeffectivepopulationsizeisthesizeoftheCSCpopulation.ThelowertheCSCfrequency

is,thegreaterthedifferenceisbetweentheeffectivepopulationsizeandthecensuspopulationsize,andthe

smallertheeffectivepopulationsizeis.Reducingtheeffectivepopulationsizeimpactsevolutionaryprocesses:

smallerpopulationsareconsideredmorepronetodrift,assamplingerrorcanmoreeasilyleadtotheexpansion

ofcloneswithnoclearsurvivalbenefit (seesection1.2).9Moregenerally, thearguments thatCSCrestriction

represses CE, favors drift, prevents complex adaptations, and favors linear evolution, could be reduced to a

question of effective population size, if the probability of dedifferentiation is null. But if non-CSCs can

dedifferentiate into CSCs at any time, then the actual CSC frequency poorly represents the cells that can

9The idea thatdriftdominates selection in smallpopulationshasbeendebated, see forexample the recentdebatebetweenClatterbuck,SoberandLewontin(2013)andBrandonandFleming(2014).


16

meaningfullycontributetoCE,andthusalsopoorlyrepresentstheeffectivepopulationsize.Theprobabilityof

dedifferentiationofnon-CSCsintoCSCsisthenoncemoreamajorfactorforCE.

Takentogether,thesedifficultieshighlightthatCSCrestrictionisnoteasilycapturedbyanyclassicalparameter

ofevolution,inparticularbecauseofthedifferencebetweenCSCfrequencyanddedifferentiation,aswellasthe

fourdifferentpossibleidentitiesofCSCs(seesection2.3).ToaccountfortheimpactofCSCrestrictiononCE,one

needstomeasureboththefrequencyofCSCsandtheprobabilityofdedifferentiationofnon-CSCs.

3.BiomedicalrelevanceoftheimpactofCSCrestrictiononCE

ThefirstsectionshowedthatthedegreeofCSCrestrictioncouldfavororimpedevariousprocessesandpatterns

of evolution, and in particular that it can favor linear or branched evolution. Scientists regularly investigate

whetherCEislinearorbranchedbutgivelimitedattentiontothedifferenceitmakes.Whyshouldwecareabout

the impact of CSC restriction on linear or branched evolution? In this section, I argue that the relationship

betweenCSC restrictionandpatternsofCE isworthexploring for thebiomedical sciences, first,because the

patternsofCEcaninformusaboutCSCrestriction,andinparticularabouttheprobabilityofdedifferentiation

that is currently difficult to measure; second, because it offers original opportunities for new therapeutic

strategies.

3.1.FromCEpatternstoCSCrestriction

CEpatternsaremainlyusedtoinfereithertherelativefitnessofeachsubcloneinagivenenvironmentortheir

age.Incasesofevolutionbynaturalselectionthebiggersubclonesaretheoneswiththegreatestreproductive

success inthatenvironment. Incasesofneutralevolution,thebiggersubclonesaretheoldest. Inadditionto

theseinferences,IhavearguedinthispaperthatthepatternofCE—linearorbranched—is,atleastinpart,a

functionofCSCrestriction.IfIamright,thenreconstructionoftheCEcouldbeinformativeaboutCSCrestriction.

This couldbevaluable incancers inwhichoneof the twoparametersofCSC restriction—CSC frequencyand

probabilityofdedifferentiation—isunknown.Forexample,inasetofcancersofagiventype(e.g.melanomas),

ifwecanmeasuretheCSCfrequencyandreconstructtheCEtree,wemaybeabletoinferifdedifferentiation

occurredornotand,ifitoccurred,whetheritisamarginalorfrequentprocess(i.e.whethertheprobabilityof

dedifferentiation is high or low). This is far from trivial given that the current technologies allow far better

approximationsofCEpatternsandofCSCfrequenciesthanoftheprobabilityofdedifferentiation.

3.2.FromCSCrestrictiontoCEpatterns:therapeuticconsequences

What difference does it make if CE is linear or branched? Is one or the other better for the patients? The

immediate relevanceofdistinguishingCEpatterns is that thecoexistenceofmultiple subclones isacauseof

therapyresistance.Themoresubclonesthereare,thegreatertheriskoftherapyresistance.Ifmyhypothesis

thatahigherCSCrestrictionshouldfavoramorelinearandsimplerpatternofCEisright,thenpatientswould

benefit from any complementary therapies that can increase CSC restriction. Opportunities to control CSC

restrictionareemerging.ThemostrecentexamplesareCimminoetal.(2017)andAgathocleousetal.(2017),

whoshowedthatvitaminCregulatesthefrequencyofnormalormalignanthematopoieticstemcells.


17

Conversely,anytherapythatwoulddecreaseCSCrestrictionshouldbeavoided.Hypomethylatingagents(HMAs)

areusedasfirstlinetreatmentinchronicmyelomonocyticleukemia(CMML),myelodysplasticsyndromes(MDS),

andacutemyeloidleukemia(AML)oftheelderly,whenpatientsarenoteligibleforstemcelltransplantation.

ButHMAshave alsobeenused in a verydifferent context, unrelated to cancer, to induce cell plasticity and

dedifferentiation(e.g.Mikkelsenetal.2008;Chandrakanthanetal.2016).SomestudiesalsosuggestthatHMAs

candecreasestemcellrestrictioninthenormalhematopoieticsystem,eitherbyincreasingtheirfrequencyorby

inducingdedifferentiation (Suzukietal.2004;Chungetal.2009). If theyalsodecreaseCSC restriction in the

patients(aquestionIamcurrentlyexperimentallyinvestigatinginmylab),thenthepredictionofthisstudyis

thatHMAtreatmentmustfavormoreCE,moreadaptability,andmorecomplexpatternsofevolution.Thiscould

helptoexplainthetransiencyofpatients’responsestoHMAsandthefrequentprogressionofCMMLandMDS

intoAML.IfprogressionundertreatmentwereduetothelossofCSCrestriction,thenonewaytomaintainthe

benefit of the drug without its counterproductive effects would be to couple it with a drug increasing CSC

restriction.Manydrugs(nonetargetingCSCs)arecurrentlytestedincombinationwithHMAswithoutyetleading

toimprovementinoverallsurvival(Balletal.2017).Inthiscurrentclinicalcontext,myworksuggeststhatdrugs

actingonCSCrestrictionareworthinvestigatingtoo.

Conclusionandperspectives

WhiletheinclusionofCEintotheCSCmodelhasbeenfairlywelladdressedintheliterature(KresoandDick2014;

NassarandBlanpain2016),showinginparticularthatCSCsareaheterogeneoustargetforanti-CSCtherapies,

the impactof theCSCmodel onCEhasbeenpoorly explored. This article shows thatCSC restriction,which

dependsonCSC frequencyandtheprobabilityofnon-CSCdedifferentiation, impactsCE inseveralways.The

highertheCSCrestriction,themoreitcan:

• limitCE;

• limitevolutionbynaturalselection;

• preventcomplexadaptations;

• favordrift;

• favorlinearevolution.

A lossofCSC restriction (higherCSC frequencyand/orhigherprobabilityofdedifferentiation)duringdisease

progressionwillthusfavormoreadaptability,makingtheinclusionofCSCrestrictionintotheCEmodelhighly

relevantforbiomedicalresearch.

Incorporating CSC restriction into the CE model faces several difficulties, opening perspectives for further

philosophical and biological research. First,within current experimental limitations, it is difficult to properly

quantifythenumberofCSCsandtheprobabilityofdedifferentiationwithinacanceratanygiventime.ThusCSC

restriction canonlybe roughlyestimated.Current improvements in lineage tracing technologieswill provide

greathelp.Estimationoftheprobabilityofdedifferentiationofnon-CSCsintoCSCsisalsobecominganurgent

needthatremainstoopoorlyaddressed.


18

Second,recentworkhasshownthatunderhomeostaticconditions,progenitorcells(thefirstdaughtercellsof

stemcells),cansupporttheproductionofallthebloodcellsformuchlongerthaninitiallythought(Sunetal.

2014;Buschetal.2015).Thisraisesanewquestionforcancers:areCSCstheonlymeaningfulunitsofselection

inCE?Dependingonthedurationofthedisease,progenitorcellsmightalsobetakenintoaccountaspartofCSC

restriction,whichwouldchangethemeasureofCSCrestriction.

Third,theanalysispresentedinthisarticleismadeceterisparibus.Mypointisthateveryotherparameterbeing

equal,adifferenceinCSCrestrictionwillaffectCE.Afurtherquestionwouldbewhetherotherparameters,like

therateofmutationandofproliferation,aswellastheroleofthemicroenvironmentandmoregenerallythe

physicalconstraintsofthetissuesoutweighsomeoftheimpactofCSCsonthepatternsandprocessesofCE.

Finally,thisarticlefocusesontheimpactofCSCrestrictiononCE.Ifcorrect,thenthenextquestionishowcan

we act on CSC restriction in order to limit CE and disease progression? To answer this question requires

investigatingthetwofactorsofCSCrestriction:CSCfrequencyandnon-CSCdedifferentiation.Whatcausesan

increase inthefrequencyofCSCswithinagivencancer,andhowcanthis increase inthenumberofCSCsbe

avoidedorreduced?Whatinducesdedifferentiationofnon-CSCsintoCSCsandhowcanwepreventit?These

questions require a thorough investigation of the role of the microenvironment and of population-level

processes.ThemicroenvironmentcanplayadecisiveroleintheproliferationofCSCsandtheirabilitytogenerate

aclone(Arranzetal.2014;Dongetal.2016).Maturecancercells(non-CSCs)canalsoplayanimportantrolein

theproliferationoftheCSCsthroughthecytokinestheysecrete(Reynaudetal.2011;Welneretal.2015),raising

the question of interactions between subpopulations, andmore generally indicating the need to investigate

multilevelselectionprocesses(Sprouffskeetal.2013;LeanandPlutynski2016).

References

AdamsJM,StrasserA(2008)Istumorgrowthsustainedbyrarecancerstemcellsordominantclones?Cancer

Res68:4018–4021.doi:68/11/4018[pii]10.1158/0008-5472.CAN-07-6334

AgathocleousM,MeachamCE,BurgessRJ,etal(2017)Ascorbateregulateshaematopoieticstemcellfunction

andleukaemogenesis.Nature549:476–481.doi:10.1038/nature23876

AlexandrovLB, JuYS,HaaseK,etal (2016)Mutationalsignaturesassociatedwithtobaccosmoking inhuman

cancer.Science354:618–622.doi:10.1126/science.aag0299

AlexandrovLB,Nik-ZainalS,WedgeDC,etal(2013)Signaturesofmutationalprocessesinhumancancer.Nature

500:415–21.doi:10.1038/nature12477

AmaroA,ChiaraS,PfefferU(2016)Molecularevolutionofcolorectalcancer:frommultistepcarcinogenesisto

thebigbang.CancerMetastasisRev35:63–74.doi:10.1007/s10555-016-9606-4

AndersonK,LutzC,vanDelftFW,etal(2011)Geneticvariegationofclonalarchitectureandpropagatingcellsin

leukaemia.Nature469:356–361.doi:nature09650[pii]10.1038/nature09650

AndorN,GrahamTA,JansenM,etal(2016)Pan-canceranalysisoftheextentandconsequencesofintratumor

heterogeneity.NatMed22:105–113.doi:10.1038/nm.3984

Arranz L, Sánchez-Aguilera A,Martín-Pérez D, et al (2014) Neuropathy of haematopoietic stem cell niche is


19

essentialformyeloproliferativeneoplasms.Nature512:78–81.doi:10.1038/nature13383

BacaSC,PrandiD,LawrenceMS,etal(2013)Punctuatedevolutionofprostatecancergenomes.Cell153:666–

77.doi:10.1016/j.cell.2013.03.021

BallB, ZeidanA,GoreSD,PrebetT (2017)Hypomethylatingagent combination strategies inmyelodysplastic

syndromes:hopesandshortcomings.Leuk.Lymphoma58:1022–1036

BapatSA (2007)Evolutionof cancer stemcells. SeminCancerBiol17:204–213 .doi: S1044-579X(06)00039-3

[pii]10.1016/j.semcancer.2006.05.001

BatlleE,CleversH(2017)Cancerstemcellsrevisited.NatMed23:1124–1134.doi:10.1038/nm.4409

BolliN,Avet-LoiseauH,WedgeDC,etal(2014)Heterogeneityofgenomicevolutionandmutationalprofilesin

multiplemyeloma.NatCommun5:.doi:10.1038/ncomms3997

Bonnet D, Dick JE (1997) Human acutemyeloid leukemia is organized as a hierarchy that originates from a

primitivehematopoieticcell.NatMed3:730–737.doi:10.1038/nm0797-730

Brabletz T, JungA, Spaderna S, et al (2005)Opinion:migrating cancer stem cells - an integrated concept of

malignanttumourprogression.NatRevCancer5:744–749.doi:nrc1694[pii]10.1038/nrc1694

BrandonR,FlemingL(2014)Driftsometimesdominatesselection,andviceversa:areplytoClatterbuck,Sober

andLewontin.BiolPhilos29:577–585.doi:10.1007/s10539-014-9437-z

BrocksD,AssenovY,MinnerS,etal(2014)IntratumorDNAmethylationheterogeneityreflectsclonalevolution

inaggressiveprostatecancer.CellRep8:798–806.doi:10.1016/j.celrep.2014.06.053

BuschK,KlapprothK,BarileM,etal(2015)Fundamentalpropertiesofunperturbedhaematopoiesisfromstem

cellsinvivo.Nature518:542–546.doi:10.1038/nature14242

CairnsJ(1975)Mutationselectionandthenaturalhistoryofcancer.Nature255:197–200

Campbell LL, Polyak K (2007) Breast tumor heterogeneity: cancer stem cells or clonal evolution? Cell Cycle

6:2332–2338.doi:4914[pii]

CaoW,WuW, YanM, et al (2015)Multiple region whole-exome sequencing reveals dramatically evolving

intratumor genomic heterogeneity in esophageal squamous cell carcinoma. Oncogenesis 4:e175 . doi:

10.1038/oncsis.2015.34

CayeA,StrulluM,GuidezF,etal(2015)Juvenilemyelomonocyticleukemiadisplaysmutationsincomponentsof

theRASpathwayandthePRC2network.NatGenet47:1334–1340.doi:10.1038/ng.3420

ChafferCL,MarjanovicND,LeeT,etal(2013)PoisedChromatinattheZEB1PromoterEnablesBreastCancerCell

PlasticityandEnhancesTumorigenicity.Cell154:61–74.doi:http://dx.doi.org/10.1016/j.cell.2013.06.005

ChandrakanthanV,YeolaA,KwanJC,etal(2016)PDGF-ABand5-Azacytidineinduceconversionofsomaticcells

into tissue-regenerative multipotent stem cells. Proc Natl Acad Sci U S A 113:E2306-15 . doi:

10.1073/pnas.1518244113

Chesnais V, Renneville A, Toma A, et al (2016) Effect of lenalidomide treatment on clonal architecture of

myelodysplasticsyndromeswithout5qdeletion.Blood127:749–60.doi:10.1182/blood-2015-04-640128

ChungYS,KimHJ,KimT-M,etal(2009)Undifferentiatedhematopoieticcellsarecharacterizedbyagenome-

wideundermethylationdiparoundthetranscriptionstartsiteandahierarchicalepigeneticplasticity.Blood

114:4968–78.doi:10.1182/blood-2009-01-197780


20

Cimmino L,Dolgalev I,Wang Y, et al (2017) Restorationof TET2 FunctionBlocksAberrant Self-Renewal and

LeukemiaProgression.Cell170:1079–1095.e20.doi:10.1016/j.cell.2017.07.032

Clappier E,GerbyB, Sigaux F, et al (2011) Clonal selection in xenograftedhuman T cell acute lymphoblastic

leukemiarecapitulatesgainofmalignancyatrelapse.JExpMed208:653–661.doi:10.1084/jem.20110105

ClatterbuckH,SoberE,LewontinR(2013)Selectionneverdominatesdrift(norviceversa).BiolPhilos28:577–

592.doi:10.1007/s10539-013-9374-2

deBruinEC,McGranahanN,MitterR,etal(2014)Spatialandtemporaldiversityingenomicinstabilityprocesses

defineslungcancerevolution.Science(80-)346:251–256.doi:10.1126/science.1253462

DingL,LeyTJ,LarsonDE,etal(2012)Clonalevolutioninrelapsedacutemyeloidleukaemiarevealedbywhole-

genomesequencing.Nature481:506–10.doi:10.1038/nature10738

DongL,YuW-M,ZhengH,etal(2016)LeukaemogeniceffectsofPtpn11activatingmutationsinthestemcell

microenvironment.Nature539:304–308.doi:10.1038/nature20131

FabianA,BarokM,VerebG,SzollosiJ(2009)Diehard:arecancerstemcellstheBruceWillisesoftumorbiology?

CytomA75:67–74.doi:10.1002/cyto.a.20690

FrancisJM,ZhangC-Z,MaireCL,etal(2014)EGFRvariantheterogeneityinglioblastomaresolvedthroughsingle-

nucleussequencing.CancerDiscov4:956–71.doi:10.1158/2159-8290.CD-13-0879

GaoR,DavisA,McDonaldTO,etal(2016)Punctuatedcopynumberevolutionandclonalstasisintriple-negative

breastcancer.NatGenet48:1119–30.doi:10.1038/ng.3641

GenoveseG,KahlerAK,HandsakerRE,etal (2014)Clonalhematopoiesisandblood-cancerrisk inferredfrom

bloodDNAsequence.NEnglJMed371:2477–2487.doi:10.1056/NEJMoa1409405

Gerlinger M, Horswell S, Larkin J, et al (2014) Genomic architecture and evolution of clear cell renal cell

carcinomasdefinedbymultiregionsequencing.NatGenet46:225–33.doi:10.1038/ng.2891

GerlingerM,RowanAJ,HorswellS,etal(2012)Intratumorheterogeneityandbranchedevolutionrevealedby

multiregionsequencing.NEnglJMed366:883–92.doi:10.1056/NEJMoa1113205

GermainP-L(2012)Cancercellsandadaptiveexplanations.BiolPhilos27:785–810.doi:10.1007/s10539-012-

9334-2

GermainPL,LaplaneL(2017)Metastasisassupra-cellularselection?AreplytoLeanandPlutynski.BiolPhilos

32:281–287.doi:10.1007/s10539-016-9555-x

Godfrey-SmithP(2009)DarwinianPopulationsandNaturalSelection.OxfordUniversityPress,Oxford

GreavesM(2010)Cancerstemcells:backtoDarwin?SeminCancerBiol20:65–70.doi:S1044-579X(10)00010-6

[pii]10.1016/j.semcancer.2010.03.002

GreavesM(2013)Cancerstemcellsas“unitsofselection.”EvolAppl6:102–108.doi:10.1111/eva.12017

GreavesM(2007)Darwinianmedicine:acaseforcancer.NatRevCancer7:213–221.doi:10.1038/nrc2071

GreavesM(2015)Evolutionarydeterminantsofcancer.CancerDiscov5:806–20.doi:10.1158/2159-8290.CD-

15-0439

Greaves M, Maley CC (2012) Clonal evolution in cancer. Nature 481:306–313 . doi: nature10762 [pii]

10.1038/nature10762

HirschP,MamezAC,BelhocineR,etal(2016)Clonalhistoryofacordblooddonorcellleukemiawithprenatal


21

somaticJAK2V617Fmutation.Leukemia30:1756–1759.doi:10.1038/leu.2016.31

HouY,SongL,ZhuP,etal(2012)Single-cellexomesequencingandmonoclonalevolutionofaJAK2-negative

myeloproliferativeneoplasm.Cell148:873–85.doi:10.1016/j.cell.2012.02.028

HughesAEO,MagriniV,DemeterR,etal(2014)Clonalarchitectureofsecondaryacutemyeloidleukemiadefined

bysingle-cellsequencing.PLoSGenet10:e1004462.doi:10.1371/journal.pgen.1004462

ItzyksonR, KosmiderO,RennevilleA, et al (2013)Clonal architectureof chronicmyelomonocytic leukemias.

Blood121:2186–2198.doi:10.1182/blood-2012-06-440347

Jaiswal S, Fontanillas P, Flannick J, et al (2014) Age-related clonal hematopoiesis associated with adverse

outcomes.NEnglJMed371:2488–2498.doi:10.1056/NEJMoa1408617

JanM,SnyderTM,Corces-ZimmermanMR,etal(2012)Clonalevolutionofpreleukemichematopoieticstemcells

precedes human acute myeloid leukemia. Sci Transl Med 4:149ra118 . doi: 4/149/149ra118 [pii]

10.1126/scitranslmed.3004315

KimTH,YoshidaK,KimYK,etal (2015)Clonaldynamics inasingleAMLcasetracked for9years reveals the

complexityofleukemiaprogression.Leukemia30:295–302.doi:10.1038/leu.2015.264

Koren S, Reavie L, Couto JP, et al (2015) PIK3CA(H1047R) inducesmultipotency andmulti-lineagemammary

tumours.Nature525:114–118.doi:10.1038/nature14669

Kreso A, Dick JE (2014) Evolution of the cancer stem cell model. Cell Stem Cell 14:275–91 . doi: S1934-

5909(14)00057-5[pii]10.1016/j.stem.2014.02.006

LagasseE(2008)Cancerstemcellswithgeneticinstability:thebestvehiclewiththebestengineforcancer.Gene

Ther15:136–142.doi:3303068[pii]10.1038/sj.gt.3303068

LandauDA,CarterSL,StojanovP,etal(2013)Evolutionandimpactofsubclonalmutationsinchroniclymphocytic

leukemia.Cell152:714–26.doi:10.1016/j.cell.2013.01.019

LangF,WojcikB,RiegerMA(2015)StemCellHierarchyandClonalEvolutioninAcuteLymphoblasticLeukemia.

StemCellsInt2015:137164.doi:10.1155/2015/137164

LaplaneL(2015)ReprogrammingandStemness.PerspectBiolMed58:229–46.doi:10.1353/pbm.2015.0022

LaplaneL(2016)CancerStemCells:PhilosophyandTherapies.HarvardUniverityPress,Cambridge,MA

LaplaneL(2014)Identifyingtheoriesindevelopmentalbiology.Thecaseofthecancerstemcelltheory.In:Minelli

A,PradeuT(eds)Towardatheoryofdevelopment.OxfordUniversityPress,Oxford,pp246–259

LatilM,NassarD,BeckB,etal (2017)Cell-Type-SpecificChromatinStatesDifferentiallyPrimeSquamousCell

CarcinomaTumor-InitiatingCellsforEpithelialtoMesenchymalTransition.CellStemCell20:191–204.e5.

doi:10.1016/j.stem.2016.10.018

LeanC, PlutynskiA (2016) Theevolutionof failure: explaining cancer as anevolutionaryprocess. Biol Philos

31:39–57.doi:10.1007/s10539-015-9511-1

Lewontin RC (1970) The Units of Selection. Annu Rev Ecol Syst 1:1–18 . doi:

10.1146/annurev.es.01.110170.000245

LewontinRC(1978)Adaptation.SciAm239:212–8,220,222passim

LingS,HuZ,YangZ,etal(2015)Extremelyhighgeneticdiversityinasingletumorpointstoprevalenceofnon-

Darwiniancellevolution.ProcNatlAcadSci112:E6496-6505.doi:10.1073/pnas.1519556112


22

LipinskiKA,BarberLJ,DaviesMN,etal(2016)CancerEvolutionandtheLimitsofPredictabilityinPrecisionCancer

Medicine.Trendsincancer2:49–63.doi:10.1016/j.trecan.2015.11.003

Mack SC,WittH, Piro RM, et al (2014) Epigenomic alterations define lethal CIMP-positive ependymomas of

infancy.Nature506:445–50.doi:10.1038/nature13108

MarkowetzF(2016)Asaltationisttheoryofcancerevolution.NatGenet48:1102–3.doi:10.1038/ng.3687

MartincorenaI,RaineKM,GerstungM,etal(2017)UniversalPatternsofSelectioninCancerandSomaticTissues.

Cell171:1029–1041.e21

Martincorena I,RoshanA,GerstungM,etal (2015)Highburdenandpervasivepositive selectionof somatic

mutationsinnormalhumanskin.Science(80-)348:880–886.doi:10.1126/science.aaa6806

Medema JP (2017) Targeting the Colorectal Cancer Stem Cell. N Engl J Med 377:888–890 . doi:

10.1056/NEJMcibr1706541

MelchorL,BrioliA,WardellCP,etal(2014)Single-cellgeneticanalysisrevealsthecompositionofinitiatingclones

andphylogeneticpatternsofbranchingandparallelevolution inmyeloma.Leukemia28:1705–15 .doi:

10.1038/leu.2014.13

MerlevedeJ,DroinN,QinT,etal(2016)Mutationalleleburdenremainsunchangedinchronicmyelomonocytic

leukaemiarespondingtohypomethylatingagents.NatCommun7:10767.doi:10.1038/ncomms10767

MerloLM,PepperJW,ReidBJ,MaleyCC(2006)Cancerasanevolutionaryandecologicalprocess.NatRevCancer

6:924–935.doi:nrc2013[pii]10.1038/nrc2013

MeyerM,ReimandJ,LanX,etal(2015)Singlecell-derivedclonalanalysisofhumanglioblastomalinksfunctional

andgenomicheterogeneity.ProcNatlAcadSciUSA112:851–6.doi:10.1073/pnas.1320611111

Mikkelsen TS, Hanna J, Zhang X, et al (2008) Dissecting direct reprogramming through integrative genomic

analysis.Nature454:49–55.doi:10.1038/nature07056

MurugaesuN,WilsonGA,BirkbakNJ,etal(2015)Trackingthegenomicevolutionofesophagealadenocarcinoma

throughneoadjuvantchemotherapy.CancerDiscov5:821–31.doi:10.1158/2159-8290.CD-15-0412

NassarD,BlanpainC(2016)CancerStemCells:BasicConceptsandTherapeuticImplications.AnnuRevPathol

11:47–76.doi:10.1146/annurev-pathol-012615-044438

NavinN,KendallJ,TrogeJ,etal(2011)Tumourevolutioninferredbysingle-cellsequencing.Nature472:90–4.

doi:10.1038/nature09807

Nik-ZainalS,AlexandrovLB,WedgeDC,etal(2012a)Mutationalprocessesmoldingthegenomesof21breast

cancers.Cell149:979–93.doi:10.1016/j.cell.2012.04.024

Nik-ZainalS,VanLooP,WedgeDC,etal(2012b)Thelifehistoryof21breastcancers.Cell149:994–1007.doi:

10.1016/j.cell.2012.04.023

NottaF,MullighanCG,WangJC,etal(2011)EvolutionofhumanBCR-ABL1lymphoblasticleukaemia-initiating

cells.Nature469:362–367.doi:nature09733[pii]10.1038/nature09733

OdouxC,FohrerH,HoppoT,etal(2008)Astochasticmodelforcancerstemcellorigininmetastaticcoloncancer.

CancerRes68:6932–6941.doi:68/17/6932[pii]10.1158/0008-5472.CAN-07-5779

OrtmannCA,KentDG,NangaliaJ,etal(2015)Effectofmutationorderonmyeloproliferativeneoplasms.NEngl

JMed372:601–12.doi:10.1056/NEJMoa1412098


23

Pantic I (2011) Cancer stem cell hypotheses: impact on modern molecular physiology and pharmacology

research.JBiosci36:957–961

PepperJW,ScottFindlayC,KassenR,etal(2009)Cancerresearchmeetsevolutionarybiology.EvolAppl2:62–

70.doi:10.1111/j.1752-4571.2008.00063.x

PepperJW,SprouffskeK,MaleyCC(2007)Animalcelldifferentiationpatternssuppresssomaticevolution.PLoS

ComputBiol3:e250.doi:10.1371/journal.pcbi.0030250

Piccirillo SG, Combi R, Cajola L, et al (2009) Distinct pools of cancer stem-like cells coexist within human

glioblastomas and display different tumorigenicity and independent genomic evolution. Oncogene

28:1807–1811.doi:onc200927[pii]10.1038/onc.2009.27

PolyakK(2007)Breastcancer:originsandevolution.JClinInvest117:3155–3163.doi:10.1172/JCI33295

PotterNE,ErminiL,PapaemmanuilE,etal(2013)Single-cellmutationalprofilingandclonalphylogenyincancer.

GenomeRes23:2115–25.doi:10.1101/gr.159913.113

QuintanaE,ShackletonM,SabelMS,etal(2008)Efficienttumourformationbysinglehumanmelanomacells.

Nature456:593–598.doi:nature07567[pii]10.1038/nature07567

Rausch T, Jones DTW, Zapatka M, et al (2012) Genome sequencing of pediatric medulloblastoma links

catastrophicDNArearrangementswithTP53mutations.Cell148:59–71.doi:10.1016/j.cell.2011.12.013

ReyaT,MorrisonSJ,ClarkeMF,WeissmanIL(2001)Stemcells,cancer,andcancerstemcells.Nature414:105–

11.doi:10.1038/3510216735102167[pii]

ReynaudD,PietrasE,Barry-HolsonK,etal (2011) IL-6controls leukemicmultipotentprogenitorcell fateand

contributes to chronic myelogenous leukemia development. Cancer Cell 20:661–73 . doi:

10.1016/j.ccr.2011.10.012

Schuh A, Becq J, Humphray S, et al (2012)Monitoring chronic lymphocytic leukemia progression by whole

genome sequencing reveals heterogeneous clonal evolution patterns. Blood 120:4191–6 . doi:

10.1182/blood-2012-05-433540

ShackletonM,QuintanaE,FearonER,MorrisonSJ(2009)Heterogeneityincancer:cancerstemcellsversusclonal

evolution.Cell138:822–829.doi:S0092-8674(09)01030-7[pii]10.1016/j.cell.2009.08.017

ShainAH,YehI,KovalyshynI,etal(2015)TheGeneticEvolutionofMelanomafromPrecursorLesions.NEnglJ

Med373:1926–36.doi:10.1056/NEJMoa1502583

ShipitsinM,CampbellLL,ArganiP,etal(2007)Moleculardefinitionofbreasttumorheterogeneity.CancerCell

11:259–273.doi:S1535-6108(07)00029-3[pii]10.1016/j.ccr.2007.01.013

SiegmundKD,MarjoramP,TavaréS,ShibataD(2009)Manycolorectalcancersare“flat”clonalexpansions.Cell

Cycle8:2187–93

SimonsBD(2016)Deepsequencingasaprobeofnormalstemcellfateandpreneoplasiainhumanepidermis.

ProcNatlAcadSciUSA113:128–33.doi:10.1073/pnas.1516123113

SottorivaA,BarnesCP,GrahamTA(2017)Catchmydrift?Makingsenseofgenomicintra-tumourheterogeneity.

BiochimBiophysActa-RevCancer.doi:10.1016/j.bbcan.2016.12.003

SottorivaA,KangH,MaZ,etal(2015)ABigBangmodelofhumancolorectaltumorgrowth.NatGenet47:209–

216.doi:10.1038/ng.3214


24

SottorivaA,SpiteriI,PiccirilloSGM,etal(2013)Intratumorheterogeneityinhumanglioblastomareflectscancer

evolutionarydynamics.ProcNatlAcadSciUSA110:4009–14.doi:10.1073/pnas.1219747110

SottorivaA,VerhoeffJJ,BorovskiT,etal(2010)Cancerstemcelltumormodelrevealsinvasivemorphologyand

increased phenotypical heterogeneity. Cancer Res 70:46–56 . doi: 0008-5472.CAN-09-3663 [pii]

10.1158/0008-5472.CAN-09-3663

SprouffskeK,AthenaAktipisC,RadichJP,etal(2013)Anevolutionaryexplanationforthepresenceofcancer

nonstemcellsinneoplasms.EvolAppl6:92–101.doi:10.1111/eva.12030

StephensPJ,GreenmanCD,FuB,etal(2011)Massivegenomicrearrangementacquiredinasinglecatastrophic

event during cancer development. Cell 144:27–40 . doi: S0092-8674(10)01377-2

[pii]10.1016/j.cell.2010.11.055

SunJ,RamosA,ChapmanB,etal(2014)Clonaldynamicsofnativehaematopoiesis.Nature514:322–327.doi:

10.1038/nature13824

SuzukiM,HarashimaA,OkochiA,etal(2004)5-Azacytidinesupportsthelong-termrepopulatingactivityofcord

bloodCD34(+)cells.AmJHematol77:313–5.doi:10.1002/ajh.20178

TakebeN,IvySP(2010)Controversiesincancerstemcells:targetingembryonicsignalingpathways.ClinCancer

Res16:3106–3112.doi:1078-0432.CCR-09-2934[pii]10.1158/1078-0432.CCR-09-2934

ThirlwellC,WillOCC,DomingoE,etal(2010)Clonalityassessmentandclonalorderingofindividualneoplastic

crypts shows polyclonality of colorectal adenomas. Gastroenterology 138:1441–54, 1454–7 . doi:

10.1053/j.gastro.2010.01.033

Tomasetti C, Li L, Vogelstein B (2017) Stem cell divisions, somatic mutations, cancer etiology, and cancer

prevention.Science(80-)355:1330–1334.doi:10.1126/science.aaf9011

TomasettiC,VogelsteinB(2015)Variationincancerriskamongtissuescanbeexplainedbythenumberofstem

celldivisions.Science(80-)347:78–81.doi:10.1126/science.1260825

VanKeymeulenA,LeeMY,OussetM,etal(2015)ReactivationofmultipotencybyoncogenicPIK3CAinduces

breasttumourheterogeneity.Nature525:119–23.doi:10.1038/nature14665

WalterMJ,ShenD,DingL,etal(2012)Clonalarchitectureofsecondaryacutemyeloidleukemia.NEnglJMed

366:1090–8.doi:10.1056/NEJMoa1106968

WelnerRS,AmabileG,BarariaD,etal(2015)Treatmentofchronicmyelogenousleukemiabyblockingcytokine

alterations found in normal stem and progenitor cells. Cancer Cell 27:671–81 . doi:

10.1016/j.ccell.2015.04.004

WichaMS,LiuS,DontuG(2006)Cancerstemcells:anoldidea--aparadigmshift.CancerRes66:1883–1886.doi:

66/4/1883[pii]10.1158/0008-5472.CAN-05-3153

WilliamsMJ,WernerB,BarnesCP,etal(2016)Identificationofneutraltumorevolutionacrosscancertypes.Nat

Genet48:238–244.doi:10.1038/ng.3489

XieM,LuC,WangJ,etal (2014)Age-relatedmutationsassociatedwithclonalhematopoieticexpansionand

malignancies.NatMed20:1472–8.doi:10.1038/nm.3733

XuX,HouY,YinX,etal(2012)Single-cellexomesequencingrevealssingle-nucleotidemutationcharacteristics

ofakidneytumor.Cell148:886–95.doi:10.1016/j.cell.2012.02.025


25

Yachida S, Wood LD, Suzuki M, et al (2016) Genomic Sequencing Identifies ELF3 as a Driver of Ampullary

Carcinoma.CancerCell29:229–240.doi:10.1016/j.ccell.2015.12.012

ZhangJ,FujimotoJ,ZhangJ,etal(2014)Intratumorheterogeneityinlocalizedlungadenocarcinomasdelineated

bymultiregionsequencing.Science346:256–9.doi:10.1126/science.1256930

ZhangXC,XuC,MitchellRM,etal(2013)Tumorevolutionandintratumorheterogeneityofanoropharyngeal

squamouscellcarcinomarevealedbywhole-genomesequencing.Neoplasia15:1371–8

Cancer stem cells modulate patterns and processes of...

Documents

Transcript of Cancer stem cells modulate patterns and processes of...