Functions of the Ikaros transcription factor and the role of gene defects in hematological malignancies

(1)

Review/Praca poglądowa

Functions of the Ikaros transcription factor and the role of IKZF1 gene defects in hematological

malignancies

Funkcje czynnika transkrypcyjnego Ikaros oraz znaczenie defektów genu IKZF1 w nowotworach hematologicznych

Anna Gorzkiewicz, Anna Walczewska *

DepartmentofCell-to-CellCommunication,MedicalUniversityofLodz,Lodz,Poland

Introduction

Interest in Ikaros, the zinc ﬁnger-containing transcription factor, hasbeengrowing since theearly 1990s [1].Initially,

in 1991,astudy of erythroid globinswitching revealedthe existence of a DNA-binding factor with a strong binding activityrestrictedtohematopoietictissues.Thisproteinwas foundtobindtomorethan95%ofthepyrimidine(PYR)-rich domain on one strand of DNA upstream of the human article info

Articlehistory:

Received:30.06.2014 Accepted:16.10.2014 Availableonline:27.10.2014

Keywords:

Ikaros

IKZF1

Acutelymphoblasticleukemia

ALL

Słowakluczowe:

Ikaros

IKZF1

ostrabiałaczkalimfoblastyczna

OBL

abstract

Discoveredintheearly1990s,thehighlyconservedDNA-andprotein-bindingtranscrip- tionfactor,Ikaros,isnowconsideredoneofthemostimportantplayersinhematopoie- sisandthedevelopmentofcertainformsofhumanmalignancies.TheIkarostranscrip- tionfactorisamultifunctionalproteinregulatinghematopoieticstemcells(HSCs)function, coordinating self-renewal, cell survival processes, cell cycle progression and lymphopoiesis. Ikaros is also considered as one of the most important antileukemic transcriptionfactors.AlterationsintheIkarosgene(IKZF1)characterizeasubsetofacute lymphoblastic leukemia withsignificant resistanceto treatment and increased risk of relapse.Hematologicalstudieshighlightshortenedandmodifieddominantnegative(DN) Ik-forms,thatplayanimportantroleinthedevelopmentandprognosisofhematological malignances.Currently,extensiveresearchinthisfieldisapriorityinthebattleagainst leukemia. This paper describes the structural and functional properties of the Ikaros protein and its family members, importantinteractions with the nuclear proteins, its influenceongenetranscriptionalprofiles,aswellasitsconsiderableinvolvementinthe keyhematopoieticprocesses.

*Correspondingauthorat:ZakładInterakcjiMiędzykomórkowych,UniwersytetuMedycznegowŁodzi,ul.Mazowiecka6/8, 92-215Łódź,Polska.Tel.:+48422725654.

E-mailaddress:anna.walczewska@umed.lodz.pl(A.Walczewska).

ContentslistsavailableatScienceDirect

Acta Haematologica Polonica

journal homepage:www.elsevier.com/locate/achaem

http://dx.doi.org/10.1016/j.achaem.2014.10.001

(2)

d-globin gene [2]. As we know now, it was Ikaros protein associatedwiththePYRcomplex.Ikaros,designatedasIk-1, Ik-VIorLyf-1,isencodedbytheIKZF1gene(also knownas ZNFN1A1)locatedonthe shortarmofhuman chromosome 7 and on mouse chromosome 11, next to the epidermal growthfactorreceptorgene[3,4]. Ikarosproteinisahighly conserved antileukemic transcription factor with a 95%

homology between mice and humans at the amino acid level[3].

Ikarosfamilymemberscomprisetwofunctionallydiffer- ent groups. The members of the ﬁrst group are the basic form of the proteins, include Ikaros encoded by the IKZF1 gene,Helios encodedbythe IKZF2gene, Aiolosencoded by theIKZF3gene,EosencodedbytheIKZF4gene,andPegasus encoded by the IKZF5 gene. These proteins possess both DNA-binding and protein-binding domains. Thefunctional Ik-forms, Ikaros,Helios and Aiolos, togetherwith Ik-Hand Ik-X act in concert withthe Ikaros protein, partially com- plementing its function, while Eos can only bind to apalindromicDNAsequence[5].

ThesecondgroupoftheIkarosfamilymembersconsists of dominant-negative (DN) Ik-forms encoded by the same genesasfunctional forms.However,these formshave lost their ability to bind DNA due to the deletion of exon 5containingtwo N-terminalzinc ﬁngers(ZF),ZF2 and ZF3 asaresultoftheiralternativesplicing.Theseshort,patholo- gical, DNA-nonbinding DN forms inhibitthe functional Ik- formsof groupone[6–8]. Theexpressionof DNsinnormal bonemarrowcellsisataverylowlevel[9],buttheirhigher levels are associatedwithhematopoietic malignancies [10].

Normal infant bonemarrow cells and thymocytesproduce onlyIkarosandHelioslocalizedinthenucleus,whereasthe DNsdisplaycytoplasmiclocalization[11].

Ithasbeendemonstratedinmicethattheheterozygous deletionof exons4 and 5, which encodethree N-terminal ZFs,causeup-regulationof DNformssuchasIk-6,Ik-7and Ik-8,andleadtodevelopmentofahighlyaggressiveformof lymphoblastic leukemia. However, the same mutation in a homozygous mouse germline completely arrested the developmentofalllymphoidlineages[11].Itisalsointerest- ing to note that Ikaros deletions are found much more frequently in p53KO lymphomas than in wild-type p53 lymphomas:bothgenesbeingonthesamechromosome[4].

Ikaros and its family members play a crucial role in hematopoiesis, particularly in the lymphoid lineage as amaster regulator of early lymphocyte development, their differentiation, homeostasis and function [12–14]. Through the regulation of target gene expression, Ikaros proteins controlcellcycleprogressionandcellsurvival[6,15],aswell asisotypeselectionduringclass-switchDNArecombination of the immunoglobulin [16]. Moreover, this protein is a crucial functional regulator and self-renewal factor of hematopoieticstemcells(HSCs)[13,17]andacriticaltumor suppressor [13,18, 19]. Ithas been alsodemonstrated that duringchemotherapyorUVtreatment,theIkarosproteinis degradedintheearlyphaseofapoptosis,presumablybythe proteasomesystem,priortotheactivationofcaspase3[20].

Theexpression proﬁle of Ikaros family members varies dependingoncelltype[21],andthe co-expressionof these proteins enablestheformationof highly stablehomo-and

heterodimers, which presumably can associate into multi- mers[21, 22].Dimerization of DNA-binding Ikarosisoforms results in their enhanced afﬁnity for DNA and increased transcriptionactivation[22].

The expression of Ikaros is primarily detected in the thymus andspleen,where it isessentialfor theregulation of T-cell speciﬁc gene transcription. Furthermore, Ikarosis expressed in HSCs, myeloid and erythroid precursors, mature T and B cells, natural killer (NK) cells, activated Tcellsandnucleatederythroidlineagecells[9],aswellasin commonlymphoidprogenitors[17,23,24].Although,Ikaros isupregulatedduringTcellactivation,theproteinoccursat a very low level in resting T cells [9, 25, 26]. The highest expression of Ikaros is observed in immature thymocytes, aswellasinself-renewingfractionsoflongterm(LT)HSCs.

Interestingly, the daughter shortterm (ST) HSC population displayssigniﬁcantlylowerIkaroslevels[17].

Although Ikaros has been previously described as a lymphoid lineage specific factor, recent studies indicate that it plays a role inother cell types, suchas developing striatal mouse neurons [27]. It has been described as inducing both cell cycle arrest of neural progenitors and neurogenesis of late precursors, i.e. ENK-positive striatal neurons[28].IthasalsobeenreportedthatIkarosoccursat highlevelsinprogenitorcellsofthecerebralcortexinearly stages of neurogenesisin mice, after which its expression systematically decreases [29]. Moreover, Ikaros expression hasbeenobservedinallearlyembryonicmousemultipotent retinal progenitor cells which, at later stages, give rise to Ikaros deficient retinal progenitorcells. The studieson the role of Ikaros in neuronal development indicate that this protein is as a mammalian homolog of Hunchback, atemporalzincfingertranscriptionfactorselectingthefate ofthefirst-bornneuronsinDrosophila[29,30].

Ikaros structure and function

The crucial features of Ikaros that contributed to its dis- covery weretheC2H2zincfinger(ZF)motifslocatedintwo Krüppel-like zinc finger domains, which are characteristic for bothDNA-bindingandprotein-bindingproteins. FourZF motifs located centrally on the N-terminal domain of the Ikarosprotein(ZF1–ZF4)areknowntopossessDNA-binding affinity,whereas 2additional zincfingers(ZF5,ZF6)located ontheC-terminaldomain,namedthedimerizationdomain, are responsible for protein interaction [10, 31, 32] (Fig. 1).

Presumably, theC-terminal zinc ﬁngersare responsible for the pericentromeric targeting of the Ikaros protein within thenucleus[31].

The latest designations of Ikaros family members are differenttotheoriginalones.Asexon1isnottranslatedand hasnotbeeninitiallyidentiﬁed,theﬁrstreportsdescribeonly sevenexons[6,33].However,allIkarosisoformspossessexon 8withprotein-binding ZF5andZF6motifs.Moreover, many of theIKZF1familymembergenes missthelast30basesof exon7andaredesignatedasminusforms[6].

Ikaros interacts with DNA, inserting the DNA-binding domainintothemajorgrooveofDNA[32].Asatranscription factor,Ikarosbindsto1or2sitescontainingthe(C/T)GGGA

(3)

(A/T)sequenceinpromotersofregulatedgenes[4,10,11,34– 36]. The Ikaros protein possesses 1 activationdomain and 2 repression domains [7]. The ability to repress gene tran- scriptiondoesnotdependonDNA-bindingafﬁnityordimer- izationpropertiesbut onthe celltype and thesequence of gene promoters [37]. The Ikaros-induced repression is mediated by chromatin modiﬁcation, co-repressor recruit- ment,aswellascompetitionforDNAsequences[13].

Ikarosislocated primaryinpericentromeric heterochro- matin (PC-HC)inclose proximity totranscriptionally silent genes[38].Thisispresumablycausedbyitsinteractionwith histone deacetylase complexes (HDACs) present at hyper- methylated DNA fragments [39]. In fact, deacetylation of histoneslocatedininaccessibleregionsofDNArequiresthe action of chromatinremodelingcomplexes.However,since the functionof these complexes dependson the cell type, thepattern of generepressiondisplays high cellspeciﬁcity [37]. Throughout chromatin remodeling, Ikaros recruits

targetgenestothepericentromericheterochromatin,which resultsinactivationorrepression oftheirtranscription[10, 12, 18, 40–42]. Ikaros can repress genes in both HDAC- dependent and independent manner,for instance,through its interactions withthe co-repressor, theC-terminal bind- ingprotein[43].

ThedifferentiationofHSCsintoalymphoid-myeloidline- agerequiresrepressionoftranscriptionalprogramsaffiliated with the fate of megakaryo-erythroid cells. It has been reported thatIkaros modulateslymphoidand myeloidgene expression,butalsorepressesgenesresponsibleforerythroid commitmentandtheself-renewalability[44].Loss ofIkaros function inlymphoid-myeloid restrictedprogenitors(LMPPs) causes deficiency of tyrosine kinase receptor Flt3 [45] and promotesmyeloidlineagebyupregulationofspecificnuclear factorsandcytokinereceptors[46].LossofIkarosfunctionin commonmyeloidprogenitors(CMPs)promoteserythro-mega- karyocyte formation, however, intermediatelevels of Ikaros Fig.1–StructuresofIkarosfamilymemberproteins.Exon1isuntranslated.Someformslacktranslationofthelast30bases ofexon7(thepartmarkedwith‘‘X’’)andaredesignatedas‘‘minus’’forms.(A)StructureofIkaros1protein.(B)Structuresof majorfunctionalIk-forms:Ik-HandIk-X.(C)StructuresofmajorDNIk-forms:Ik-6,Ik-9andIk-10.ZF:zincfinger;N-:N- terminalend;C-:C-terminalend

Ryc.1–StrukturybiałekzrodzinyIkaros.Ekson1niejesttłumaczony.Wniektórychformachostatnich30parzasadeksonu7nie jesttłumaczone(fragmentprzekreślony,,X’’),aichnazwysąoznaczaneminusem.(A)StrukturabiałkaIkaros1.(B)Strukturydwóch głównychbiałekfunkcjonalnychzrodzinyIkaros:Ik-HiIk-X.(C)Strukturytrzechgłównychformbiałekdominującychnegatywnych zrodzinyIkaros:Ik-6,Ik-9iIk-10.ZF:paleccynkowy;N-:koniecaminowy;C-:konieckarboksylowy

(4)

were foundto generateerythroid andmyeloid lineagecells [45]. Ikaros is also a crucial regulator of the self-renewal machinery active in the pool of long term-reconstituting HSCs. A homozygous mutation of Ikaros in mouse fetus resultsinlossofthe(CD150+)longterm-HSCs,leavingnon- long term self-renewing cellular pools, which causes rapid andfatalanemia,aswellasadysregulationoferythroidcell differentiation and proliferation from reduced progenitors [17].

Moreover, Ikaros playsa crucialroleinthe regulationof cellcycleandcelldifferentiation.Inthenucleus,speciﬁcally inthehigher-orderchromatinstructure,itiscolocalizedwith methyltransferaseandCyclinA,suggestingthatitisaccessi- bletobothtranscriptionandreplicationmachineries[25].Cell division, especiallyduring replication or themoment after- wards, provides an opportunity for gene maintenance to occur. This phenomenon may genetically distinguish the daughter from the mother cell, which is the principle of differentiation[25]. Ikarosisknowntohavethepotentialto maintainheterochromaticcodethroughcelldivisions,ashas beendescribedinastudyontheroleofchromatinremodel- inginhematopoieticlineagecommitmentanddifferentiation:

Ikaros appears to coordinate the correct differentiation of daughter cells from the mother cell, and act as a factor governingthepatternoffate-speciﬁcgeneexpression[47].

Ikarosisalsoinvolved inthe controlof Tcellactivation in which Ikaros co-localizes with the DNA replication machinery. Ithas been reportedthat reducedor abolished IkarosactivityleadstoacceleratedentryintotheSphaseby reductionof theactivation threshold,reduction ofsensitiv- itytocellcycleinhibitorsandhyper-responsetoIL-2,which is requiredfor G1-S progression [25]. Thisleads toaberra- tions in chromosome structures including an oversized chromosome 1. The absence of Ikaros in mice results in rapiddevelopmentofT-cellneoplasia[25].

Another important function of Ikaros is shaping the courseofanimmuneresponsewhichrequiresthepresence ofIL-10fornormaldirection.Ikarosisknowntobeacritical activatorofIL-10geneexpressioninCD4+Tcells[48].Ikaros null T cells display aberrant differentiation and low IL-10 gene expression. Furthermore, recent reports have shown thatIkarosisinvolvedintherepressionof theIL-2genein anergicTcells,aswellasintheactivationof IL-4,IL-5,and IL-13 genes inTh2 cells suggesting that Ikarosis a critical factorforTcelleffectorfunctions[49].

Ikarosisphosphorylatedpredominantlybycaseinkinase II(CKII).Asa consequenceofthis protein phosphorylation, therecruitmentofIkarostoPC-HC,itsDNA-bindingafﬁnity, therepressionofsomegenes,andthenegativeregulationof G1-Stransitionisreducedorabolished[18,50–52].Although Ikaros is dephosphorylated by the late G1 phase, Ikaros becomeshyperphosphorylatedasitenterstheSphaseand remains so throughout the M phase [50]. Inappropriate levels of activeor inactive proteins regulating G1-S transition,acriticalcellcyclecheckpoint,inducecellapoptosisor neoplasticstateinprimarylymphocytes[50].

Moreover, Ikaros has been established as a centralreg- ulator of the transition of pre-B or pre-T cells to mature lymphocytes[14].LackofIkaros,inbothBandTcells,causes adeﬁcitinantigen-receptorrearrangement[53].Thisprotein

alsoregulatesthetranscriptionofgenes,suchasCd4orCd8, andhasbeendemonstratedtoactivatetheCD8agenelocus [41, 54]. Lack of Ikaros activity during the progression of double negative T cells (CD4CD8) to double positive (CD4⁺CD8⁺)thymocytesresultsininappropriatepre-TCRand TCR signaling [14, 54], nonetheless the T cells progression occurs. However, the rateof differentiation into the double positivestage isincreasedandthenexttransitionisheavily skewed toward CD4 single-positive T cells, which do not appearintheperiphery, presumablybecausetheyhave not undergonetheﬁnalselectionstepsrequiredfortheirexporta- tion.Moreover,thenormalproliferativeexpansionofTcells doesnotoccur,resultinginahighlyhypocellularthymus[14].

Ikaros is also required in mature B cells to ensure the speciﬁcity and efﬁciency of the class-switchrecombination.

In thisprocess,Ikaros isaregulator ofimmunoglobulin(Ig) isotype selection. It has been shown thatIkaros deﬁciency promotes ectopic class-switch recombination toward IgG2b and IgG2a and reduces class-switch recombination into all othertypes[16].

There are several lines of evidences that Ikaros also promotesthedifferentiationandsurvivalofhumanerythroid cells.Furthermore,itiscrucialforerythroidmaturationand, mostimportantly,foradulterythroidglobingeneexpression and globin switch [2, 55, 56]. Ikaros is required for the formation andDNA bindingof developmental-stagespeciﬁc complexessuchaschromatinremodelingcomplexesandthe polypyrimidine (PYR) complex, which occur only in adult hematopoietic cells[2,57–59].Lackof Ikarosinmiceresults inno PYRcomplex formationand delayedglobinswitching [57,58,60].

Ikaros alternations in hematological malignancies

The body of evidence indicating the role of Ikaros in leukemias has accumulated steadily since the mid-1990s [61].ThefocalorbroaddeletionsofIKZF1gene[62]arenow known to be the cause or the result of some human hematologicaldiseases, suchasacutelymphoblastic leukemia (ALL) [63]. Moreover, Ikaros genealterations indirectly leadtomyeloproliferativeneoplasms(MPNs)orits progression to acute myeloid leukemia (AML) [6]. Diverse IKZF1 mutations are found during blastic progression of chronic myeloid leukemia (CML) [64]. Presumably,activation of the JAK-STAT pathway is responsible for the leukemogenic potentialofIKZF1genemutations[65,66].

Homozygous IKZF1 gene deletions are embryonic lethal and areassociated with thedefective orabolisheddevelop- mentofalllymphoidcells,theearliestlymphoidprogenitors, aswell asexcessivemacrophageformationandfully defective erythrocyte and granulocyte differentiation. Heterozy- gous alterations of the IKZF1 gene mostly lead to rapid developmentofaggressiveleukemiasandlymphomas[7,61, 67]. Substitution of an amino acid in the DNA-binding ZF domaincausedbyapointmutationinonealleleoftheIkaros geneleadstocongenitalpancytopenia.Suchmutationsresult inprofoundBlymphopeniaanddecimatedNKcells,however, theTcellnumberremainsnormal,whichsuggeststhatother IkarosfamilymembersmaycompensateforthelossofIkaros

(5)

itself[68].Inaddition,Ikaros-dependentchangesoflympho- poiesiscanalsobecausedbyitsabolishedexpression.Ithas beenrevealedthathumanpituitaryglandcellslackingIkaros mRNA transcripts exhibit signiﬁcant IKZF1 promoter DNA hypermethylation[69].

Acute lymphoblastic leukemia

Totalor partialdeletionsof IKZF1 arevery frequentinALL.

Mutations of this gene were found in 20–30% cases of childhoodB-cellprecursorALL(BCP-ALL)andinabout50%of adult BCP-ALL and in over 80% of BCR-ABL1⁺ lymphoid leukemia(includingdenovoALLandCMLtransformationto ALL – lymphoid blast crisis), in 58% of BCR-ABL1-negative BCP-ALLcasesandapproximatelyin5%ofT-cellALL[18,62, 64,70–72]. However,Ikarosdeletionsarenotdetectedinthe acceleratedorchronicphaseofCML[64].Thefrequenciesof thevarioustypesofIKZF1mutationsinBCP-ALLwasreported asfollows: large deletions of chromosome 7are presentin 15% of cases, deletions of exons 2–7 in 15%, deletions of exons 4–7 in 30%, other deletions, including single exon deletions, appear in 30% of cases, and less frequent point mutationsarepresentin10%ofcases[70].Inrecentstudies, allBCP-ALLcaseswithIKZF1mutationsweresubdividedinto three functional categories characterized by:(1) monoallelic deletionsleadingtohaploinsufﬁciency(55%of allBCP-ALL cases), (2) 15–200kb deletions restricted to the IKZF1 gene leadingtoproductionofDNforms(D4–7accountfor33%of allIKZF1mutations),and(3)anIkarosnullphenotypecaused bybiallelicdeletions(12%ofallBCP-ALLcases)[35].

AhighriskofchildhoodALLdevelopmentduetoasingle nucleotide polymorphism (SNP) in IKZF1 (7p12.2 variant rs4132601)hasbeenreported[73,74].Interestingly, another SNP, rs10272724 (T>C), near IKZF1 (at 7p12.2) has been identiﬁed and is associated with a reduced risk of type 1diabetes[75].Therefore,itishypothesizedthattheseSNPs affectthemRNAlevelinterruptinglymphocytedevelopment and leadingtoreducedimpaired immuneresponse,result- ingindiabetes-protectiveandleukemia-susceptibleeffects.

A strong correlation has been shown between IKZF1 deletions and a poor outcome inhuman leukemias arising fromthedisruptionofearlyB-celldevelopment.Forinstance, Ikaros restricts the expression of hematopoietic stem cell- speciﬁc genes and negatively regulatescellproliferation. As aconsequence,Ikarosappearstobeacentralregulatorofthe high-riskgene-expressionsignature[14,61,64,71].

The susceptibilityof leukemias totreatmentdepends on the nature of themutated transcriptional regulator. Apoor prognosisassociatedwiththehigh-riskBCP-ALLresultsfrom twomajorgeneticmodiﬁcations.The ﬁrstbeing overexpression of hematopoieticstem cellgenes,giving risetomalig- nantclonesthatexhibitsuch stemcell-likefeaturesasself- renewal and drug resistance. The second being the down- regulationofgenesresponsibleforlymphocytedevelopment, whichcontributestoB-celldevelopmentalarrestatthepoint of high proliferative potential and, hence, to increased susceptibilitytomalignanttransformation[64,71].

The presence of BCR-ABL1 rearrangement is associated withahighriskofleukemiarelapse.Thistypeofmutationis

foundinonly5%ofpediatricBCP-ALLandinapproximately 40%ofadultALL.Inthegroup of43patientswithBCR-ABL1 ALL the IKZF1 gene deletions were detected in 76.2% of childrenand 90.9% ofadults[64]. Lacobucciet al.examined a group of 106 de novo BCR-ABL1⁺ adult ALL patients for oncogenic lesions that cooperate with BCR-ABL1 to induce ALL. The most frequent,identified in 75% of patients, was afocaldeletionon7p12ofIKZF1gene.Furthermore,patients wereexaminedfortwomajorbroadIKZF1deletions:exons4– 7 and exons 2–7. Among patients with any IKZF1 deletion identified, thelossof exons4–7wasfound in55%,whereas 24% of patients suffered from a lack of exons 2–7 [76]. In anotherstudyfrom2012,agroupof144adultB-ALLpatients was analyzed for total or partial IKZF1 deletions. In this cohort, 106 patients were BCR-ABL1⁺ with the frequency of IKZF1deletionshigherthanintherestof38patientsnegative for known molecularrearrangements (B-NEGpatients). The frequenciesofIKZF1deletionsreached75%and58%,respec- tively [34]. In BCR-ABL1⁺ ALL, 59–70% of all examinedIKZF1 mutations weresevere D4–7 deletionsor biallelicmutations [64, 70, 76]. In contrast, 57% of the IKZF1 mutations inthe high-risk BCP-ALL pediatric cases negative for BCR-ABL1 demonstrated IKZF1 haploinsufficiency [77]. Loss of Ikaros functionduetoitsgenemutationcausesB-lymphoidmatura- tion arrest in BCR-ABL1⁺ ALL by alteration of the earliest lymphoidlineagespecification,especiallytheB-celldevelopment.Moreover,frequentco-deletionsof PAX5andCDKN2A were found together with IKZF1 deletionsinBCR-ABL1⁺ ALL [64, 71]. AlthoughBCR-ABL1translocation wasconsidered to appearindependentlyof Ikarosmutations,theIkarosaltera- tions directly contribute to the pathogenesis of BCR-ABL1⁺ ALL resulting in an increased relapse rate and a greater numberofadverseevents[64,71,77–79].Moreover,theIKZF1 gene alterations occur in 40% of ALL with a BCR-ABL1-like phenotype andthissubset isassociated witha highrisk of relapse [72]. Another study of BCR-ABL1-like ALL subtype showed ahigh rateof deletions ingenes involvedinB-cell development(includingIKZF1,TCF3,EBF1,PAX5,andVPREB1) occurringin82%ofexaminedpatients[80].Somaticdeletions andmutationsoftheIKZF1genehavealsobeenidentifiedin Philadelphia-like ALL,whichisan ALLsubtypewitha gene expressionprofilesimilartoPhiladelphia-positiveALL.Itwas recently found that in 73% of Philadelphia-like ALL cases, aGATA3variant(rs3824662)ispresenttogetherwithaCRLF2 lesion,JAKmutationandIKZF1deletion[72,81].

It has been previously reported that broad exon deletions resulting in DNs expression have been found to originate from gene alterations and non-aberrant post- transcriptionalsplicinginducedbyBCR-ABL1.MajorDNIk- forms of aberrant IKZF1 expression are enumerated in Fig.1.Ithasbeensuggestedthatdeletionofexons4–7from IKZF1 arisesdue toaberrant RAG-mediatedrecombination [64, 76]. This DN Ik-6 form has been identiﬁed in nearly a thirdofhumanT-cellacute leukemiacases[69].Among others, the Ik-6DN formis the most commonproduct of IKZF1 mutations (Table I). Overexpression of Ik-6 favors myelopoiesis bysimultaneouslyalteringthe humanglobin switch and inhibiting erythroid differentiation [55]. In AML Ik-6 up-regulation enhances Bcl-XL activation in myeloid precursor cells [15]. A highIk-6 levelalso causes

(6)

up-regulationoftheantiapoptoticsignalin40%ofpituitary tumors by Bcl-XL promoter selective acetylation,whereas Ik-1attenuatesBcl-XLpromoteractivity[82].

In2012,ananalysisof422genesinbothB-NEGandBCR- ABL1⁺ adultB-ALL patientsrevealed thatthecoexistence of IKZF1deletionscontributedtoup-regulationof294genesand down-regulation of 128 genes. The group of up-regulated genes included genes responsible for cell-cycle progression, JAK-STATsignaling,stemcellself-renewal,apoptosisregula- tion, as well as the production of early primed erythroid factor KLF9 and the late myeloid gene ID2 [34]. A recent genome-wide analysis revealed that 50% of all genes up- regulatedduringB-celllineagespeciﬁcationinvivoareIkaros targets [83]. The group of down-regulated genes included genes responsible for DNA repair processes (MSH2, MSH6, UBE2V2), the B-cell differentiation pathway (EBF1, BLK, BTK, IGLL1,CD22,PLCG2,MAP3K1,VPREB1),apoptosisandcellcycle progression, as well as RAG gene. Accumulation of DNA damage was conﬁrmed by markedly increased basal phos- phorylationofhistoneH2A.X[34].

Chronic myeloid leukemia

The expression of tyrosine kinase that is constitutively activated due to BCR-ABL1 translocation underlies chronic myeloidleukemia(CML).ThisdefectcanaloneinduceaCML- like myeloproliferative disease, however, the generation of

blastic leukemia requires additional oncogenic lesions.

AlthoughCMLrespondswelltotherapyinthechronicphase, it acquiresresistancetotreatmentafterprogressingtoblast crisis. Cooperating cytogenetic aberrations and tumor suppressor gene mutations occur in CML progressing to blast crisis.ThemostfrequenteventsinthetransformationofCML to lymphoid blast crisis are IKZF1 diverse mutations (66%), such asdeletionsofexons4–7,2–7,oreventheentiregene [64,76].AlsotheIKZF1nonsensemutationhasbeenidentiﬁed inthe C-terminal zinc-ﬁnger domainof exon 7inthenon- deletedallele,togetherwithCDKN2AdeletionandPAX5copy numberalterations[64].

It is important to note that IKZF1 deletions are present only in lymphoblast crisis and have not been detected in myeloidblastcrisisorthechronicphaseofCML.Ananalysis of42newlydiagnosedCMLIk-6positivepatientsinlympho- blast crisis showed complete remission in only 40%, with a66.7%rateofrelapseafter2–18months.NoexpressionofIk- 6wasfoundinhealthycontrols,norinchronicoraccelerated phaseofCML,norinpatientswithmyeloblastcrisis[76,84].

Myeloproliferative neoplasm and acute myeloid leukemia

The most serious consequence of myeloproliferative neoplasm (MPN)is its transformationto AML.TheJAK2-V617F genegain-of-functionmutation occursinmorethanhalfof TableI–ReviewofthemostcommonDNproductsofIKZFfamilygenesmutationsreportedoverlast14years.Initially, researcheswerebasedonRT-PCRanalysisandafter2007large-scalecomparativegenomichybridization(CGH)technology usinghigh-densitymicroarraysemerged

TabelaI–PrzeglądnajczęściejwystępującychformbiałekdominującychnegatywnychpowstałychwwynikumutacjigenuIKZF1, opisanychwprzeciąguostatnich15lat.PoczątkowebadaniaopierałysięnatechniceRT-PCR,apo2007rokustosowanabyła technologiaporównawczejhybrydyzacjigenomowejzużyciemmikromacierzyowysokiejgęstości

Publication Year Typeofdisorder Detected

DNIk-forms

Subgroupwith detected spliceforms(%)

Researched group (detected/all)

Sunetal.[11] 1999 ALL Ik-4,Ik-7,Ik-8 100 7/7infants

Sun[63] 1999 T-ALL Ik-4orIk-4D 80 8/10infants

Ik-7orIk-7D 0 0/10

Ik-8D 60 6/10

BCP-ALL Ik-4orIk-4D 45 5/11infants

Ik-7orIk-7D 18 2/11

Ik-8D 27 3/11

Iacobucci etal.[85]

2000 BCR-ABL1⁺ALL Ik-6 49 23/47adults

Yagietal.[15] 2002 AML Ik-6 29.2 7/24pediatrics

Ezzatetal.[8] 2003 Pituitaryadenomas Ik-6 36 18/50

Ikcytoplasmic localization

56 28/50

Mullighan[64] 2008 BCR-ABL1⁺ALL Ik-6,Ik-9,Ik-10 76.2 16/21pediatrics

90.9 20/22adults

Iacobucci etal.[86]

2008 BCR-ABL1⁺ALL Ik-6 41 19/46adults

Martinelli[79] 2009 BCR-ABL1⁺ALL Ik-6 37 31/83(allpatients)

59 31/52(IKZF1deleted

patients)

Ik-10 20 17/83(allpatients)

33 17/52(IKZF1deleted

patients)

Mietal.[87] 2012 BCP-ALL Ik-6 14.7 56/379pediatrics

31.3 64/203adults

(7)

allMPN patients,however, nosigniﬁcant associationexists with post-MPN leukemic transformation, as both JAK2- V617F-dependentand JAK2-V617F-independentpathwaysof transformation have been observed. Moreover, a strong association was observed between del7p and leukemic transformation, where the common deleted region con- tainedonlytheIKZF1gene.Sixof sevenexaminedpatients with del7p progressed to AML whereas only one showed features of advanced phase of primary myeloﬁbrosis with pancytopenia and increased blasts inthe peripheral blood.

Deletionsontheshortarmofchromosome7(del7p)involve such clinical symptoms as anemia, thrombocytopenia and increased percentage of blasts in the peripheral blood.

Ikarosgene alterationsdo notinduce differentiation arrest of myeloid cell types.IKZF1deletions, but notpoint mutations,are acquired late inthe disease progression and are strongly associated with JAK2-V617F mutation. IKZF1 hap- loinsufﬁciency (including monosomy7) occurred in21% of post-MPNleukemiaand0.2%ofnonleukemicMPNpatients.

The IKZF1 gene haploinsufﬁciency was found to induce elevated Stat5 phosphorylation and increase cytokine- dependent growth. Monosomy 7 is known to occur in approximately10%ofadult and5%ofchildhoodAMLcases [65].Anotherstudyof childhoodAML revealedthat 7of10 casesofmyelomonocyticormonocyticleukemiasexpressed Ik6.ItislikelythatIk-6playsanexclusiveroleinmonocyte/

macrophagedifferentiation,asitup-regulatestheantiapop- toticproteinBcl-XL,whichisinvolvedinthedifferentiation andsurvivalofmonocytes/macrophages[15].

Summary

Ikaros is a multifunctional zinc finger DNA-binding tran- scriptionfactorencodedbytheIKZF1genewhichisrequired fortheearliestspecificationofthelymphoidlineageduring hematopoiesis. The Ikaros protein and the transcripts of fourother genescomprisethe basicformsof Ikarosfamily proteins. In addition, numerous variations of the Ikaros family genes transcripts may occur including defective formsofproteinsbeinginvolvedinarangeofhematological disorders such as leukemias. TheIKZF1 gene is subject to suchaberrationsaspointmutations andbiallelic deletions, the most common haploinsufficiency, as well as broad deletionsofexons2–7,and4–7.Thelastofthesemutations causes production of dominant negative Ik-forms, which lackthecoreoftheDNA-bindingdomain.Overexpressionof these dominant negative Ik-forms is detected in about ahalfof adultBCP-ALLand inmostof BCR-ABL1⁺ALL,BCR- ABL1-likeALLand BCR-ABL1-negativeBCP-ALLcases.Domi- nant negative Ik-forms not only inhibit the functional Ik- forms,butalsopreventthe basicIkarosforms fromoccup- ing a close proximity to PC-HC and their target genes.

However,IKZF1mutationscanariseassecondarymutations during disease progression, as in CML transformation to BCR-ABL1⁺ ALL, or MPN transformation to AML. Dominant negative Ik-forms occur in CML blast crisis and remain after transformationtoBCR-ABL1⁺ALL. Ikarosdeletionsare also acquired as a late event in the clonal evolution of myeloid progenitors, and the leukemogenic potential of

these mutations is presumably induced via JAK-STAT sig- nalingpathway.Moreover, JAKmutations,particularly JAK2 mutations, are known to be signiﬁcantly associated with IKZF1defectsinchildhoodALL.The7p12.2variationofIKZF1 increasestheriskofchildhoodALL,whereasthe BCR-ABL1- like signature and/or IKZF1 deletions are associated with high risk of relapse among BCP-ALL patients. The broad research of the Ikaros transcription factor identiﬁed this protein as a central regulator of numerous hematopoietic processesand determinedIkarosasapotential therapeutic targetofaggressiveleukemiatreatment.

Authors' contributions/Wkład autorów

Study design was done by AG and AW.AG contributedin data collection and interpretation, manuscript preparation, andliteraturesearch.

Conﬂict of interest/Konﬂikt interesu

Nonedeclared.

Financial support/Finansowanie

There hasbeennoﬁnancialsupportfor theconductof the researchand/orpreparationofthearticle.

Ethics/Etyka

The work described in this article hasbeen carried out in accordance withThe Code of Ethics of the World Medical Association (Declarationof Helsinki)for experiments invol- ving humans; EU Directive 2010/63/EU for animal experiments;UniformRequirementsformanuscriptssubmittedto Biomedicaljournals.

references/pi smiennictwo

[1] GeorgopoulosK,MooreDD,DerﬂerB.Ikaros,anearly lymphoid-speciﬁctranscriptionfactorandaputative mediatorforTcellcommitment.Science1992;258 (5083):808–812.

[2] O'neillD,BornschlegelK,FlammM,CastleM,BankA.

ADNA-bindingfactorinadulthematopoieticcellsinteracts withapyrimidine-richdomainupstreamfromthehuman d-globingene.ProcNatlAcadSciUSA1991;88(20):

8953–8957.

[3] MolnárA,WuP,LargespadaDA,etal.TheIkarosgene encodesafamilyoflymphocyte-restrictedzincﬁngerDNA bindingproteins,highlyconservedinhumanandmouse.

JImmunol1996;156(2):585–592.

[4] RebolloA,SchmittC.Ikaros,AiolosandHelios:

transcriptionregulatorsandlymphoidmalignancies.

ImmunolCellBiol2003;81(3):171–175.

[5] MolnarA,GeorgopoulosK.TheIkarosgeneencodesa familyoffunctionallydiversezincﬁngerDNA-binding proteins.MolCellBiol1994;14(12):8292–8303.

(8)

[6] FrancisOL,PayneJL,SuR,PayneKJ.Regulatorofmyeloid differentiationandfunction:thesecretlifeofIkaros.World JBiolChem2011;2(6):19–125.

[7] SunL,LiuA,GeorgopoulosK.Zincﬁnger-mediated proteininteractionsmodulateIkarosactivity,amolecular controloflymphocytedevelopment.EMBOJ1996;15 (19):5358–5369.

[8] EzzatS,YuS,AsaSL.Ikarosisoformsinhumanpituitary tumors–distinctlocalization,histoneacetylation,and activationofthe5⁰ﬁbroblastgrowthfactorreceptor-4 promoter.AmJPathol2003;163(3):1177–1184.

[9] PayneKJ,HuangG,SahakianE,etal.Ikarosisoformxis selectivelyexpressedinmyeloiddifferentiation.JImmunol 2003;170(6):3091–3098.

[10] LiZ,Perez-CasellasLA,SavicA,SongC,DovatS.Ikaros isoforms:thesagacontinues.WorldJBiolChem2011;2 (6):40–145.

[11] SunL,HeeremaN,CrottyL,etal.Expressionofdominant- negativeandmutantisoformsoftheantileukemic transcriptionfactorIkarosininfantacutelymphoblastic leukemia.ProcNatlAcadSciUSA1999;96(2):680–685.

[12] PayneKJ,DovatS.Ikarosandtumorsuppressioninacute lymphoblasticleukemia.CritRevOncog2011;16(1–2):3–12.

[13] SellarsMcL,KastnerP,ChanS.IkarosinBcelldevelopment andfunction.WorldJBiolChem2011;2(6):32–139.

[14] WinandyS,WuL,WangJ,GeorgopoulosK.Pre-Tcell receptor(Tcr)andTcr-controlledcheckpointsinTcell differentiationaresetbyIkaros.TheRockefellerUniversity Press.JExpMed1999;190(8):1039–1048.

[15] YagiT,HibiS,TakanashiM,etal.HighfrequencyofIkaros isoform6expressioninacutemyelomonocyticand monocyticleukemias:implicationsforup-regulationofthe antiapoptoticproteinBcl-XLinleukemogenesis.Blood 2002;99(4):1350–1355.

[16] SellarsMcL,Reina-San-MartinB,KastnerP,ChanS.

Ikaroscontrolsisotypeselectionduringimmunoglobulin classswitchrecombination.TheRockefellerUniversity Press.JExpMed2009;206(5):1073–1087.

[17] PapathanasiouP,AttemaJL,KarsunkyH,etal.Self-renewal ofthelong-termreconstitutingsubsetofhematopoietic stemcellsisregulatedbyIkaros.StemCells2009;27 (12):3082–3092.

[18] SongC,LiZ,ErbeAK,etal.RegulationofIkarosfunctionby caseinkinase2andproteinphosphatase1.WorldJBiol Chem2011;2(6):26–131.

[19] KathreinKL,LorenzR,InnesAM,GrifﬁthsE, WinandyS.IkarosinducesquiescenceandT-cell

differentiationinaleukemiacellline.MolCellBiol2005;25 (5):1645–1654.

[20] HeLC,XuHZ,GuZM,etal.Ikarosisdegradedby proteasome-dependentmechanismintheearlyphaseof apoptosisinduction.BiochemBiophysResCommun 2011;406(3):430–434.

[21] HahmK,CobbBS,McCartyAS,etal.Helios,aTcell- restrictedIkarosfamilymemberthatquantitatively associateswithIkarosatcentromericheterochromatin.

GenesDev1998;12(6):782–796.

[22] KelleyCM,IkedaT,KoipallyJ,etal.Helios,anovel dimerizationpartnerofIkarosexpressedinthe earliesthematopoieticprogenitors.CurrBiol1998;8(9):

508–515.

[23] MorganB,SunL,AvitahlN,etal.Aiolos,alymphoid restrictedtranscriptionfactorthatinteractswithIkarosto regulatelymphocytedifferentiation.EMBOJ1997;16 (8):2004–2013.

[24] WangJ,AvitahlN,CariappaA,etal.AiolosregulatesBcell activationandmaturationtoeffectorstate.Immunity 1998;9(4):543–553.

[25] AvitahlN,WinandyS,FriedrichC,etal.Ikarossets thresholdsforTcellactivationandregulateschromosome propagation.Immunity1999;10(3):333–343.

[26] RonniT,PayneKJ,HoS,etal.HumanIkarosfunctionin activatedTcellsisregulatedbycoordinatedexpressionof itslargestisoforms.JBiolChem2007;282(4):2538–2547.

[27] AgostonDV,SzemesM,DobiA,etal.Ikarosisexpressedin developingstriatalneuronsandinvolvedin

enkephalinergicdifferentiation.JNeurochem2007;102 (6):1805–1816.

[28] Martín-IbáñezR,CrespoE,UrbánN,etal.Ikaros-1couples cellcyclearrestoflatestriatalprecursorswith

neurogenesisofenkephalinergicneurons.JCompNeurol 2010;518(3):329–351.

[29] AlsiöaJM,TarchinibB,CayouettebM,LiveseyaFJ.Ikaros promotesearly-bornneuronalfatesinthecerebralcortex.

PNAS2013;110(8):E716–E725.

[30] ElliottJ,JolicoeurC,RamamurthyV,etal.Ikarosconfers earlytemporalcompetencetomouseretinalprogenitor cells.Neuron2008;60(1):26–39.

[31] CobbBS,Morales-AlcelayS,KleigerG,etal.Targetingof IkarostopericentromericheterochromatinbydirectDNA binding.GenesDev2000;14(17):2146–2160.

[32] PayneMA.Zincﬁngerstructure-functioninIkaros.WorldJ BiolChem2011;2(6):61–166.

[33] HahmK,ErnstP,LoK,etal.Thelymphoidtranscription factorLyF-1isencodedbyspeciﬁc,alternativelyspliced mRNAsderivedfromtheIkarosgene.MolCellBiol1994;14 (11):7111–7123.

[34] IacobucciI,IraciN,MessinaM,etal.IKAROSdeletions dictateauniquegeneexpressionsignatureinpatientswith adultB-cellacutelymphoblasticleukemia.PLoSONE2012;7 (7):e40934.

[35] KastnerP,DupuisA,GaubMP,etal.FunctionofIkarosasa tumorsuppressorinBcellacutelymphoblasticleukemia.

AmJBloodRes2013;3(1):1–13.

[36] YapW,YeohE,TayA,etal.STAT4isatargetofthe hematopoieticzinc-ﬁngertranscriptionfactorIkarosinT cells.FEBSLett2005;579(20):4470–4478.

[37] KoipallyJ,RenoldA,KimJ,etal.RepressionbyIkarosand Aiolosismediatedthroughhistonedeacetylasecomplexes.

EMBOJ1999;18(11):3090–3100.

[38] BrownKE,GuestSS,SmaleST,etal.Associationof transcriptionallysilentgeneswithIkaroscomplexesat centromericheterochromatin.Cell1997;91(6):845–854.

[39] NanX,NgHH,JohnsonCA,etal.Transcriptionalrepression bythemethyl-CpG-bindingproteinMeCP2involvesa histonedeacetylasecomplex.Nature1998;393(6683):

386–389.

[40] KoipallyJ,HellerEJ,SeavittJR,etal.Unconventional potentiationofgeneexpressionbyIkaros.JBiolChem 2002;277(15):13007–13015.

[41] HarkerN,NaitoT,CortesM,etal.TheCD8Genelocusis regulatedbytheIkarosfamilyofproteins.MolCell2002;10 (6):1403–1415.

[42] SabbattiniP,LundgrenM,GeorgiouA,etal.Bindingof Ikarostothelambda5promotersilencestranscription throughamechanismthatdoesnotrequire

heterochromatinformation.EMBOJ2001;20(11):

2812–2822.

[43] KoipallyJ,GeorgopoulosK.IkarosinteractionswithCtBP revealarepressionmechanismthatisindependentof histonedeacetylaseactivity.JBiolChem2000;275 (26):19594–19602.

[44] NgSY,YoshidaT,ZhangJ,etal.Genome-widelineage- speciﬁctranscriptionalnetworksunderscoreIkaros- dependentlymphoidpriminginhematopoieticstemcells.

Immunity2009;30(4):493–507.

(9)

[45] YoshidaT,NgSY,Zuniga-PﬂuckerJC,etal.Early hematopoieticlineagerestrictionsdirectedbyIkaros.Nat Immunol2006;7(4):382–391.

[46] YoshidaT,NgSY,GeorgopoulosK.Awakeninglineage potentialbyIkaros-mediatedtranscriptionalpriming.

Immunology2010;22(2):154–160.

[47] GeorgopoulosK.Haematopoieticcell-fatedecisions, chromatinregulationandIkaros.Immunology2002;2 (3):162–174.

[48] UmetsuSE,WinandyS.Ikarosisaregulatorofil10 expressioninCD4+Tcells.JImmunol2009;183(9):

5518–5525.

[49] QuirionMR,GregoryGD,UmetsuSE,etal.Cuttingedge:

IkarosisaregulatorofTh2celldifferentiation.JImmunol 2009;182(2):741–745.

[50] Gómez-delArcoP,MakiK,GeorgopoulosK.

PhosphorylationcontrolsIkaros'sabilitytonegatively regulatetheG1-Stransition.MolCellBiol2004;24(7):

2797–2807.

[51] PopescuM,GurelZ,RonniT,etal.Ikarosstabilityand pericentromericlocalizationareregulatedbyprotein phosphatase1.JBiolChem2009;284(20):13869–13880.

[52] GurelZ,RonniT,HoS,etal.RecruitmentofIkarosto pericentromericheterochromatinisregulatedby phosphorylation.JBiolChem2008;283(13):8291–8300.

[53] WinandyS.IkarostotherescueofTCR-achaingene rearrangement.EurJImmunol2013;43(2):314–317.

[54] CollinsB,ClambeyET,Scott-BrowneJ,etal.Ikaros promotesrearrangementofTCRagenesinanIkarosnull thymomacellline.EurJImmunol2013;43(2):521–532.

[55] DijonM,BardinF,MuratiA,etal.TheroleofIkarosin humanerythroiddifferentiation.Blood2008;111(3):

1138–1146.

[56] BottardiS,RossJ,BourgoinV,etal.IkarosandGATA-1 combinatorialeffectisrequiredforsilencingofhumang- globingenes.MolCellBiol2009;29(6):1526–1537.

[57] O'neillDW,SchoetzSS,LopezRA,etal.AnIkaros- containingchromatin-remodelingcomplexinadult-type erythroidcells.MolCellBiol2000;20(20):7572–7582.

[58] LopezRA,SchoetzS,DeAngelisK,O'NeillD,BankA.

Multiplehematopoieticdefectsanddelayedglobin switchinginIkarosnullmice.PNAS2002;99(2):602–607.

[59] O'neillD,YangJ,Erdjument-BromageH,etal.Tissue- speciﬁcanddevelopmentalstage-speciﬁcbindingofaSWI/

SNFcomplexassociatedwithhumanfetaltoadultglobin.

ProcNatlAcadSciUSA1999;96(2):349–354.

[60] KeysJR,TallackMR,ZhanY,etal.AmechanismforIkaros regulationofhumanglobingeneswitching.BrJHaematol 2008;141(3):398–406.

[61] WinandyS,WuP,GeorgopoulosK.Adominantmutationin theIkarosgeneleadstorapiddevelopmentofleukemiaand lymphoma.Cell1995;83(2):289–299.

[62] MullighanCG.Themoleculargeneticmakeupof acutelymphoblasticleukemia.Hematology2012;2012:

389–396.

[63] SunL,GoodmanPA,WoodCM,etal.Expressionof aberrantlysplicedoncogenicIkarosisoformsinchildhood acutelymphoblasticleukemia.JClinOncol1999;17 (12):3753–3766.

[64] MullighanCG.BCR-ABL1lymphoblasticleukaemiais characterizedbythedeletionofIkaros.Nature2008;453 (7191):110–114.

[65] JägerR,GisslingerH,PassamontiF,etal.Deletionsofthe transcriptionfactorIkarosinmyeloproliferative

neoplasms.Leukemia2010;24(7):1290–1298.

[66] TefferiA.Novelmutationsandtheirfunctionalandclinical relevanceinmyeloproliferativeneoplasms:JAK2,MPL, TET2,ASXL1,CBL,IDHandIKZF1.Leukemia2010;24 (6):1128–1138.

[67] PapathanasiouP,PerkinsAC,CobbBS,etal.Widespread failureofhematolymphoiddifferentiationcausedbya recessiveniche-ﬁllingalleleoftheIkarostranscription factor.Immunity2003;19(1):131–144.

[68] GoldmanFD,GurelZ,Al-ZubeidiD,etal.Congenital pancytopeniaandabsenceofBlymphocytesinaneonate withamutationintheIkarosgene.PediatrBloodCancer 2012;58(4):591–597.

[69] ZhuX,AsaSL,EzzatS.Ikarosisregulatedthroughmultiple histonemodiﬁcationsanddeoxyribonucleicacid

methylationinthepituitary.MolEndocrinol2007;21 (5):1205–1215.

[70] DupuisA,GaubMP,LegrainM,etal.Biclonalandbiallelic deletionsoccurin20%ofB-ALLcaseswithIKZF1 mutations.Leukemia2013;27(2):503–507.

[71] GeorgopoulosK.Acutelymphoblasticleukemia–onthe wingsofIKAROS.NEnglJMed2009;360(5):524–526.

[72] VeerA,WaandersE,PietersR,etal.Independentprognostic valueofBCR-ABL1-likesignatureandIKZF1deletion,but nothighCRLF2expression,inchildrenwithB-cellprecursor ALL.Blood2013;122(15):2622–2629.

[73] PastorczakA,GórniakP,SherborneA,etal.Roleof657del5 NBNmutationand7p12.2(IKZF1),9p21(CDKN2A),10q21.2 (ARID5B)and14q11.2(CEBPE)variationandriskof childhoodALLinthePolishpopulation.LeukRes2011;35 (11):1534–1536.

[74] PapaemmanuilE,HoskingFJ,VijayakrishnanJ,etal.Locion 7p12.2,10q21.2and14q11.2areassociatedwithriskof childhoodacutelymphoblasticleukemia.NatGenet2009;41 (9):1006–1010.

[75] SwaffordAD,HowsonJM,DavisonLJ,etal.Analleleof IKZF1(Ikaros)conferringsusceptibilitytochildhoodacute lymphoblasticleukemiaprotectsagainsttype1diabetes.

Diabetes2011;60(3):1041–1044.

[76] IacobucciI,StorlazziCT,CilloniD,etal.Identiﬁcationand molecularcharacterizationofrecurrentgenomicdeletions on7p12intheIKZF1geneinalargecohortofBCR-ABL1- positiveacutelymphoblasticleukemiapatients.Blood 2009;114(10):2159–2167.

[77] MullighanCG,SuX,ZhangJ,etal.DeletionofIKZF1and prognosisinacutelymphoblasticleukemia.NEnglJMed 2009;360(5):470–480.

[78] KuiperRP,WaandersE,vanderVeldenVH,etal.IKZF1 deletionspredictrelapseinuniformlytreatedpediatric precursorB-ALL.Leukemia2010;24(7):1258–1264.

[79] MartinelliG,IacobucciI,StorlazziCT,etal.IKZF1(Ikaros) deletionsinBCR-ABL1-positiveacutelymphoblastic leukemiaareassociatedwithshortdisease-freesurvival andhighrateofcumulativeincidenceofrelapse:aGIMEMA ALWPreport.JClinOncol2009;27(31):5202–5207.

[80] DenBoerML,vanSlegtenhorstM,DeMenezesRX,etal.A subtypeofchildhoodacutelymphoblasticleukaemiawith poortreatmentoutcome:agenome-wideclassiﬁcation study.LancetOncol2009;10(2):125–134.

[81] Perez-AndreuV,RobertsKG,HarveyRC,etal.Inherited GATA3variantsareassociatedwithPh-likechildhoodacute lymphoblasticleukemiaandriskofrelapse.NatGenet 2013;45(12):1494–1498.

[82] EzzatS,ZhuX,LoeperS,FischerS,AsaSL.Tumorderived Ikaros6acetylatestheBcl-XLpromotertoupregulatea survivalsignalinpituitarycells.MolEndocrinol2006;20 (11):2976–2986.

[83] Ferreirós-VidalI,CarrollT,TaylorB,etal.Genome-wide identiﬁcationofIkarostargetselucidatesitscontributionto mouseB-celllineagespeciﬁcationandpre-B-cell

differentiation.Blood2013;121(10):1769–1782.

[84] XiaoM,WuY,YangY,etal.AnalysisofIkaros6expression in73patientswithchronicmyelogenousleukemia.

ZhongguoShiYanXueYeXueZaZhi2011;19(4):907–910.

(10)

[85] IacobucciI,LonettiA,MessaF,etal.Expressionofspliced oncogenicIkarosisoformsinPhiladelphia-positiveacute lymphoblasticleukemiapatientstreatedwithtyrosine kinaseinhibitors–implicationsforanewmechanismof resistance.Blood2008;112(9):3847–3855.

[86] IacobucciI,LonettiA,CilloniD,etal.Identiﬁcationof differentIkaroscDNAtranscriptsinPhiladelphia-positive

adultacutelymphoblasticleukemiabyahigh-throughput capillaryelectrophoresissizingmethod.Haematologica 2008;93(12):1814–1821.

[87] MiJQ,WangX,YaoY,etal.Newlydiagnosedacute lymphoblasticleukemiainChina(II)–prognosisrelatedto geneticabnormalitiesinaseriesof1091cases.Leukemia 2012;26(7):1507–1516.