MoiseDanielpour,M.D.VeraandPaulGuerinFamilyChairinPediatricNeurosurgeryMedicalDirector,PediatricNeurosurgeryandCenterforPediatricNeurosciencesDepartmentofNeurosurgeryandTheRegenerativeMedicineInstituteCedars-SinaiMedicalCenter

¡  ConsultantforBiomarinPharmaceuticalsforclinicaldrugtrialforforamenmagnumstenosisinachondroplasia

¡  Noconflictofinterest

0 500 1000 1500 2000

CancerRemainsaLeadingCausesofDeathamong1-14Year-OldsU.S.,1991vs.1998

AccidentsCancer*

BirthDefectsHomicide

HeartDiseasesPneu&Influenza

CerebralPalsySuicide

HIVInfectionObstruc.LungDis.

Dis.ofInfancyCerebrovasc.Dis.

SepticemiaViralDiseases

Meningitis

19911998

1991:Ca-ACaJforClinJan-Feb,'951998:Ca-ACaJforClin51:15-36,'01

*includesfatal'benign'tumors

51896325

¡ Morethan7000newcasesofchildhoodcancersarediagnosedeachyearintheUnitedStates

¡  CentralNervousSystem(CNS)Tumorsarethemostcommonsolidtumorofchildhood

¡  ThemorbiditycausedbyCNStumorsandtheirtreatmentexceedsthatofmostotherchildhoodcancers

¡  EarlydiagnosisandextentofsurgicalresectionofCNStumorsinchildrenarethestrongestpredictorsofincreasedsurvivalorcure§  SignsandSymptoms§  RoleofSurgeryandChemotherapy▪  AdvancesleadingtodecreasedMorbidity

§  PrecisionMedicineandPersonalizedModelingofCancers

§  Advancesinthelaboratory

¡  Seizures¡  Ataxia/Dysmetria¡  Vomiting¡  CranialNeuropaties¡  Macrocephaly

§  Nausea§  Headache§  VisualDisturbance§  AlteredMentalStatus§  Pain§  Weakness

¡  Occursin84%ofpediatricpatientswithbraintumors

¡ MaybeduetogeneralizedelevatedICP,orcompressionofvagalnucleiorvomitingcentersinflooroffourthventricle

¡  Earlymorningheadachesandvomiting¡ Waxingandwaningsymptomsoftenwithimprovementofheadachesafteremesis

¡  Rareearlyindicationofamasslesion:§  Initialpresentationin15%ofchildrenwithsupratentoriallesions

¡  Temporallobemostcommonsitefortumorsproducingseizuresasanearlysign§  Canpresentasabsenceseizures§  Difficulttodetect:staring,Lackofawareness,suddenhaltinactivity,lipsmackingetc.

§  behavioralissuesanddeclineinschoolperformance

¡  Gaitdisturbancesmaysuggesteitherabrainstemorcerebellarprocess

¡  Triadoflong-tractsigns,cranialneuropaties,andataxiasuggestsbrainstempathology

¡  Cerebellarlesions§  Midlinelesions=truncalataxia

§  Cerebellarhemisphere=Appendicularataxia

¡  Seeninmultipletypesofbraintumors¡  Producedbydirectcompression/invasionor¡  IndirectlyduetoincreasedICP

§ MostcommonlyCNVI▪  Diplopiawithcomplimentarystrabismusorheadtilt

¡  MostCommon--Diplopia,asymmetriccranialnervedeficitsanddifficultywalking

¡  HydrocephalusSyndrome--Headaches,vomitingandventriculomegaly

¡  PsychiatricSyndrome1--Apathy,depression,declineinschoolperformanceandmemoryloss

¡  PsychiatricSyndrome2--Behavioralchange,nightmaresandenuresis

¡  StrokeorhemorrhagicSyndrome--Acutestrokelikeonsetofhemiplegia,quadraparesis,intra-nuclearophthalmoplegiaandupbeatnystagmus

¡  --Cerebello-pontineanglepresentationwithinvolvementofcranialnervesV,VIIandataxia

¡  Backpaininachildmaybewarningofundelyingspinalcordlesion§ Whenpresentingwithintracranialsignsorsymptomsmayrepresentadropmetastasisfromintracranialtumor

Tumorwithdropmetastasis

¡ Weaknessofextremetiescommonlyseenwithsupratentorialtumors

¡  Canbepresentingsignofspinalcordcompression

¡  Otherassociatedsignsandsymptomsmustbeelicitedtodifferentiatebetweenthetwotypesofpresentation

¡  YoungchildrencanaccommodatelargetumorswithminimalsignsofraisedICP

¡  PresenceofopensuturesorfrontalbossingisalongstandingsignofincreasedICP.

¡  Imperativetomeasureachild’sheadcircumferenceifanysignsofachangeinmentalstatus,irritabilityorlethargyoccurs.

¡  Directinvolvementofvisualpathway§  Opticglioma,hypothalamicgliomas,visualcortexandcraniopharyngioma:visualfieldcuts

§  pinealandtectalplatetumors:perinaud’ssign¡  Papilledemasecondarytoraisedintracranialpressure§  Presentinupto90%ofpatientswithcerebellarastrocytoma

¡  Growthdisturbances¡  Precociouspuberty¡  Amenorrhea¡  Hypothyroidism¡  Diabetesinsipidus

¡  42%ofpatientspresentwithhormonaldeficiency(D.I.)andprecociousordelayedpuberty

¡  Ophthalmologicexaminationiscriticalbecausepatientsrarelycomplainofvisualdisturbance

¡  Visualcutsarecommonlyfoundonexam

¡  Frequentlypresentwithhormonaldeficiency¡  Over50%presentwithgrowthhormonedeficiency

¡  Over91%ofpatientshavehypopituitarismwithmultiplehormonaldeficiencies

¡  DiabetesInsipidusisalatesignandfrequentlyunderappreciated

Post-op

Pre-op

¡  Parinaud’ssign§  Paralysisofupwardgazewithretractionnystagmus,pupillarylight-near

dissociation,lidretraction¡  Raisedintracranialpressure:hydrocephalus¡  Diplopia¡  Diabetesinsipidus¡  Hypopituitarism

Pre-opPost-op

0102030405060708090100

resection>90% Residual<1.5cc neurologicalcomplications

GNSDPNSASPN

SurgicalResection

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Neuroanatomy Microneurosurgery Imaging

PRE-OPERATIVEIMAGE

POST-OPERATIVEIMAGE

BRAINTUMOR

¡  Realtimeimages¡  Canaccountforbrainshift¡  Ideallysuitedforuseinchildren

§  Smallersurgicalfield

§  Higherincidenceoflow-gradetumors

§  Donotneedtousetraditionalheadholdingapparatus

¡  AFlexibleLaserapplicatorisguidedtointendedtargetedarea

¡  IntheMRIunitaphysicalpreciselymonitorstreatmentusingsoftwaretomeasuretemperaturechange

¡  Laserlightheatsanddestroystargetarea.Temperaturemapsshowextentoftumortissuebeingdestroyed,minimizingriskofdamagetosurroundinghealthytissue

¡  DeterminantsofDrugEfficacy§  Sensitivityoftumortothedrug

§  Pharmoracokineticsofthedrug

§  Drug’sabilitytocrosstheBBB§  Abilityoftumorcellstoaccumulateandretainthedrug

§  Abilityofthetumorcellstorepairdamagecausedbythedrug

¡  MalignantGlioma§  Increasesthediseasefreesurvivalinbothanaplasticastrocytoma(30%)andGlioblastoma(20%)whenusedfollowingradiotherapyascomparedtothosetreatedwithpostsurgicalradiotherapyalone.

¡  Medulloblastoma§  Upto80%fiveyeardiseasefreesurvivalfollowinggrosstotalresection,radiationandchemotherapycomparedtoabout40%withoutchemotherapy

¡  Ependymoma§  Chemotherapoyhasnotimprovedsurvival

¡  Oligodendrogliomas§  Verylittledataexistregardinguseofchemotherapyforchildrenwithnewlydiagnosedorrecurrentdisease

¡  GermCellTumors§  Dependsonthetypeofgermcelltumor

Leptomeningealtumor

Chemotherapy

Drop-Mets?

TUMORPAINTOptides;targetedtreatmentstokillcacercellswhilesparing

patientsfromtoxiceffectsofchemotherapy

-1993teamatHarvardidentifiedcomponentofscorpianvenomthatblockschloridechannels-1994NicoleUllrichandHaraldSontheimerusedCholorotoxintoblockmovementofGliomacellsandfoundcompoundonlybindsmalignantcells-TransmolecularPhase1andPhase2trials-JimOlsonandBlazeBiosciencewithtumorpainttovisualizetumorinsurgery-Makesurgeonsabletoremovemoretumorwithouthdisturbingnormaltissue-Attachchemotherapeuticdrugstoabindingagentthatonlyattachestotumorcells

¡  Matchingtherightpatient,totherightmedicine,attherighttime.

¡  Mosttumorsdon’thaveasinglegenemutationorsingleabnormalpathway

¡  Mostbraintumorsareheterogenousandconstantlyevolving

Needtogainadeeperunderstandingofmolecularcharacteristicsthataredrivingtumorgrowthand

matchingtreatmenttothosespecificabnormalities

¡  Pediatricgliomasareassociatedwithuniquemutations

¡  Pediatricgliomasoftenresemble“normal”developmentalprograms(e.g.EGLprogenitorsinmedulloblastoma)

¡  PediatricdiseasemorefrequentlypresentsaslowgradewhileadultpresentsasGBM

¡  Notablypediatricdiseasepresentsuniquechallengesduetocoincidentbraindevelopment

¡  Next-gensequencinghasleadtorapiddeterminationofthesomaticmutationsinlowandhighgradegliomas

¡  PediatricGlioma§  Schwartzentruberetal.2012,

Nature§  Zhangetal.2013,NatureGenetics

¡  AdultGlioma§  Brennanetal.2013,Cell§  WilliamsParsonset.al.2008,

Science§  Frattiniet.al.2013,NatureGenetics§  TCGA2008,Nature§  Verhaaketal.2010,CancerCell

¡  Identifytarget¡  Findspecificdrugsorcombinationsofdrugsthatattackthosetargets

¡  Identifybiomarkersthatcanpredictwhichcombinationofdrugswillbeeffectiveforsubgroupofpatient(s)

¡  Thereareanaverageof15somaticmutationsinpediatricglioma—relevanceofeachmutationnotclear

¡  Foradultthisnumberrisesto36somaticmutations

¡  Manymutationsdonothavegeneticallyengineeredmousemodels

Schwartzentrubereta.2012;Nature

¡  EngineeredMouseModels§  Costly§  Takemonths/yearstodesign§  Moretimetocross§  EvenwithatransgenicpromotersuchasNestin,a“non-

physiological”amountofcellsexpressthemutationthroughoutdevelopment

¡  ViralModels§  Safetyconcerns(pantropic)§  Needskillinachievinghightiters§  BreedingneededforRcas-Tvamodels(ecotropic)

¡  TransplantModels§  Immuneinteractionsanissue§  Notautochthonicandlikelydoesnotrecapitulatetumorigenic

process

Developmentofanovelmethodologyforrapidandefficientgenerationofbraintumormodels

¡  ElectroporationistheapplicationofcurrentacrosstheheadtodriveplasmidDNAintothenucleiofdividingcells

¡  Postnatalelectroporationisasimple,“non-invasive”proceduretakinglessthan1hourperlittertocreatesomatictransgenics

¡  Veryfewplasmidsstablyintegrate

¡  Employs“pBase”transposasetostablyintegratethe“geneofinterest”intothegenomeinatargetedfashion

¡  AdaptedbyLoturcoetal.foruseintheembryonicCNS

¡  Postnatalelectroporationallowsfortemporalandspatialcontrolofsomatictransgenesis

¡  PiggyBactranspositionprovidesforpermanenttransgeneintegration

CRISPR/Cas9systemcanbeusedforgeneediting•  Genesilencing•  Replacingnormalgenewithmutantgene

https://www.neb.com/tools-and-resources/feature-articles/crispr-cas9-and-targeted-genome-editing-a-new-era-in-molecular-biology

¡  Stable,somatictransgenicmutation¡  Labelandmutatesimultaneously(i.etumorcellsare

fluorescentagainstawildtype,unlabeledbackground)

¡  Flexible-Multiplegenes/shRNAscanbedeliveredsimultaneously(i.e.patientmutationsignatures)

¡  Autochthonic–fromendogenouscells,allowingnaturalprogressionoftumors

¡  Non-viral–nosafety/immuneconcerns¡  Rapid–requiresonlyPiggyBac-baseddonorplasmids¡  Definedinitialmagnitudeofsomatictransgenesisas

wellasthespatialandtemporalgenesisofthetumor

¡  TheMAPKpathwayishighlyimplicatedingliomagenesis

¡  Rasmutationsareseenin20-25%ofcancers

¡  Erbb2(HER2/Neu)formsaheterodimerwithotherErbb2familymembersandmutationsarefoundinglioma

http://www.biooncology.com/images/therapeutic-targets/

RachelleLevyShawnWagner

RachelleLevy

RachelleLevy

¡  Thecriticalpathologicalhallmarksarenoted

SergueiBannykh,M.D.

RoelVerhaak,Ph.D.

•  miR-Etechnologyisanevolutionofmir30microRNAtechnology•  CanbeusedwithPolIIpromoters(CAG,TRE-Bi,tissuespecificpromoters,etc.)•  Nf1functionstoinhibitRasactivationandisafrequentmutationinpediatricglioma

Schwartzentruberetal.2012,Nature

¡  InhibitionofFattyAcidSynthaseDecreasesExpressionofStemnessMarkersinGliomaStemCells.Yasumotoetal.PloSOne2016Jan25;11(1)

¡  TERTpromotermutationsingliomas,geneticassociationsandclinico-pathologicalcorrelations.LabussièreM1etal,BrJCancer.2014Nov11;111(10):2024-32.doi:10.1038/bjc.2014.538.Epub2014Oct14.

¡  Smallmolecules§  Fasn

¡  Geneticproofofprinciple§  Etsfamily

¡  Immunotherapy§  Tgr1-3

ADDRESSINGTUMORRECURRANCE

74

¡  Cedars-Sinai§  Moise

Danielpour,M.D.§  JoshuaBreunig,

Ph.D.§  RachelleLevy§  C.Danielle

Antonuk§  JessicaMolina-

Arevena§  GibumKim§  AslamAkhtar§  MarinaDutra-

Clarke§  HannahPark

§  SergueiBannykh,

M.D.§  CliveSvendsen,

Ph.D.§  JavierRodriguez§  DavidGate§  PaskoRakicM.D.,

Ph.D.§  Svendsenlab§  ShawnWagner,Ph.D.§  Townlab

¡  Plasmids§  JoeLoturcoPh.D.§  NenadSestanPh.D.

¡  Addgene

¡  Funding

§  SOCCICancerCenterForumAward

§  VeraandPaulGuerinFamilyFoundation

§  SmidtFamilyFoundation

§  RMIandBiomedicalSciences