PEDIATRICPATHOLOGY
SPECIMENGROSSINGANDSPECIALHANDLINGOVERVIEW
JeffreyD.Goldstein,MD
May2017
Whenyougetacasethatshould/couldbeforPediatricPathology,showittothegrossroomsupervisororpageDr.Goldsteinat31418beforeyoucutitin.Ifitistheweekendorlateatnight,pagetheon-callattendingtodiscussthecase.IfDr.Goldsteinisnotavailableduringtheday,pagethepersonwhoiscoveringPediatricPathology,ortheappropriate“organ-service”pathologist.Itdoesnotmatterhowtrivialyoufeelthecaseis;ifitistrivial,asimplediscussionwillconfirmyourassessmentandyoucanproceedunabated.If,however,itappearsnottobetrivial,itcanbeagreatlearningexperienceforyouandprovidebettercareforyourpatient.
SpecialConsiderations
UCLAisaparticipatingmemberoftheChildren'sOncologyGroup(COG)andatissuebankforpediatricneoplasmsmaintainedattheCOGBiopathologyCenter(BPC)atNationwideChildren’sHospital/OhioStateUniversity.Clinicalprotocolschangeallthetimeandthefollowingtumorshaveorstillrequirespecialtissuehandling:WilmsTumor,neuroblastoma,rhabdomyosarcoma,lymphoma,germcelltumors,andmalignantbraintumors.Manypediatriconcologypatientswillberandomizedintotherapeuticprotocols.Sincetheprotocolsandtrialstudiesareinflux,bettertocheckifanyspecialthingsneedtobedoneBEFOREproceedingwithcasesinthesecategoriesorothersituationsthatmayseemunusualorrare(whichoccursinpediatric-agedpatients!)
AllCOGtreatmentprotocolsrequirecentralpathologyreview,andinsomecases,anexpeditedrapidreviewisnecessarytodeterminethecorrectinitialtreatmentregimenforthechild.Therefore,forallchildrenregisteredonprotocol,acompleteduplicatesetofsequentialslidesfromeachblockshouldbeorderedatthetimeofinitialhistologicprocessing.
Forallpediatrictumorsforwhichthereissufficientmaterialavailable,aftersatisfyingprotocolrequirementsandourneeds(includingourTPCL),additionalfrozentissuecanbesubmittedtotheBPC.TPCLpersonnelwillbeavailableduringregularworkhourstoassistwiththeprocurementoftissueforCOGprotocolsandtissuebanking.
SpecimenProcurementKit(NOTE:Thisisusedsporadically,notallcases!):
AspecialbiologykitforpediatrictumorsmaybeavailableintheforpediatrictumorstobesenttoBPC.Usually,itisprovidedbyrequest,typicallyfromthePedsHeme/OncteamrelayedtoDr.Goldsteinoraserviceattending,whowillcontactyouintheeventthisisneeded.AkitmaybeobtainedbycallingthePedsHeme/OncClinicalResearchAssociate(CRA)throughtheiroffice,x56708.Thekitisequipedwith:
1.plastictubesoraluminumforfrozentissue
2.truncatedembeddingmoldsfortumorfrozeninOCT
3.formalincontainersforfixedtissue
4.chargedslidesfortouchimprints
5.culturetubewithmediaforfreshtumorwithseparatemailingcontainer
6.pre-printedFederalExpressairbill
7.kitinstructions,SpecimenTransmittalForm,reimbursementinvoice,Biohazardsticker,dry-icelabelandaFederalExpressstickerforSaturdayDelivery
Additionally,alaminatedwallchartispostedinthecuttingareawhichillustrateshowtissueshouldbeprocessedusingthisspecialkit.Thedetailsforusageofkitmaterialsisdescribedintheinstructionsaccompanyingthekit.
Certainpediatrictumorcaseshavespecializeddiagnostictermsandcriteriathatassistdecisionswithtreatmentandprotocolenrollment.Theseincludeneuroblastoma,Wilmstumor,hepatoblastoma,Ewingsarcoma/PNETandpotentiallyothertumors,suchasrhabdomyosarcoma,germcellneoplasmsandleukemias/lymphomas.CollegeofAmericanPathologists’pediatrictumorsynoptic reports should beused, when applicable,andthefullCAPprotocolsmaybe reviewed foradditionalinformation. Synoptic reports shouldbe usedfor othertumorsthatoccurinchildrenandadolescents.ForpediatricsolidtumorsnotsignedoutbyDr.Goldstein,outsidereviewandconsultationmayberequiredbaseduponadiscussionwithyoursignoutattending.
Inquiriesrelatedtoanypediatrictumoraretobedirectedto:
1.JeffreyGoldstein,M.D.,x57443,Beeper31418;
2.TheHematopathologyandNeuropathologyfelloworattending-on-call,forthosecases.
3.PedsHem/OncClinicalResearchAssociate(CRA),x56708..
4.ThePedsHeme/OncFellowatx56708,orthepageoperatorforthefellowon-call
5.NoahFedermanp21525orx56708forsolidtumorserviceandWilliamMay(leukemia/lymphoma)officex56708pager10205.
ChromosomalAnalysis
Itisadvisabletosavethetissueforchromosomaland/ormolecularanalysisofthefollowingneoplasticdisorders:
1.Wilmstumor
2.Neuroblastoma
3.Rhabdomyosarcoma(especiallyalveolarsubtype)
4.Ewing’ssarcoma/PNET/Demoplasticsmallroundcelltumor
5.Burkittandothernon-Hodgkinlymphomas
6.Acuteleukemiaandgranulocyticsarcoma
7.Germcelltumors
8.Malignantbraintumors
9.Synovialsarcoma
10.Anyrare,unusualorundiagnosedpediatrictumor
Ifchromosomeanalysisisneededonanypediatrictumor,obtainRPMImediumfrom tubesprovidedbytheFlow CytometryLaboratoryintheSurgical Pathology refrigerator.Alternatively,theCytogeneticsLaboratory(300MedPlaza,Room3158)canprovideRPMImedia.YoumaycalltheCytogeneticsLabatx56678 andtheywillprovideyouwiththeRPMImedia.ThislabisopenMonday throughFriday.PleasecontactDr.NageshRao(Pgr#92239)forafterhoursor weekendrequestsiftheSurgicalPathologysupplyisoutorold.Freshtissueof2- 3mmsizeisOKforthestudy.
SpecificSpecimenProcessing
Eachofthefollowingspecimenshasauniqueprotocolforprocessingas outlinedbelow.Pleaserefertothediagramsattachedforgrossing illustrationsandsampledictations(kindlyprovidedbyDr.FloretteK,Gray Hazard,LucillePackardChildren’sHospital,StanfordUniversitySchoolof Medicine.)“Pilot”sectionsoftumorsobtainedpriortofixationmaybesubmittedfornextdaypreviewandpreliminarydiagnosis.Blockmapsonphotographssimilartothoseintheillustrationsareencouragedforlargeand complexspecimens.
WILMS
SmallBiopsies(Usuallycontraindicatedastheyresultinupstaging):
1.Thesmallbiopsyspecimensshouldbesubmittedentirelyinformalin.
2.If there is adequate tissue, a portion should be frozen in liquid nitrogen.
3.Ifthereisadequatetissue,aportionshouldbesavedforchromosomeanalysis.
4.Ifthereisadequatetissue,aportionshouldbesavedforEMin glutaraldehyde.(ThedecisiontoprocesstheEMspecimencanbemadelater.)
Nephrectomies
1.Photographallsurfacesoftheintactspecimenbeforeinkingorbivalving,
2.Weighthespecimenbeforefurthermanipulation,
3.Applyinktothesurface,andletthisdrybeforemakingtheinitialbivalvingcut,
4.Bivalvethespecimencarefully,choosingtheplaneofincisiontoprovideoptimaldemonstrationoftherelationshipbetweenthetumorandthekidney,
5.Donotstripthecapsulefromthecorticalsurfaces
6.Followingtheinitialbivalvingcut,submitfreshtissueforspecialstudies
- jE_SnapfreezenormalkidneyandtumorforsubmissiontotheBiopathologyCenterand/orsubmittedtoourTPCL
- jE_Submittissueforcytogeneticevaluation,whenindicated
- jE_Placetissueingluteraldehydeforelectronmicroscopy,whenindicated
7.Submitinitialpilotsectionsformicroscopicdiagnosis,whennecessary
8.Submitureteralandvascularmarginsofresection,
9.Searchcarefullyinthehilarregionforanylymphnodesthatmightbepresent.
10.Wheneverpossible,makeparallelcutsofthespecimeninaplaneparalleltotheoriginalbivalvingincisionintoslabs2-3cm.thickandplaceinalargecontainerofformalinorothersuitablefixativeinarefrigeratedenvironment,fixingseveralhourstoovernightbeforetakingstagingsections.Theadditionaltimerequiredforthisstepwillenhancethequalityofthestagingsections,andfacilitateaccurateevaluationofcapsulesandmargins.
Samplingofnephrectomyspecimenforhistologicevaluation:
1.Severalimportantconsiderationswillenhancethequalityofthefinalpathologicevaluation.
2.Beforesampling,prepareadrawing,photographorothermappingdeviceonwhichtomarktheexactsitefromwhicheachsectionwasobtained.Thisisofgreatimportanceinviewof thedefinitionsoffocalanddiffuseanaplasia.Includeanydistinctiveinternaltumorfociinthesample.
3.Takemostrandomtumorsectionsfromtheperipheryofthelesion,toshowrelationshipbetweenthetumorandtherenalcapsule,thespecimensurfaces,therenalparenchymaandtherenalsinus.Lookcloselyforanydistinctivechangesintherenalparenchymathatmightbenephrogenicrests.Samplenormalrenalparenchymagenerously.
4.Submitaminimumofonegeneroussectionoftumorforeverycentimeterofgreatesttumordiameter.Formulticentrictumors,thisrulecanbeappliedinsamplingeachindividualtumor.
Furtherinformationregardingmicroscopy,gradingandstagingmaybeobtainedfromtheCOGprotocol;contactDr.Goldsteinforadigitalcopy.
•
NEUROBLASTOMAANDRELATEDTUMORS
Specimensmaybesmallbiopsies,primaryresectionsorpost-treatmentresections
1.Describeappearanceanddimensions;weighexcisionalspecimens
2.Photographlargerspecimenspriortoinkingandslicing
3.SubmitfreshtissueforcytogeneticsandFISHforMYCamplification
4.IfthepatientisregisteredonaCOGprotocol,aportionofthetumor,ideallyatleast1g,shouldbesnap-frozeninliquidnitrogen(withoutOCT).Atleastonepieceoftissuefromtheprimary(ifpresent)andfrommetastaticareas(ifpresent)shouldbecutinto3-5mmslicesandwrappedinthefoilandsnapfrozeninthevaporphaseofliquidnitrogen(donotsubmergethetissueinliquidnitrogen)orisopentane/dryice.TissuemayalsobesubmittedtoourTPCL.
5.ForCOGprotocolpatients,optionalfreshtissuemayberequested.
6.Contact PedsHem/OncClinicalResearchAssociate(CRA),x56708fordistributionofmaterials.
7.SavesmallportionforEM.
8.Submit portions for routine histology, after overnight fixation forlargerspecimens
BIOPSIESANDSMALLSPECIMENSOFPEDIATRICSARCOMASANDLOCALLY AGGRESSIVENEOPLASMS
1.SaveasmallportionforEM. Iftissueisscanty,removea1mmcubeforthisstep.
2.Submit fresh tissue for cytogenetics and FISH studies as may beindicated.
3.ForCOGprotocolpatients,ideallyatleast1g,oftumorandadjacentnormaltissue,ifavailableshouldbecutinto3-5mmslicesandwrappedseparatelyinfoilandsnapfrozeninthevaporphaseofliquidnitrogen(donotsubmergethetissueinliquidnitrogen)orisopentane/dryice.TissueshouldalsobefrozeninOCTinanembeddingmold,andwet,formalinfixedtissueoradditionalparaffinblockspreparedforreferralarealsorequested.
PedsHem/OncClinicalResearchAssociate(CRA),x56708fordistributionofmaterials.
4.TissuemayalsobesubmittedtoourTPCL.
5. Submit adequate tissue for routine histology.This step takesprecedenceover#2,3&4,iftissueveryscanty.
TheBoneandSoftTissueServicehasproceduresforhandlingmajorresectionsofsarcomasandotherneoplasms.SpecificrequirementsforCOGprotocols,ifthepatientisregistered,willbeprovidedpriortospecimenreceipt.
LYMPHOMA
Pleasehandlelymphomaspecimensas per Hematopathology procedures.Thediagrambelowmaybeusedtoguidetissuetriage.SpecificrequirementsforCOGprotocols,ifthepatientisregistered,willbeprovidedpriortospecimenreceipt.
MalignantBrainTumorsPleasehandlebraintumorspecimensasperNeuropathologyproceduresforcollectionoftissueforcytogeneticsandFISHstudies,moleculargenetics,andTPCLtissuebanking.
ForchildrenenrolledonCOGprotocols,snapfreeze100-200mgoftissueinliquidnitrogen(withoutOCT)and/orOCTembedded(1mold–100mgtissue) Ifavailable,50mgofwetformalinfixedtissueand50-200mgoffreshtissueintissueculture
mediaarealsorequested. Otherprotocolspecificrequirementswillbeprovidedpriortospecimenreceipt.ContactthePedsHem/OncClinicalResearchAssociate(CRA),x56708fordistributionofmaterials.
Neuroblastoma
Pre-treatment
Pre-treatmentwithhemorrhagicnodules
Representativesectionsaretakenandsubmittedasfollows:A1 –tumor(pilotsection)
A2–Representativesections
A4
A5,etc…
SampleGrossTemplatePediatricNeuroblastoma
Received[withoutfixative/in formalin] labeledwith the patient's name"[#]",medical record numberanddesignated"[***]"isa[***]g,[***x***x***]cm[irregular/round/oval]portionof[red/yellow/tan/brown]softtissue. Itisentirelycomprisedoftumor[ORitiscomprisedoftumorandnormal-appearingadrenalgland,smallbowel,kidneylocatedalongtheperiphery].Theexternalsurfaceis[intact/disrupted]and[does/doesnot]showafocusofpossiblerupture.Theexternalsurfaceis inked[insertcolor][andthefocusofpossibleruptureis inked [insertcolor]].Thespecimenisbisectedalongitslongestaxistoreveal[homogeneous/heterogeneous],[insertcolor],[firm,soft]tumoraltissue. Fociofhemorrhageandnecrosis[are/arenot] present.[If present, describeloca8on:distributedthroughout,alongtheperiphery].Well-circumscribed,dis8nct,hemorrhagicnodules[are/arenot]present.Thetumor[directlyabuts/ispresent#cmfrom]theinkedresectionmargin. [Photographthe cut surface.] Representative portionsoffreshtissuearefrozenat-80Cforpossible future ancillary studies andportionsaresubmi;edinRPMIforcytogeneticanalysis. Followingfixation,thespecimenisseriallysectionedtoreveal[noadditionallesions/additionallesions(describeifpresent)].[Photographanyunusualfeatures.]
Representativesectionsaretakenandsubmittedasfollows:A1 –tumor(pilotsection)
A2–Gallbladder(ifattached)
A3– Tumortoresection margin (if segmentalresection)A4–Tumor
A6,etc…
SampleGrossTemplatePediatricLivertumor
Received[fresh/informalin]labeledwiththepatient'sname,medicalrecordnumberanddesignated"[#]"isa[***]g,***x***x***cm[irregular/round/oval]liver[explant/segmentalresection].Thecapsuleis[intact/disrupted]and[does/doesnot]showafocusofpossiblerupture.Thegallbladderis[present/notpresent][andmeasures***x***x***cm.][Ifpresent:Itisopenedtoreveal[insertcolor],[thick/watery]bileanda[smooth/rough]mucosalsurface.]Theexternalsurfaceoftheliveris[insertcolor]and[smooth/nodular].[If segmentalresection:Theresection marginisinked[insertcolor].]Thecapsuleisinked [insertcolor][andthe focusofpossibleruptureisinked[insertcolor].Theliverisbisectedfromsuperiortoinferior(coronalplane)throughthe hepaticveintoreveal[homogeneous/heterogeneous],[insertcolor],[firm, soft]tumoraltissue.The liver is thenserially sectionedshowing [#]tumor nodules.Thetumormeasures[***]x[***]x[***]cm.[Ifmorethanonenodule,measureeachnodule.]Fociofhemorrhageandnecrosis [are/are not]present.[Ifpresent,describelocation:distributedthroughout,alongtheperiphery].Thehepaticvein[is/isnot] involved bytumor.Tumor[directlyabuts/ispresent#cmfrom]theinkedresectionmargin.Theuninvolvedliverparenchymais[insertcolor]andis[unremarkable/diffuselynodular].Photograph the cut surface.] Representative portions of fresh tissuearefrozenat -80CforpossiblefutureancillarystudiesandportionsaresubmittedinRPMIforcytogeneticanalysis. PilotsectionsoffreshtumoraresubmittedincassettesA1andA2. Followingfixation,representativesectionsaresubmittedasdescribedbelow. [#/No]candidatehilarlymphnodes are identified. [Photograph anyunusual features.]
PediatricKidneyTumor
CapsuleCentralHilarfat
Representative sections are submitted as described below and illustrated by the accompanying block map.
A1 – Random tumor (pilot section)
A2 – Hilar margins (ureter, renal artery, renal vein)A3-AX –Tumor to capsule (most sections from here)
Center (1-2 sections) Hilar fat (all)
AY– Normal kidney
Additional sections:Adrenal gland, nephrogenic rests, other abnormalities
Sample Gross Template PediatricKidney tumor
Received[fresh/informalin]labeledwiththepatient'sname,medicalrecordnumberanddesignated"[***]" isa[***] g, [***]x[***]x[***]cm[irregular/round/oval]kidney [with/without]perinephricadiposetissue.The capsule is [intact/disrupted] and [does/does not] show a focus of possible rupture. The adrenalgland is [present/notpresent].The ureter,renalartery and renalveinarelocated in the renal hilum [ifnot, please modify].The distalmargin of the ureter,renalartery and renalveinareremoved andsubmittedwithincassetteA3,asdescribedbelow. Theexternalsurfaceofthespecimenisinked[insertcolor] [and the focus of possible rupture is inked [insert color].The specimen is bisectedthrough theuretertoreveal[homogeneous/heterogeneous],[insertcolor],[firm,soft]tumoraltissuelocatedinthe[upper pole/middle/lower pole/diffusely effacing the renalparenchyma].[#] tumor nodules arepresent. The tumor measures [***] x [***] x [***] cm.Foci of hemorrhage and necrosis[are/are not] present.[Ifpresent,describelocation:distributedthroughout,alongtheperiphery]. [Iftheadrenalglandispresent:Tumor [does/does not] invade the adrenal gland.]Tumor [does/does not] invade the renalcapsule [ifso,statewhereand towhatextent]. Therenal sinusadiposetissue[is/isnot] involvedbytumor. [Iftherenalsinusadiposetissueisinvolved,statethe%ofadiposetissue replacedbytumor.] Tumor[directlyabuts/ispresent#cmfrom]theinkedresectionmargin. Theuninvolvedrenalparenchyma is [unremarkable/diffuselynodular/contains [#] wellcircumscribed nodules suspicious fornephrogenicrests]. [Photographthecutsurface.] Representativeportions offreshtissuearefrozenat-80Cforpossiblefutureancillarystudiesandpor8onsaresubmittedinRPMIforcytogeneneticanalysis.PilotsectionsoffreshtumoraresubmittedincassettesA1andA2. Following fixation,thespecimenisseriallysectionedfromanteriortoposteriorinthecoronalplanetoreveal[noadditionallesions/additionallesions(describeifpresent)].[#/No]candidatehilarlymphnodesareidentified. Theentirerenalsinusadiposetissueissubmittedasdescribedbelow. [Photographanyunusualfeatures.]
Bone
Afullcrosssectionofthetumorissubmittedasdescribedbelowandillustratedintheaccompanyingblockmap.
A1–Surgicalresectionmarginskin,softtissue
A2–Surgicalresectionmarginbone marrowA3,etc…
SampleGrossTemplatePediatricBonetumor
Received[fresh/informalin]labeledwiththepatient'sname,medicalrecordnumberanddesignated"[***]"isa[***]x[***]x[***]cm[listbonereceived]amputated[above/below]the[nameclosestjoint:elbow,knee,shoulder].
Theexternalskinis[insertcolor]with[noscars/multiplescars/adominantscar]located[#]cmfromtheresectionmargin.[No]obvioustumorisidentifiedalongthesurfaceofthespecimen.[Iftumorisseen,measureanddescribe.] Representative sections ofthe skin and soft tissueas well asportionsofbonemarrowscoopedfromtheboneatthesurgicalresectionmarginaretakenandsubmittedincassettesA1andA2,asdescribedbelow.Thespecimenisfrozenovernightandseriallysectionedlongitudinallyusingabandsaw.Thesefrozenlongitudinalsectionsreveal[homogeneous/heterogeneous],[insertcolor],tumoraltissuemeasuring[***x***x***]cm.
There[is/are][#]tumornodule(s)present.[Ifmultipletumornodulesare
present,describe.]Thetumorislocatedwithinthemedullarycavity[with/without]extensionintotheadjacentsofttissue.Thejointspace[is/isnot]involvedbytumor.Thetumorispresent[#]cmfromthesurgical resectionmarginand[#]cmfromtheclosestsofttissuemargin.Frozenlongitudinalsectionsthatcontainthegreatestcrosssectionoftumorarefixedinformalinanddecalcified.