3 Prognostic factors used to direct therapy Fixed factorsAge, sex, white cell count at diagnosis, cytogeneticsDynamic factorsresponse to treatment correlates with prognosisslow early response (SER) assessed by microscopy predicts relapsepredicts outcome within groups of children with the same fixed risk factors receiving the same therapyBUT the microscope is an insensitive tool for detection of residual leukaemia and most destined to relapse have a rapid response
4 Analysis of outcome for MRC UKALL 97/99 Trial - Stratification of Treatment based on “clinical risk” CharacteristicsNArm A<10 yrs AND WCC <50AND RER62Arm B>10 yrs orWCC >5022Arm CA or B ANDSEROr poor cytos16IntensityofTherapy
5 Analysis of outcome for MRC UKALL 97/99 Trial - Stratification of Treatment based on “clinical risk” CharacteristicsNEFS (%)Arm A<10 yrs AND WCC <50AND RER6284Arm B>10 yrs orWCC >502270Arm CA or B ANDSEROr poor cytos16IntensityofTherapy
6 Analysis of outcome for MRC UKALL 97/99 Trial - Stratification of Treatment based on “clinical risk” CharacteristicsNEFS (%)RelapseArm A<10 yrs AND WCC <50AND RER628411 (52%)Arm B>10 yrs orWCC >5022706 (28%)Arm CA or B ANDSEROr poor cytos164 (20%)IntensityofTherapyOnly 20% of relapse comes from highest risk groupMany of those cured are over-treated
7 Relative frequency of leukaemic cells Haematologicremission10121011107MRD“cure”RelapseMRD AnalysisFollow-upIn yearsDetection limit ofPCR techniquecytomorphologyRelative frequency of leukaemic cellsMinimal Residual Disease
8 Prognostic value of MRD MRD shown to have independent prognostic value in ALLChildhood ALLvan Dongen et al 1998Cavé et al 1998Coustan-Smith et al 1998Relapsed childhood ALLKnechtli et al 1998Eckert et al 2001Goulden et al 2003Adult ALLBruggeman et al 2006Raff et al 2007
10 Quantitative MRD Detection Three methods currently available :Flow cytometric immunophenotypingUtilises tumour associated aberrant immunophenotypesE.g. Presence of myeloid markers on leukaemia blastsReverse transcriptase (RT) PCRUtilises tumour specific RNA targetsE.g. Fusion gene transcriptsReal-time Quantitative (RQ) PCRUtilises patient-specific gene rearrangementsE.g. Immunoglobulin and T-cell receptor gene rearrangementsUtility of method chosen depends upon the aim of the studyImportant considerations: applicability, stability, sensitivity & quantitationRQ-PCR methodology is method of choice in most European MRD-based clinical trials
11 RQ-PCR for MRD analysis Methodology identifies unique Immunoglobulin and/or T-cell receptor gene rearrangements that are clone-specific98% of patients will have at least one clonal rearrangementTwo patient-specific RQ-PCR assays are designed for each patientcapable of detecting one leukaemic cell in a background of 10,000 marrow cellsImportant to prevent false-negative results due to clonal evolution80-85% of patients will have two assays quantitative to 1 in 10,000
12 Scheme of Investigation - identification of patient-specific MRD markerPCR analysis of diagnostic DNAHeteroduplex analysis of PCR productsPurify and sequence clonal rearrangementTTGTAGTAGTTACCAGCTGGGCTATGAATACTTCCAGCACTGGGD regionJ regionN regionIdentification of V, D and J segment usageSynthesis of base patient-specific oligonucleotideIn Patient-specific RQ-PCR for MRD Analysis84
15 MRD risk groups by Regimen - January 2009 CharacteristicsRegisteredMRDHigh riskLow riskRegimen A<10 yrs AND WCC <50AND RER947264 (28%)288 (30%)Regimen B>10 yrs orWCC >50637214 (34%)155 (24%)Total1584478 (30%)443 (28%)
16 Event Free Survival By Trial 100ALL2003 (2003-)88%ALL97-99 ( )80%7574%ALL97 ( )PERCENT502512345TIME IN YEARSAt risk:ALL97 ( )997919865801757732ALL97-99 ( )938889849814769745ALL2003 (2003-)18281368967551201
17 Relapse Risk By Trial ALL PATIENTS ALL97-99 (1999-2002) 10075No.No.Obs./PatientsEventsExp.ALL97-99 ( )ALL97-99 ( )9321621·2ALL2003 (2003-)1839700·7ALL2003 (2003-)PERCENT502P = 0·00012516%8%12345TIME IN YEARSAt risk:ALL97-99 ( )932889848814769744ALL2003 (2003-)18191368953551201
18 Event Free Survival by MRD Risk Group LOW = 95%100INDETERMINATE = 85%75HIGH = 78%PERCENT50No.No.Obs./25PatientsEventsExp.HIGH638681·5LOW604120·3INDETERMINATE738701·212345TIME IN YEARSAt risk:HIGH63845131718768LOW604467311184893INDETERMINATE738566449307161715-JAN-09 17:51:15
19 Proposals for ALL 2010All patients on ALL 2020 will have therapy allocated based on MRDTreatment on ALL 2003 is currently randomised based on MRDIncluding risk groups A, B and CMRD Analysis done at day 28 and week 11Sequential analysis of High Risk disease at 2 or 3 extra time-pointsIdentification of patients with Very High Risk diseaseNovel therapies employedBMT?
20 Acknowledgements Members of UKMRD network: - Barts Gary Wright Maggie CorboSheela MedahunsiUlrika JohannsonSusannah AkikiBristol Jerry HancockPaul ArcherRichard HathwayKayleigh McDonaghPaula WaitsNigel WoodGlasgow Sandra ChudleighMary GardinerFrances FeeLinda SmithNicola CraigAnne SproulSteve McKaySheffield Gill WilsonHelen StuartShilpa HaridasAmal AfifiMiranda DurkieJane HoldenRichard KirkJames BlackburnClinical Co-ordinator: -Nick GouldenUKMRD Steering Committee:-Nick CrossAjay VoraBrenda GibsonDavid GrantChristine HarrisonFinbarr CotterJohn MoppettCLWP and ALL Task ForceESG-MRD-ALL:-Jacques van DongenVincent van der VeldenI-BFM MRD Group
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