RAS, KIT, FLT3 and JAK2 gene mutations in Acute Myeloid Leukemia (AML) with inv(16) and t(8;21) [Elektronische Ressource] : incidence and relevance on clinical outcome / presented by Juan Du

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Sujets

• Gesundheit

Informations

• Publié par: universitat_ulm
• Publié le: 01 janvier 2007
• Nombre de lectures: 7
• Langue: English
• Poids de l'ouvrage: 1 Mo

Extrait

UniversitätsklinikumUlm
ZentrumfürInnereMedizin
KlinikfürInnereMedizinIII
Hämatologie,Onkologie,Rheumatologieund
Infektionskrankheiten
ÄrztlicherDirektor:Prof.Dr.med.HartmutDöhner


RAS,KIT,FLT3andJAK2 GeneMutationsin
AcuteMyeloidLeukemia(AML)withinv(16)
andt(8;21):IncidenceandRelevanceon
ClinicalOutcome



Dissertation
fortheattainmentofthe
DoctoralDegreeofMedicine(Dr.med.)
attheFacultyofMedicine,UniversityofUlm,Ulm,Germany



PresentedbyJuanDu
borninMudanjiang,HeilongjiangProvince,P.R.China


2007









AmtierenderDekan:
1.Berichterstatter:
2.Berichterstatter:



TagderPromotion:


Content
Content
Abbreviations………………………………………………………………III
1. Introduction........................................................................................... 1
1.1. Acutemyeloidleukemia.......................................................................... 1
1.2. Prognosticsubgroupsbasedoncytogeneticfindings............................. 3
1.2.1. Corebindingfactor(CBF)acutemyeloidleukemia................................. 5
1.3. MutationclassesinAML......................................................................... 9
1.3.1. ClassImutations .................................................................................. 10
1.3.2. ClassIImutations ................................................................................. 15
1.4. Theaimofthestudy ............................................................................. 16
2. MaterialandMethods ......................................................................... 18
2.1. Material................................................................................................. 18
2.1.1. DNAextraction...................................................................................... 18
2.1.2. Polymerasechainreaction(PCR)......................................................... 18
2.1.3. Agarosegelelectrophoresis ................................................................. 20
2.1.4. Denaturinghighperformanceliquidchromatography(DHPLC)............ 20
2.1.5. PurificationofPCRproducts................................................................. 21
2.1.6. Cyclesequencingreaction(CSR)......................................................... 21
2.1.7. Dyeremoval.......................................................................................... 21
2.1.8. DirectDNAsequencing ........................................................................ 21
2.2. Patients................................................................................................. 21
2.3. Treatment ............................................................................................. 23
2.4. Cytogeneticandmoleculargeneticanalysis......................................... 25
2.5. AnalysisofRAS,KIT,FLT3andJAK2genemutations ........................ 25
2.5.1. DNAextraction...................................................................................... 25
2.5.2. Polymerasechainreaction(PCR)......................................................... 26
2.5.3. PCRwithrestrictionfragmentlengthpolymorphism(PCRCRFLP)........ 26
2.5.4. DetectionofJAK2mutationsusingalleleCspecificPCR....................... 27
2.5.5. Agarosegelelectrophoresis ................................................................. 28
2.5.6. IsolationoftheDNAfragmentsfromagarosegels ............................... 28
2.5.7. Denaturinghighperformanceliquidchromatography(DHPLC)............ 28
2.5.8. PurificationofPCRproducts................................................................. 30
2.5.9. Cyclesequencingreaction(CSR)......................................................... 31
IContent
2.5.10. DyeremovalusingDyeExspinkit ........................................................ 31
2.5.11. DirectDNAsequencing ........................................................................ 31
2.5.12. SensitivityofDHPLCandsequencingassay........................................ 31
2.6. Statisticalanalyses ............................................................................... 32
3. Results................................................................................................. 33
3.1. IncidenceofRAS,KIT,FLT3andJAK2genemutations ...................... 33
3.1.1. RASmutations...................................................................................... 34
3.1.2. KITmutations........................................................................................ 35
3.1.3. FLT3mutations ..................................................................................... 36
3.1.4. JAK2mutations..................................................................................... 36
3.2. Associatedgenemutations................................................................... 36
3.2.1. Associatedgenemutationsininv(16)Cpositivepatients ........................ 36
3.2.2. Associatedgenemutationsint(8;21)Cpositivepatients......................... 37
3.3. Patientscharacteristics......................................................................... 38
3.3.1. Pretreatmentcharacteristicsforinv(16)Cpositivepatients ..................... 38
3.3.2. Pretreatmentcharacteristicsfort(8;21)Cpositivepatients...................... 43
3.4. Responsetoinductiontherapy ............................................................. 48
3.4.1. Responsetoinductiontherapyforinv(16)Cpositivep atients ................. 48
3.4.2. Responsetoinductiontherapyfort(8;21)Cpositivep atients.................. 48
3.5. Survivalanalysis................................................................................... 51
3.5.1. Survivalanalysisforinv(16)Cpositivepatients ...................................... 51
3.5.2. Survivalanalysisfort(8;21)Cpositivepatients........................................ 56
4. Discussion .......................................................................................... 60
4.1. RASanalysis ........................................................................................ 60
4.2. KITanalysis .......................................................................................... 62
4.3. FLT3analysis ....................................................................................... 64
4.4. JAK2analysis ....................................................................................... 66
4.5. Conclusion............................................................................................ 67
5. Summary ............................................................................................. 69
6. References .......................................................................................... 71
7. Curriculumvitae ................................................................................. 87
8. Acknowledgements............................................................................ 90
IIAbbreviations
Abbreviations
ACN Acetonitrile
Allo6SCT AllogeneicstemCcelltransplantation
AML Acutemyeloidleukemia
AMLSG AMLStudyGroup
APL Acutepromyelocyticleukemia
ATRA alltransretinoicacid
Auto6SCT AutologousstemCcelltransplantation
BM Bonemarrow
bp Basepair
CBF CoreCbindingfactor
CBFB CoreCbindingfactorβ
CBFs CoreCbindingfactors
CEP AntiCcancercompound
CI Confidenceinterval
CIR Cumulativeincidenceofrelapse
CALGB CancerandLeukemiaassociatedGroupB
CMPD Chronicmyeloproliferativediseases
CNS Centralnervoussystem
CR Completeremission
CRi Completeremissionwithincompletehematologicalrecovery
CSR Cyclesequencingreaction
DFS DiseaseCfreesurvival
DHPLC Denaturinghighperformanceliquidchromatography
DNA Deoxyribonucleicacid
dNTP Deoxynucleotidetriphosphate
ED Earlydeath
EDTA Ethylenediaminetetraaceticacid
EFS EventCfreesurvival
EML Extramedullaryinvolvement
ETO EighttwentyCone
EUN NCethylCNCnitorsourea
FAB FrenchCAmericanCBritish
IIIAbbreviations
Fluorescenceinsituhybridization FISH
FTIs Farnesyltransferaseinhibitors
GISTs Gastrointestinalstromaltumors
HAM HighCdosecytarabineandmitoxantrone
HiDAC HighCdosecytarabine
HOVON HematoCOncologievoorVolwassenenNederland
HPV Humanpapillomavirus
HR Hazardratio
HSC Hematopoieticstemcell
HSCs Hematopoieticstemcells
HSPCs Hematopoieticstem/progenitorcells
HUGO HumanGenomeOrganization
IC Idarubicin,cytarabine
ICE Idarubicin,cytarabine,etoposide
IDAC IntermediateCdosecytatabine
IE Idarubicin,etoposide
ins/del Insertion/deletionmutations
inv(16) Inversioninv(16)
ITD Internaltandemduplication
JAK Januskinase
JM Juxtamembrane
KI Kinaseinsert
LDH Lactatedehydrogense
LSCs Leukemiastemcells
MCD Mastcelldisease
MDS Myelodysplasticsyndrome
MgCl Magnesiumchloride2
MgSO Magnesiumsulfate4
MPD Myeloproliferativedisorders
MRCAML10 MedicalResearchCouncil's10thAMLtrial
MYH11 Myosinheavychain11
NCAM Neuralcelladhesionmolecule
OR Oddsratio
OS Overallsurvival
IVAbbreviations
PB PeripheralBlood
PCR Polymerasechainreaction
PCR6RFLP PCRwithrestrictionfragmentlengthpolymorphism
PDGFRa PlateletCderivedgrowthfactorCreceptorCα