pediatric AML

1. Approach to
Pediatric AML
By: Mohamadreza Sadri

2. • Acute myeloid leukemia (AML) is a hematopoietic malignancy that
is the culmination of genetic and epigenetic alterations in the
hematopoietic stem/progenitor cells and resulting in the
expansion of undifferentiated myeloid cells
• Childhood acute myeloid leukemia is a rare and heterogeneous
disease, with an incidence of 5-7 cases per million children
younger than 15 years
• Early peak incidence with gradual rise during adolescence
• Incidence of ALL (acute lymphoblastic leukemia) is 5 times higher
than incidence than AML (acute myeloid leukemia)

4. • there are a wide range of inherited chromosomal and gene
defects and that predispose to the development of child
AML:
 trisomy 21 (Down syndrome [DS])
 Fanconi anemia
 dyskeratosis congenita(DC)
 Shwachman-Diamond syndrome (SDS)
 Kostmann syndrome(severe congenital neutropenia or SCN).

5. • Include prognostic factors for AML
Include cytogenetics and molecular data
• Two groups:
- De novo AML
- Secondary AML
• AML that evolves without a prior cytotoxic exposure is
referred to as de novo AML
• Secondary AML refers to evolution of AML subsequent to prior
exposure to cytotoxic therapy or antecedent hematopoietic
insufficiency(eg, myelodysplastic syndrome [MDS]

9. • Diagnosis in childhood AML are :
 morphology with cytochemistry
 immunophenotyping
 karyotyping
 FISH, and specific molecular genetics
in the bone marrow aspiration , which is comparable with
common practice in adults
• The initial diagnostic tests may be done on peripheral
blood if the patient’scondition contraindicates a bone
marrow aspirate

10. • investigation of CNS Involvement at diagnosis is not practiced
routinely in adults but is considered necessary in children
• CNS involvement at diagnosis and at relapse is seen in
5%-10% of pediatric patients with AML
• specific treatment(cranial radiotherapy ) is required to eradicate
hidden AML blasts in the CNS
• CNS positivity is generally define by > 5 × 106/L white blood cells
(WBCs) in the cerebrospinalfluid , with blasts present in a non
bloody tap

11. • The morphologic classification of AML is based on the lineage
associated phenotype
 undifferentiated,
 Myeloid,
 Monoblastic ,
 erythroblastic,
Or
 Megakaryoblastic
• Cytochemistry confirms lineage affiliation and classifies myeloid
(myeloperoxidase [MPO]-positive) and monoblastic differentiation
(nonspecific esterase-positive)

12. • Differentiation between AML and MDS :
• Differentiating between AML and advanced MDS
may be difficult in children with a low percentage
of blasts
• In this case of a low blast count (<20%) and MDS
serial bone marrow aspirates and biopsies are
required, as well as detailed cytogenetic analyses
• Differentiating between AML and MDS is important
for treatment allocation
• MDS can only be cured by hematopoietic stem cell
transplantation (HSCT)

13. • Differentiation between AML and MDS :
• In adults, a blast threshold of 20% is used to
differentiate between AML and MDS
• In children blast percentages between 20% and
30% may be seen in MDS(refractory anemia with
excess of blasts)
• AML-specific genetics, hyperleukocytosis,
extramedullary disease, and progression within a
short time frame (2-4 weeks) are supportive of AML
rather than MDS

14. • ambiguous morphology and cytochemistry,
immunophenotyping support the lineage definition
 Acute megakaryoblastic leukemia (AMKL, FAB M7)
 Minimally differentiated AML(FAB M0)
 between AML and acute lymphoblastic leukemia
(ALL)
 biphenotypic leukemia, bilineage leukemia with
distinctly differentiated blast populations
 undifferentiated leukemia without any lineage
commitment
 minimal residual disease (MRD)

16. • Conventional cytogenetics can detect structural and
numerical cytogenetic abnormalities in 70%-80% of
children with AML
• Cytogenetic alterations have been a cornerstone in
the diagnosis of AML, and best tool for stratification
for treatment
• Certain fusion genes, products from cryptic
translocations, loss of chromosome material can only
be reliably detected using FISH

17. • The most frequent chromosomal abnormalities in children with
AML include :
 25% t(8;21)(q22;q22), inv(16)(p13.1q22) (together referred
as core binding factor [CBF]-AML)
 12% t(15;17)(q22;q21)/PML-RARA
 20% 11q23/MLL-rearranged abnormalities
 20% do not have a discernible karyotypic abnormality
(normal karyotype)
 t(1;22)(p13;q13)/RBM15(OTT)-MKL1(MAL) are more
predominant in pediatric AML

18. • Cryptic translocations, those not amenable to
identification by conventional karyotyping
• require more specialized techniques such as PCR
or (FISH)
t(7;12)(q36;p13)/ETV6(TEL)- HLXB9(MNX1)
t(5;11)(q35;p15.5)/NUP98-NDS1
NUP98/KDM5A
CBFA2T3/GLIS2

20. • Several gene mutations and aberrantly expressed
genes have been recognized in pediatric AML
• AML is result from at least 2 classes of cooperating
mutations, These classes can be categorized as :
type I mutations inducing proliferation, such as
abnormalities in tyrosine kinases
type II mutations, inducing maturation arrest,
comprising most of the translocations

21. • The frequency and nonrandom associations of type I and
type II mutations in pediatric AML differ from adults
• mutations in 3 genes (FLT3, NPM1, and CEBPA) have
been shown to have clinical implications in childhood
AML

22. • The most commonly mutated gene in childhood AML
• constitutive activation of the receptor kinase activity
• internal tandem duplication (FLT3/ITD) of the
juxtamembrane domain coding sequence(15% of all
children with AML)
• missense mutation in the activation loop
domain(FLT3/ALM)

23. • FLT3/ITD be highly associated with poor response to induction
chemotherapy and high relapse rate
• FLT3/ALM do not have increased rates of treatment failure
• Mutations in the WT1 gene are found mainly in CN-AML and are
often associated with FLT3-ITD mutations
• The frequency of activating mutations of tyrosine kinase receptor
genes, such as :
 FLT3
 t(15;17)PML-RARA
and
 t(5;11)NUP98-NSD1, increases with age

24. • NPM1 encodes a ubiquitously molecular chaperone that shuttles
rapidly between the nucleus and cytoplasm
• prevalence of 30% in adult and 8% to 10% in pediatric AML
• mutations, characterized by 4 base insertions in exon 12 of the NPM
gene, lead to impaired nuclearlocalization of the nucleophosmin
protein
• NPM1 mutations appear to be more prevalent in AML with normal
karyotype
• the presence of NPM1 mutations overlap in a subset of patients with
FLT3/ITD, and its coexpression ameliorate the poor prognosis by
FLT3/ITD

25. • The CEBPA gene encodes CCAAT/enhancer binding protein
alpha (C/EBPa)
• transcription factor that regulates granulocytic proliferation and
terminal differentiation
• Mutations in the CEBPA in 5% of childhood AML
• CEBPA mutations occur in patients with normal cytogenetics
• associated with decreased relapse risk and improved survival
• C-KIT mutations occur in 25% of children with CBF-AML

26. • Alterations in DNA methylation can lead to silencing of
genes
 Silencing of the CEBPA gene by promoter methylation
• Aberrant methylation can be caused by genomic
alterations (mutations, deletions,or translocations) in
genes regulating methylation
• methyltransferase genes :
 MLL1, DNMT3A, TET2
• somatic mutations in TET2, IDH1, IDH2, and DNMT3A
are highly prevalent in adult AML but not in childhood
AML

31.  Response to the first course of treatment
 cytogenetics
 molecular genetics
• Cytogenetics with favorable outcome :
CBF-AML
 t(15;17)(q22;q21)
t(1;11)(q21;q23)/MLL-MLLT11(AF1Q)

32.  Cytogenetics with adverse outcome
• t(6;11)(q27;q23)/MLL-MLLT4(AF6),
• t(10;11)(p12;q23)/MLLMLLT10( AF10),
• t(7;12)(q36;p13)/ETV6(TEL)-HLXB9(MNX1),
• t(6;9) (p23;q34)/DEK-NUP214,
• t(5;11)(q35;p15.5)/NUP98-NSD1
• -12p
 Adverse cytogenetics described in adult AML,such as -
5q, inv(3)(q21q26.2) or t(3;3)(q21;q26.2)/RPN1-EVI1,
are very rare in children

34. • Residual disease can be monitored by :
Morphology
immunophenotyping
quantification of molecular aberrations and gene
expression levels

35. • Immunophenotyping :
 MRD assessment by immunophenotyping can be done in up
to 96% of children with AML
 The heterogeneity of leukemia-associated
immunophenotypes and frequent antigen shifts over time can
limit the sensitivity and specificity of immunophenotypic
detection of MRD
 Current technologic advances, such as 6-color flow
cytometry, may overcome any limitations

36. • Fusion genes :
The high specificity and sensitivity (up to 105) of real-time quantitative
PCR of AML fusion genes of :
 RUNX1(AML1)-RUNX1T1(ETO),
 CBFB-MYH11,
 PML-RARA,
 MLLT3(AF9)-MLL
• Mutation-specific MRD :
 NPM1,
 FLT3-ITD, or
 GATA1s
Additional candidate mutations :
WT1, C-KIT, N-/K-RAS, PTPN11, or FLT3- point mutations.

37. • The intensification of conventional chemotherapy along
with improvements in supportive care has improved the
prognosis in childhood AML
• side effects and toxicity still remain a major concern
• New compounds, such as :
 epigenetically active agents,
 tyrosine kinase inhibitors,
 antibody-mediated treatment,
might be effective but less toxic approache in AML