AML with Recurrent Genetic Abnormalities

Question 1

What specific gene mutations are a category

in the WHO for AML ?

Answer 1

• mutated NMP1
• biallelic CEBPA mutation
• mutated RUNX1 (provisional entity)

* Note: there are other mutations, FLT3-ITD and KIT mutations are common and are prognostic.

Question 2

What is the definition of

AML with t(8;21)

RUNX1-RUNX1T1?

Answer 2

• defined by the fusion
• shows predominantly neutrophilic maturation
• large myeloblasts with basophilic cytoplasm and azurophilic granules
• high rates of complete remission and favorable long-term outcome

Question 3

What is the epidemiology and the

clinical features of AML with t(8;21) ?

Answer 3

• about 1-5% of all AMLs
• can present with a myeloid sarcoma
  
  • if that is the case then the bone marrow blasts may be low in number

Question 4

What are some key morphologic

features of AML with t(8;21) ?

Answer 4

• large blasts with deep basophilic cytoplasm and numerous azurophilic granules and perinuclear hofs
  
  • granules may be pseudo-Chediak Higashi like
  • may also see some smaller blasts predominantly in the peripheral blood
• single, long tapered Auer rods
  
  • may be seen in neutrophils as well
• Neutrophils with variable dysplasia
  
  • hyposegmentation
  • abnormal cytoplasmic granulation ( homogeneous pink)
• IMP:
  
  • dysplasia in other cell lines is uncommon

Question 5

What is common about the other

cell lines in AML with t(8;21) ?

Answer 5

• no significant dysplasia of the erythroid or megakaryocytes
• a monocytic component is usually minimal or absent
• eosinophil precursors are frequently increased but different from the inv(16) ones
• basophils and mast cells can be increased
  
  • concurrent mastocytosis is seen in some cases

Question 6

What is the immunophenotype of

AML with t(8;21) ?

Answer 6

• high intensity expression of CD34, HLA-DR, MPO and CD13
• relatively weak expression of CD33
• neutrophilic differentiation can be seen:
  
  • positive for CD15 and/or CD65
• maturation asynchrony can sometimes be seen:
  
  • coexpression of CD34 and CD15
• Other Key Markers positive:
  
  • CD19, Pax-5 and CD79a
  • TdT- weak expression
  • CD56 (sometimes)- adverse prognostic significance
    
    • may be due to KIT mutations in these cases

Question 7

What is the genetic profile of

AML with t(8;21) ?

Answer 7

• core binding factor genes involved
  
  • essential for hematopoiesis
• 70% of cases show additional chromosome abnormalities and mutations
  
  • loss of sex chromosome or del 9q are most common
  • KIT mutations occur in 20-30%
  • KRAS and NRAS occur in 30% of pediatric and 10-20% adult cases

Question 8

What is the prognosis of AML

with t(8;21) ?

Answer 8

• associated with high rate of remission when treated with high-dose Cytarabine
  
  • and long-term disease free survival
• Adverse prognostic factors
  
  • KIT mutations
  • CD56 expression

Question 9

What is the definition of AML

with inv(16) or t(16;16)

CBFB-MYH11 ?

Answer 9

• shows monocytic and granulocytic differentiation
• abnormal eosinophil component

Question 10

What are the epidemiology and

clinical features of AML

with inv(16) ?

Answer 10

• represents 5-8% of younger patients with AML
  
  • lower frequency in adults
• myeloid sarcoma may be initial or relapse presentation
• significantly higher WBC count

Question 11

What are the microscopic findings

of AML with inv(16) ?

Answer 11

• usually AML with monocytic features
• Abnormal eosinophils
  
  • without significant maturation arrest
  • promyelocyte and myelocyte stage with large, basophilic granules (EOS-BASO)
  • mature eosinophils sometimes show hyposegmentation
  • sometimes can be difficult to find
• Auer rods
• Neutrophils are decreased

Question 12

What is an unusual cytochemistry finding

in AML with inv(16) ?

Answer 12

• napthol AS-D chloracetate esterase
  
  • normally negative in normal eosinophils
  • faintly positive in the abnormal eosinophils in this AML

Question 13

What is the immunophenotype of

AML with inv(16) ?

Answer 13

• complex immunophenotype with multiple blast populations
• Immature blasts
  
  • CD34 and CD117 high expression
• Granulocytic lineage
  
  • CD13, CD33, CD15, CD65 and MPO
• Monocytic lineage
  
  • CD14, CD4, CD11b, CD11c, CD64, CD36 and lysozyme
• Other markers often positive
  
  • CD2
  • non-specific

Question 14

What is the genetic profile of

AML with inv(16) ?

Answer 14

• inv(16) is the most common and t(16;16) is rarely seen
  
  • both lead to a fusion of CBFB to MYH11
• conventional cytogenetics
  
  • inv(16) is very subtle
  • FISH and RT-PCR may be necessary
    
    • FISH:
      
      • fusion is normal
      • break apart shows the split CBFB

Question 15

What are frequent secondar genetic

alterations in AML with inv(16)?

Answer 15

• 40% of cases have secondary genetic alterations (most common below)
  
  • gains of chromsome 22 and 8 (10-15% of cases)
    
    • IMP: Trisomy 22 is fairly specific for inv(16)
  • gain of chromsome 21 and del 7q
• secondary gene mutations are very common
  
  • KIT, NRAS, KRAS, FLT3
  • ASXL2 (common in t(8;21)) but not common in inv(16)

Question 16

What are the prognostic and predictive factors

in AML with inv(16) ?

Answer 16

• high rate of complete remission and favorable overall survival
  
  • when treated with high dose cytarabine
• KIT mutations (especially exon 8) have higher risk of relapse and worse survival
  
  • but still not as significant as AML with t(8;21)
• Gain of chromosome 22
  
  • frequently seen associated with an improved outcome
• Worse outcome:
  
  • older age, elevated WBC
  • FLT3 mutations (esp. TKD)
  • trisomy 8

Question 17

What is the definition of AML

with t(15;17) ?

Answer 17

• PML-RARA
• abnormal promyelocytes predominate
  
  • hypergranular (typical)
  • hypogranular (or microgranular)

Question 18

What is the epidmiology and clinical features

of APL ?

Answer 18

• accounts for 5-8% of all AML
• often in younger patients with lower frequency in elderly patients
• often associated with DIC and increased fibrinolysis
• microgranular
  
  • very high WBC which is different from the typical kind

Question 19

What are the microscopic findings of

the hypergranular (classic) type of APL ?

Answer 19

• abnormal promyelocytes are kidney shaped or bilobed
• cytoplasm has densely packed granules and auer rods (bright pink or purple)
  
  • myeloblasts with single Auer rods may also be seen
  • auer rods are larger in APL compared to other AMLs and have a periodicity of 250 nm compared to 6-20 for the others
• hypergranular APL
  
  • usually presents with very low WBC

Question 20

What are the microscopic findings of

the hypogranular variant of APL ?

Answer 20

• generally presents with a high WBC
• promyelocytes have a bilobed nucleus and a paucity of cytoplasmic granules
  
  • can mimic a monocytic leukemia
  • rare Auer rods can be found
  • at relapse (after treatment with tretinoin) the cytoplasm can be deeply basophilic
• bone marrow is usually hypercellular

Question 21

What are the cytochemical findings in APL ?

Answer 21

• MPO is strongly positive in both the hypergranular and microgranular variants
  
  • helpful in differentiating from monocytes
• Non-specific esterase
  
  • weakly positive in ~25% of cases

Question 22

What is the characteristic immunophenotype

of APL ?

Answer 22

• Hypergranular variant
• low or absent : CD34, HLA-DR
• positive: CD11a, CD11b, CD117 (can be weak), bright CD33, CD64, and heterogeneous CD13
• negative for granulocytic markers: CD15 and CD65
• Microgranular variant
  
  • some CD34 positivity and CD2 positive
  • CD11c also positive
• CD11b and CD11c can be upregulated after treatment
• CD2 expression
  
  • associated with FLT3-ITD
• CD56 - positive in 10% of cases
  
  • associated with worse outcome

Question 23

What is the genetic profile of

APL ?

Answer 23

• RARA gene (chromosome 17) fuses with PML (chromsome 15)
• complex variant translocations are also seen, must know
  
  • can have submicroscopic insertions of the PML-RARA gene into different chromsomes
    
    • considered cryptic or masked genes
• Secondary cytogenetic alterations (40% of cases)
  
  • gain of chromsome 8 most frequent
  • FLT3-ITD and FLT3-TKD (30-40% of cases)
    
    • more often microgranular variant (high WBC count)
    • involve the bcr3 breakpoint of PML

Question 24

What are the key variant translocations

seen in APL ?

Answer 24

• Variant partners
  
  • ZBTB16 (previously called PLZF) - 11q23.2
  • NUMA1 - 11q13.4
  • NPM1- 5q35.1
  • STAT5B - 17q21.2
• in general most of the variants have APL morphology, with some exceptions

Question 25

What is the morphology of variant APL

t(11;17) ZBTB16-RARA ?

Answer 25

• predominance of cells with regular nuclei and many granules
• usually an absence of Auer rods
• increased number of plegeroid neutrophils
• strong MPO activity

Question 26

Which variant APL translocations are

resistant to ATRA ?

Answer 26

• ZBTB16-RARA
• STAT5B-RARA

Question 27

What are the prognostic and predictive

factors of APL ?

Answer 27

• very sensitive to ATRA and arsenic trioxide
  
  • allows the cells to differentiate
• best prognosis of all AML subtypes if treated appropriately
• anthracycline is added for high risk patients

Question 28

What is the definition, epidemiology

and clinical features of AML t(9;11)

KMT2A-MLLT3 ?

Answer 28

• usually associated with monocytic features
• IMP:
  
  • more common in children
  • only 2% of adult AMLs
• can present with DIC or myeloid sarcoma

Question 29

What is the immunophenotype if

AML with t(9;11) ?

Answer 29

• in children:
  
  • positive: CD33, CD65, CD4 and HLA-DR
  • low expression of: CD13, CD34, and CD14
• in adults:
  
  • positive: monocytic markers
    
    • CD14, CD4, CD11b, CD11c, CD64, CD36 and lysozyme
  • variable to no expression of markers indicative of immaturity
    
    • CD34, CD117, and CD56

Question 30

What is the genetic profile of

AML with t(9;11) ?

Answer 30

• KMT2A protein
  
  • histone methyltransferase that assembles protein complexes
• most common is t(9;11)
• secondary cytogenetic abnormalities are frequent
  
  • most common: gain of chromsome 8
    
    • usually MECOM negative
  • MECOM (aka EVI1) is reported in 40% of cases
    
    • these cases seem to behave differently than MECOM-negative ones

Question 31

What are the variant KMT2A translocations

in acute leukemia ?

Answer 31

• there are >120 translocations involving KMT2A
• review this section p. 137
• some KMT2A translocations are subtle
  
  • may need FISH or other molecular studies to identify the variants
• MECOM overexpression is frequently seen

Question 32

What are the prognostic and predictive factors

for AML with t(9;11) ?

Answer 32

• AML with this translocation has an intermediate survival
  
  • better than the other KMT2A translocations
• MECOM
  
  • if present portends a poor prognosis

Question 33

What is the definition of AML with

t(6;9) DEK-NUP214 ?

Answer 33

• AML with >20% blasts
• generally has monocytic features
• basophilia and multilineage dysplasia are frequent

Question 34

What is the epidemiology and clinical features

of AML with t(6;9) DEK-NUP214 ?

Answer 34

• translocation is seen in 0.7-1.8% of AMLs
• seen in both kids (median age 14) and adults (35-44 years)
• usually present with:
  
  • anemia and thrombocytopenia
  • can present with pancytopenia
  • IMP: generally the WBC is lower than most AMLs and may be barely above normal

Question 35

What are the microscopic findings of

AML with t(6;9) ?

Answer 35

• blasts typically have features of other AMLs (except for APL and megakaryoblastic)
  
  • generally myelomonocytic and with maturation
• Auer rods (~1/3 of cases)
• marrow and peripheral blood basophilia ~2% (up to 60% of cases)
• granulocytic and erythroid dysplasia
• ring sideroblasts in some cases

Question 36

What are the immunophenotypic findings

of AML with t(6;9) ?

Answer 36

• non-specific myeloid phenotype
  
  • MPO, CD9, CD13, CD33, CD38, CD123 and HLA-DR
  • most cases also positive for CD117 and CD34 and CD15
• some cases express: CD64 and TdT
• other lymphoid antigens are uncommon
• Basophils
  
  • CD123, CD33 and CD38 +
  • HLA-DR (-)

Question 37

What are the genetic findings

in AML with t(6;9) ?

Answer 37

• t(6;9) creates a fusion protein
  
  • acts as an aberrant transcription factor and alters nuclear transport by binding to soluble factors
  • the sole clonal abnormality in most cases
• FLT3-ITD mutations are very common
  
  • not the TKD mutations

Question 38

What are the prognostic and predictive factors

of AML with t(6;9) ?

Answer 38

• generally a poor prognosis
• increased WBC and increased bone marrow blasts are most predictive of shorter overall survival
• despite the high numbers of FLT3-ITD
  
  • does not appear to alter the OS
  • may benefit from FLT3 inhibitors

Question 39

What is the definition of AML with inv(3) or

t(3;3), GATA2-MECOM ?

Answer 39

• dereguated MECOM (also called EVI1) and GATA2 expression
• requires 20% peripheral blood or bone marrow blasts
• often associated with elevated platelet counts

Question 40

What is the typical morphology of

AML with inv(3) or t(3;3)?

Answer 40

• increased dysplastic megakaryocytes
  
  • unilobed or bilobed
• multinlineage dysplasia

Question 41

What is the epidemiology of AML

with inv(3) or t(3;3) ?

Answer 41

• accounts for only 1-2% of AMLs
• occurs most commonly in adults
• no sex predilection

Question 42

What are the clinical features

associated with AML with inv(3) or t(3;3) ?

Answer 42

• most present with anemia and thrombocytopenia
  
  • 7-22% of cases can present with significant thrombocytosis
• hepatosplenomegaly may be present but lymphadenopathy should not occur

Question 43

What are the morphologic features in the

peripheral blood of AML inv(3) ?

Answer 43

• hypogranular neutrophils
  
  • can have pseudo-pelger huet cells as well
• peripheral blood blasts can also be seen
• RBC anomalies are usually mild
  
  • no teardrop cells
• Giant and hypogranular platelets are common
• bare megakaryocytic nuclei can be seen

Question 44

What are the morpologic features of

blasts in AML inv(3) ?

Answer 44

• they have variable morphologic and cytochemical features
  
  • AML without maturation
  • AML with maturation
  • acute megakaryoblastic leukemia
  • these three morphologies are the most common

Question 45

What are the bone marrow findings for

AML with inv(3) ?

Answer 45

• multilineage dysplasia is frequent with dysplastic megakaryocytes being the most common finding (small, hypolobated forms)
  
  • megas are usually also increased in number
• marrow eosinophils, basophils and mast cells can be increased in number
• fibrosis and cellularity are variable

Question 46

What is the immunophenotype fo the blasts

in AML with inv(3) ?

Answer 46

• Positive:
  
  • CD34, CD33, CD13, CD117 and HLA-DR
    
    • Note: high CD34 expression is more often seen with inv(3) than with t(3;3)
  • CD38
  • CD7 (aberrant)- other lymphoid marker expression uncommon
  • CD41 and CD61 (megakaryocyte markers) can occasionally be seen

Question 47

What is important about the genetic alterations

of AML with inv(3) or t(3;3) ?

Answer 47

• MECOM gene is on chromosome 3q26.2
• the genetic alterations reposition the GATA2 enhancer to activate MECOM expression
• IMP:
  
  • MECOM overexpression is not specific to this AML it can be seen in other AMLs particularly therapy-related disease such as t(3;21) MECOM-RUNX1

Question 48

What other genetic abnormalities can be seen

in AML with inv(3) ?

Answer 48

• secondary karyotypic abnormalities are frequent
• most common is monosomy 7 in 50% of cases
• second most common is del 5q and complex karyotypes
• secondary gene mutations are seen in almost all cases
  
  • mutations activating RAS/receptor tyrosine kinase signalling pathways occur in 98%
    
    • most common in descending order: NRAS, PTPN11, FLT3, KRAS, NF1, CBL, and KIT
  • other common genes that are mutated: RUNX1, GATA2, and SF3B1 (usually associated with GATA2 mutation)

Question 49

How is BCR-ABL1 positive CML with

inv(3) or t(3;3) best classified?

Answer 49

• it is best considered as accelarated or blast phase of CML and not AML
  
  • presence of the inversion or translocation is indicative of progression

Question 50

What are the prognostic and predictive factors

of AML with inv(3) or t(3;3) ?

Answer 50

• aggressive disease with short survival
• regardless of blast percentage:
  
  • complex karyotype and additional monosomy 7 are associated with an even worse prognosis

Question 51

What is the definition of AML with t(1;22)

RBM15-MKL1 ?

Answer 51

• AML that generally shows maturation in the megakaryocytic lineage

Question 52

What is the epidemiology of AML

with t(1;22), RBM15-MKL1 ?

Answer 52

• uncommon abnormality in AML (<1%)
• most common in children with Trisomy 21
• female predominance
• can be congenital

Question 53

What are the clinical features of

AML t(1;22) RBM15-MKL1 ?

Answer 53

• unique AML most often seen in children <3 years old
  
  • most cases occur in the first 6 months of life
• present with:
  
  • anemia and thrombocytopenia
  • moderately elevated WBC
  • organomegaly , particularly hepatosplenomegaly

Question 54

What are the microscopic features of

AML with t(1;22)?

Answer 54

• the peripheral blood and bone marrow blasts are similar to those seen in acute megakaryoblastic leukemia
  
  • small and large megakaryoblasts may be present and they may be admixed with more undifferentiated blast cells that resemble lymphoblasts

Question 55

What is the morphology of the megakaryoblasts

seen in AML with t(1;22) ?

Answer 55

• medium to large in size
• round to slightly irregular and indented nucleus with fine reticular chromatin and 1-3 nucleoli
• cytoplasm is basophilic, often agranular with possible blebs or cytoplasmic projections
• micromegas are common but dysplastic features in the granulocytes and erythroids are not common

Question 56

What are the other bone marrow

findings in AML with t(1;22) ?

Answer 56

• can be normocellular or hypercellular
• often there is dense fibrosis similar to that seen with metastatic tumors
  
  • because of this the aspirate may be less helpful in getting 20% blasts on the differential count

Question 57

What is a key cytochemical staining finding in

the megakaryoblasts of AML with t(1;22) ?

Answer 57

• the megakaryoblasts are negative for Sudan Black B and MPO

Question 58

What is the immunophenotype of the

blasts in AML with t(1;22) ?

Answer 58

• express one or more of the platelet glycoproteins:
  
  • CD41 (GP IIb/IIIa)
  • CD61 (GP IIIa)
  • CD42b (GP Ib)
  • IMP: cytoplasmic staining for CD41 and CD61 are more specific and sensitive for megakaryocytic differentiation
• Positive for:
  
  • CD13 and CD33
  • CD36, not specific
• Negative for:
  
  • CD45, CD34, and HLA-DR
  • MPO, lymphoid markers and TdT

Question 59

What are the genetics of AML

with t(1;22) ?

Answer 59

• should have the translocation or at least evidence of RBMK15-MKL1 gene fusion
  
  • generally this is the sole karyotypic abnormality
  • RBM15
    
    • encodes RNA recognition motifs and split-end (spen) paralogue and orthologue C-terminal domains
  • MKL1
    
    • encodes a DNA-binding motif involved in chromatin organization
  • Fusion gene
    
    • may modulate chromatin organization
    • HOX-induced differentiation and extracellular signalling pathways

Question 60

What are the prognostic and predictive factors

for AML t(1;22)?

Answer 60

• most studies have shown this to be a high risk disease as compared to acute megakaryocytic leukemia without the t(1;22)