Acute Myeloid Leukemia

Acute Myeloid Leukemia

- Also known as
Acute myelocytic leukemia
Acute myelogenous leukemia
Acute nonlymphocytic leukemia
Stem cell disorder characterized by Clonal expansion of myeloid
precursor cells with reduced capacity to differentiate i.e,
MATURATION ARREST.

Common myeloid progenitor

Predisposing factors
Congenital factors : Down syndrome.
Bloom syndrome.
Monosomy 7 syndrome
Klinefelter syndrome
Turner syndrome
Neurofibromatosis
Congential dysmorphic syndrome

Predisposing factors
Marrow failure syndrome:

Fanconi anemia
Dyskeratosis congenita.
Schwchman Diamond Syndrome
Amegakaryocytic thrombocytopenia
Blackfan Diamond syndrome
Kostmann Agranulocytosis
Familial aplastic anemia
Familial platelet disorder.

Predisposing factors

Environmental factors:Solvents ( benzene )

Smoking
Ionizing radiation
Non ionizing radiation
Chemotherapy
Alkylating agent
Topoisomerase II inhibitor

Biological features
Leukemogenesis- result from block of differentiation as
well as altered proliferation and impaired apoptosis
through genetic dysregulation.

Genetic Associations
Research states that AML is caused by genetic
aberrations such as translocations between
chromosomes that alter the function of transcriptory
regulatory factors
These translocations are a direct result of chimeric
fusion proteins which are caused by the abnormal cells
and its inability to allow further growth, proliferation,
maturation and differentiation.
Class 1 and 2: mutations responsible for the
development of the neoplastic process of
myeloproliferation and de-differentiation

Genetic Associations
Class 1: mutations that give rise to proliferation and/or
differentiation.
Class 2: mutations that interfere with terminal
differentiation and apoptosis thereby providing survival
advantage for the mutated cells.

Classification of AML
WHO classification.
FAB classification.

Differences between FAB and WHO

FAB-classification:
1)Heavily used Morphologic Findings
2)Special staining (SBB, MPO, NSE, etc), if required

WHO-classification:
1)Morphologic findings
2)Special staining (decreased role)
3)Immunophenotyping (in the form of FC and IHC) heavil
4)Cytogentics and Molecular genetics studies frequently

WHO Classification of Acute Myelocytic

Leukemias

FAB Classification of AML

M0 undifferentiated acute myeloblastic leukemia (5%)

M1 AML with minimal maturation (20%)
M2 AML with maturation (30%)
t(8;21)

M3 Acute promyelocytic leukemia (5%)

t(15;17)

M4 Acute myelomonocytic leukemia (20%)

M4 eos Acute myelomonocytic leukemia with eosinophilia (5%)
inv (16)

M5 Acute monocytic leukemia (10%)

t(9;11)

M6 Acute erythroid leukemia (3%) (DiGuglielmo's disease)

M7 Acute megakaryoblastic leukemia (3%)

Clinical features
Due to Bone Marrow Failure
Due to Organ Infilteration
Others

History from patient with leukemia

Increasing fatigue or decreased exercise tolerance
(anemia)
Excess bleeding or bleeding from unusual sites (DIC,
thrombocytopenia)
Fevers or recurrent infections (granulocytopenia)
Headache, vision changes, nonfocal neurologic
abnormalities (CNS leukemia or bleed)
Early satiety (splenomegaly)
Family history of AML (Fanconi, Bloom or Kostmann
syndromes or ataxia telangiectasia)
History of cancer (exposure to alkylating agents,
radiation, topoisomerase II inhibitors)
Occupational exposures (radiation, benzene,
petroleum products, paint, smoking, pesticides)

Physical Examination
Ecchymosis and oozing from IV sites (DIC, possible
acute promyelocytic leukemia)
Fever and tachycardia (signs of infection)
Papilledema, retinal infiltrates, cranial nerve abnormalities
(CNS leukemia)
Poor dentition, dental abscesses
Gum hypertrophy (leukemic infiltration)(M4)
Skin infiltration or nodules (leukemia infiltration)(M4)
Lymphadenopathy, splenomegaly, hepatosplenomegaly
Back pain, lower extremity weakness [spinal granulocytic
sarcoma, most likely in t(8;21) patients]

Acute Myeloid Leukemia:

Diagnostic Steps
1. Evaluation of an abnormal CBC for possible AML

Confirm bone marrow failure, assess for blasts/blast equivalents and dysplasia
WBC: non-specific; in AML can be low, normal, or high
ANC: severe neutropenia characteristic of HP failure; typical in AML, but exceptions
occur
Circulating blasts: variable number and percent in AML, but key feature to assess
in blood
RBC features: severe anemia characteristic of HP failure, an expected feature of AML
Polychromasia: reduced, since anemia is result of bone marrow production failure
Other RBC pathology: non-specific
Platelets: severe thrombocytopenia characteristic of HP failure

2. Identify morphologic blasts and blast equivalents in blood (and

subsequent bone marrow, if performed)
Morphologic assessment of nuclear features is key for blast
designation
A cell can be a blast if it is exhibiting finely dispersed rather than
condensed nuclear chromatin. Other useful nuclear features include
overall size, nucleoli and nuclear configuration.

Cytoplasmic features are very helpful in lineage determination, ie,

sparse fine granules and Auer rods in myeloblasts, cytoplasmic
blebbing in megakaryoblasts, and deeply basophilic, vacuolated
cytoplasm in erythroblasts

Myeloblast
Relatively high nuclear / cytoplasmic ratio.
Finely dispersed chromatin and variably
prominent nucleoli
Variable number of cytoplasmic granules,
may be concentrated in limited portion of
cytoplasm

Promyelocyte
Nuclear chromatin slightly condensed;
Nucleoli variably prominent;
Nucleus often eccentric, and Golgi zone
may be apparent
Numerous cytoplasmic granules that may
be more dispersed throughout cytoplasm
Blast equivalent in APL only
In APL, intense cytoplasmic granularity
usually present .Nuclear configuration
variable, but nuclear folding and lobulation
characteristic of microgranular variant of
APL

Monoblast

Moderate to low nuclear to

cytoplasmic ratio,
Nuclear chromatin finely dispersed
with variably prominent nucleoli;
nuclei round to folded
Abundant, slightly basophilic
cytoplasm containing fine granulation
and occasional vacuoles

Promonocyte

Slightly condensed nuclear chromatin;

Variably prominent nucleoli
Abundant finely granular blue/gray
cytoplasm that may be vacuolated
Very monocytic appearance with nuclear
immaturity
Consistent blast equivalent in AML

Erythroblast

Relatively high nuclear/cytoplasmic ratio

Nucleus round with slightly condensed
chromatin;
Nucleoli variably prominent
Moderate amounts of deeply basophilic
cytoplasm that may be vacuolated
Included in blast percentage only in acute
erythroid leukemia

Megakaryoblast
Highly variable morphologic features;
May be lymphoid-appearing with high
nuclear to cytoplasmic ratio
Nuclear chromatin fine to variably
condensed
Cytoplasm may be scant to moderate, is
usually agranular or contains a few
granules;
Blebbing or budding of cytoplasm may be
evident
Blasts may form cohesive clumps

Lymphoblast
Morphology
Coarse

Myeloblast

Nucleoli

1-2

3-5

N:C ratio

High

High

Auer rod

Absent

Present

Accompanying
cells

Lymphocytes

Myeloid
precursor

Nuclear
chromatin

Fine

3. Bone marrow examination often performed to address differential

diagnoses from blood assessment or for protocol requirements.
4. Enumeration of blasts/blast equivalents by morphology and differential
cell count

Previously >30% blasts on BM aspirate (per FAB criteria)

As per recent WHO criteria, AML is defined by greater than 20% blasts on BM
aspirate.
patients with certain cytogenetic abnormalities are considered to have
AML regardless of blast percentage
t(8;21)(q22;q22), inversion (16)(p13q22)
t(16;16)(p13;q22), and t(15;17)(q22;q12)

Unique situations compromising blast count:

Fibrosis and/or necrosis
Predominance (50%) of erythroid lineage
Marked hypocellularity
Technically poor specimen

5. Determine lineage of blasts/blast equivalents (can be performed on

blood or bone marrow)
Morphology (nucleus and cytoplasm)
Cytochemistry
Immunophenotype

Myeloperoxidase stain
Basis- breakdown of hydrogen peroxide by enzyme
myeloperoxidase releasing an oxygen radical that reacts
with a soluble substrate to form colored precipitate.

MPO located in peroxisomes of neutrophils and

monocytes and specific granules of eosinophils.
Staining is more pronounced in golgi region.

MPO positive

Sudan Black B
It is a direct stain phospholipid in granular membrane.
Auer rods are MPO and SBB positive.

Esterase stains
Non specific esterase reactivity is found in monocytes.
Basis- Enzymatic release of a side chain from a naphthol
ring with subsequent reaction of the free ring with a
soluble colour develops to generate a coloured
precipitate.
Most common used substrate for Non specific esterase
are Alpha- naphthyl butyrate and Alpha naphthyl
acetate.

NSE positive in MONOCYTIC CELLS

PAS staining
Periodic Acid Schiff stain reacts primarily with glycogen,
generating a fuchsian coloured precipitate.

PAS+

MEGAKARYOCYTES

Cytochemical staining for myeloperoxidase is important in establishing

the lineage of myeloblasts

Immunophenotyping
FAB

Immunological marker

AML with minimally differentiated

CD13,CD34, HLA-DR,
CD33,CD117,CD2,CD7,TdT

AML without maturation

CD13,CD14,CD33, CD34

AML with maturation and with

t(8;21)

CD34,CD56

Acute promyelocytic leukemia

CD13,CD33, HLA-DR absent, CD34

negative

Acute myelomonocytic leukemia

with abnormal eosinophils and
inversion 16

CD13,CD34,CD11b,CD11c,CD14,CD33

Acute monocytic leukemia and

11q23 abnormalties

CD14,CD4,CD36,CD64

Erythroleukemia

Glycophorin 7, Transferrin receptor

CD71

Acute Megakaryocytic leukemia

cCD41,cCD42b,cCD61

Minimally Differentiated Acute

Myeloid Leukemia
5% of AML cases
No definite evidence of myeloid differentiation can be given
by morphology & cytochemistry.
CRITERIA FOR DIAGNOSIS
<3% of blast which are MPO/SBB+(evident on EM)
>20 % of leukemia cells expressing myeloid antigens.

Morphologically undifferentiated
blasts with distinct nucleoli are
peroxidase-negative and do not
show the esterase reaction typical of
monocytes

Bone marrow smear from the same

patient. Immunocytochemical detection
of CD13. A large proportion of the
blasts are positive (red).

AML without maturation

10 20% of AML cases
CRITERIA FOR DIAGNOSIS
Predominance of myeloblast ( > 90% ) without evidence of
maturation ( < 10% promyelocytes or others) in marrow .
IF no auer rods , at least 3% of blast must be MPO OR
SBB positive .
Median age : 45-50 yrs.
Generally chemosensitive and prognostically favourable
unless hyperleukocytosis or complex karyotype present.

BLOOD SMEAR

LARGE & SMALL BLAST WITH

NORMAL PLATELETS

BONE MARROW SMEAR

BLAST WITH PALE TO BASOPHILIC AGRANULAR

CYTOPLASM , NUCLEI WITH FINE CHROMATIN &
PROMINENT NUCLEOLI

MYELOBLAST TYPES

TYPE 1
NO cytoplasmic granules

TYPE 2
15 20 CYTOPLASMIC
GRANULES

 TYPE 3
>20 CYTOPLASMIC
GRANULES

AML with maturation and with t(8;21)

30-45% of AML cases (Most frequent).
Genes involved in t(8;21) are AML1 at 21q22 and ETO
(eight twenty one) at 8q22.
CRITERIA FOR DIAGNOSIS
Blast 20% or more(20-89) of all nucleated cells in bone
marrow
Mature cells (promyelocytes to granulocytes) > 10%
Monocytic cells < 20%.

BONE MARROW SMEAR

BLAST WITH NUCLEOLI AUER RODS

Acute promyelocytic leukemia

Median age 30-38 yrs (young patient).
It is generally not preceded by myelodysplastic
syndrome.
Most patient present with hemorrhagic manifestation
secondary to DIC.
Associated with t(15;17).
Retinoic acid receptor (RAR- alpha) gene on
chromosome 17q12.
Promyelocytic gene (PML gene) on chromosome 15q22.

AML and disseminated intravascular

coagulation (DIC).
Extensive purpura is present on the soles
of a patient with acute promyelocytic
leukemia and DIC

Acute promyelocytic leukemia

Either HYPERGRANULAR OR MICROGRANULAR.

Hypergranular type is most common.
Leukopenia is seen in Hypergranular APL.
Leukocytosis in Microgranular APL.

HYPERGRANULAR

BONE MARROW
SMEAR

Nucleus : Folded, lobulated, granular obscure border.

Cytoplasm: Prominent Azurophilic granules.
Auer rods: Frequent, FAGGOT cells ( cells with bundles of auer rods)

Variations on the appearance of faggot cells in several different cases of APL.

BONE MARROW BIOPSY

MICROGRANULAR

Nucleus : Irregular, Folded. Mostly binucleated.

Cytoplasm : Fine small granules, Dusky appearance.
Auer rods: Rare.

ACUTE MYELOMONOCYTIC LEUKEMIA with ABNORMAL

EOSINOPHILS and INVERSION Of Chrosome 16

15-25% of AML cases

CRITERIA FOR DIAGNOSIS
Blast >20%
Monocytic cells & their precursor
>20%
Neutrophil & their precursor
Median age : 40 45 yrs.
Leukocytosis is present in most of the patients.
Prognosis is better than M1, M2, or M3.

Peripheral smear

myeloblast

PROMONOCYTE

MYELOCYTE

BONE MARROW SMEAR

MONOCYTES & NEUTROPHIL AT VARIOUS STAGES OF MATURATION

M4Eo

EOSINOPHIL

Immature eosinophils have a monocytoid nucleus and a mixture of eosinophilic

and large atypical basophilic granules. M4Eo(CBFb/MYH11)

Acute monocytic leukemia & 11q23

abnormalities
Two types : M5a and M5b
M5a :Poorly differentiated
Trisomy 8 is most common abnormality seen.
M5b :Well differentiated.
FLT3 mutation is most common abnormality seen.
Extramedullary disease occur in > 50% of the patient.
It has a very poor prognosis , 6 -12 months

Gingival Hyperplasia
Leukemia cutis most commonly occurs in
monocytic forms of AML and represents
skin infiltration by leukemic blast cells

M5a( Acute Monoblastic Leukemia )

BLOOD SMEAR

BONE MARROW SMEAR

MONOBLAST

80% or more are MONOBLAST

Abundant cytoplasm
Round nuclei with nucleoli

MONOBLAST WITH ABUNDANT CYTOPLASM

WITH FINE GRANULES

M5b( Monocytic Leukemia )

BLOOD SMEAR
PROMONOCYTES

<80% Monoblast
Mature monocytes or
promonocytes predominate

BONE MARROW SMEAR

ACUTE ERYTHROID LEUKEMIA

M6a (ERYTHROLEUKEMIA)
5% of AML cases
More COMMON THAN pure erythroid leukemia.
Bimodal distribution- <20 yrs and >60yrs.

CRITERIA FOR DIAGNOSIS

>50% of nucleated marrow cells are erythroid lineage
>20% of nonerythroid cells are myeloblast
Dyserythropoiesis is prominent

BLOOD SMEAR

ERYTHROID PRECURSOR

BONE MARROW SMEAR

ACUTE ERYTHROID LEUKEMIA

M6b (PURE ERYTHROID LEUKEMIA )
Very rare
Also called ERYTHEMIC MYELOSIS ,
ACUTE Di GUGLIELMO SYNDROME
>80% of marrow cells are erythroblast
No significant myeloblastic component

Bone marrow smear

Abnormal erythroid

ACUTE MEGAKARYOBLASTIC
LEUKEMIA
10% of AML in children & 5% of adult AML
Bimodal distribution- Infancy and elderly
CRITERIA FOR DIAGNOSIS
Megakaryoblast 20% or more in BM
Bone marrow fibrosis
Megakaryoblast are either small to round with scanty
cytoplasm & coarse chromatin (resembling
lymphoblasts) or medium to large with fine chromatin &
prominent 1-3 nucleoli

ACUTE MEGAKARYOBLASTIC
LEUKEMIA
Morphologically confused with
- L2 subtype of ALL
- AML M1.
Diagnosis depends on expression of at least one platelet
antigen ( i.e., CD41,CD42b, CD61 or factor VIII related
antigen)
Most common leukemia seen in Downs Syndrome.
Platelet show impaired aggregation response.
Elevated serum Lactate Dehydrogenase level.

 Blast show distinct cytoplasmic blebs or psedopods

formation
Peripheral blood fragments of megakaryoblast
micromegakaryocytes Or dysplastic large platelets seen

BONE MARROW SMEAR

Promegakarocytes , irregular nuclei , coarse

BONE MARROW BIOPSY

PROLIFERATION OF MEGAKARYOCYTES

AML WITH MULTILINEAGE

DYSPLASIA
Multilineage dysplasia dysplasia present
>50% of cells in 2 or more myeloid cell lines
Occur in elderly
With / without prior h/o MDS
Poor prognosis
Chromosomal abnormalities similar to MDS

An aspirate smear with increased

blasts and a dysplastic mature
erythroid precursor displays
irregular (cookie cutter) nuclear
contours
(arrow).

This aspirate smear shows several

giant hypogranular bands (arrows)
and a dysplastic erythroid
precursor with asymmetric binucleation is situated just below the
centrally located hypogranular band.

An aspirate smear reveals increased

blasts and two dysplastic
micromegakaryocytes (arrows).

AML & MDS therapy related

Different from denovo AML

Characteristic cytogenetic abnormalities
Multilineage dysplasia
Refractoriness to therapy
Short survival
Follow TOPOISOMERASE II INHIBITOR
( myeloid/lymphoid) OR
ALKYLATING AGENTS

ACUTE BASOPHILIC LEUKEMIA

Basophilic differentiation
Blast contain basophilic granules

IMMATURE BASOPHIL PRECURSOR

Positive metachromatic staining with TOLUIDINE BLUE

ACUTE PANMYELOSIS WITH

MYELOFIBROSIS

Very rare type.

Median age 57 to 67 yrs.
Pancytopenia with < 5% blast.
No history of preceding myeloproliferative disorder.
Proliferation of all major myeloid cell lines
Dyspalstic changes are present along with fibrosis of bone
marrow

Bone marrow biopsy

Relacement by blast & fibrous tissue

MYELOID (granulocytic) SARCOMA

(Myeloblastoma)
Isolated tumour mass.
Also known as Chloroma because some appear green or
turn green in dilute acid secondary to expression of MPO.
Composed of myeloblast or immature cells in
extramedullary site
Sign of relapse in a treated case of AML
Common sites orbits and the paranasal sinuses.
The diagnosis should be suspected if eosinophilic
myelocytes are present in H & E stained biopsy sections.

Differential diagnosis

1. Leukaemoid reaction
2. Myelodysplastic Syndrome
3. Acute Lymphoblastic Leukemia
4. Blast crisis of Chronic Myeloid
Leukemia

LEUKAEMOID REACTION
Refers to the presence of markedly increased leucocyte count
(>50,000/mm3) and immature white blood cells in peripheral
blood resembling leukemia but occurring in non-leukaemic
conditions.
Causes of leukaemoid reaction Severe bacterial or viral infection.
Severe acute haemolysis.
Severe haemorrhage
Cancer metastatic to bone marrow.
Tuberculosis

LEUKAEMOID REACTION
Differentiation from AML is made by following features:

Clinical presentation.
Presence of underlying disease.
Morphology on blood smear.
% of blasts in bone marrow.
Correction of leukaemoid blood picture after treatment of
underlying disease.

Myelodysplastic syndrome
Differentiation of AML from MDS depends on proportion
of myeloblasts in the bone marrow.
In AML, myeloblasts are greater than 20%.
In MDS, myeloblasts are less than 20%.
MPO staining may also be useful for diagnosis of MDS
wherein granulocytes may lose MPO reactivity.

ALL Vs AML
ALL

AML

Age

Mainly children

Mainly adults

Lymphadenopathy

Usually present

Usually absent

Gum hypertrophy

-ve

+ve in M4/M5

Skin infiltration

-ve

+ve in M4/M5

Granulocytic
sarcoma

-ve

+ve in few cases

Mediastinal mass

+ve in T-ALL

Associated DIC

-ve

+ve in M3

Blast crisis of CML

Presence of marked splenomegaly, basophilia and
Philadelphia chromosome are suggestive of CML .
These features differentiate blast crisis of CML with AML.

Prognosis
Factors

Favourble

Unfavourable

Age

< 45 yrs

< 2yrs , > 60 yrs

Leukemia

De novo

-Antecedent
hematological disorder
-Myelodysplastic disorder
-- Myeloproliferative
disorder

Infection

Absent

Present

Prior chemotherapy

No

Yes

Leukocytosis

< 25,000 / mm3

> 100,000 /mm3

Serum LDH

Normal

Elevated

Extramedullary disease

Absent

Present

CNS disease

Absent

present

Clinical factors

Prognosis
Factors

Favourble

Unfavourable

Auer rods

Present

Absent

Eosinophils

Present

Absent

Megaloblastic erythroid

Absent

Present

Dysplastic
megakaryocytes

Absent

Present

FAB type

M2,M3,M4

M0,M6,M7

Morphology

Prognosis
Factors

Favourble

Unfavourable

Myeloid

CD34 ve , CD 14 ve,
CD 13 -ve

CD 34 +ve

HLA- DR

Negative

Positive

TdT

Absent

Present

Lymphoid

CD 2 +ve

CD 7 +ve , CD 56 +ve,
Biphenotypic

Multidrug resistance
gene

Absent

Present

Cytogenetics

t (15;17), t (8;21),
inv (16)

Monosomy 7, del (7q)

Monosomy 5, del (5q),
Complex karyotype

Surface / Enzyme
Marker

THANK YOU

Presented by Dr. Monika Nema