HEMA2 Lec 1st Shift Reviewer • The system groups them further into:

Acute Leukemias o AML into 9 subtypes (M0-M7)

o ALL into 3 categories (L1-L3)
Prepared by: C3
• This system is based on the type of cell
from which the leukemia developed and
EPIDEMIOLOGY OF ACUTE the maturity of cells
LEUKEMIAS • Classifications are based largely on how
• According to the National Cancer the leukemia cells look under the
Institute data for the USA in 2009: microscope
o 44,970 new cases of leukemia
o 21,870 deaths from leukemia
• As a group, the acute leukemias are
characterized by the presence of blasts
and immature leukocytes in the
peripheral blood and bone marrow
• Anemia is usually present and caused
by bleeding and the replacement of
normal marrow elements by leukemic
blasts
• Although the total leukocyte count is
usually elevated, some patients may
demonstrate normal or decreased
leukocyte counts
• Thrombocytopenia is also usually
present in patients with acute leukemia

PROGNOSIS OF ACUTE LEUKEMIAS

• Modern treatment methods have
produced a high rate of survival in Figure 1A & 1B. FAB classification of acute
children but the only significant progress leukemias.
in treatment for the last three decades
has been confined mostly to younger Table 1. The FAB Classification System
than the majority of patients who are
older M0 Myeloid Undifferentiated blasts,
AML–not otherwise
• The best time to achieve the longest categorized
remission and possible cure of acute M1 Myeloid Blasts and
leukemia via maximum cell kill is when promyelocytes
the disease is first diagnosed predominate without
further maturation of
myeloid cells
FRENCH-AMERICAN-BRITISH AND M2 Myeloid Myeloid cells
WORLD HEALTH ORGANIZATION demonstrate maturation
beyond the blast and
CATEGORIES promyelocyte stage
• FAB classifies acute leukemias into two M3 Promyelocytic Promyelocytes
major divisions: predominate in the bone
o Acute Myeloid Leukemias marrow
(AMLs) M4 Myelomonocytic Both myeloid and
monocytic cells are
o Acute Lymphoblastic Leukemias present to the extent of
(ALLs) at least 20% of the total
leukocytes

TRIPLE C NOTES 1
M5 Monocytic Most cells are
monocytic; two
subtypes (a and b) are
recognized, one
characterized by large
blasts in bone marrow
and peripheral blood,
the other (differentiated
type) by monoblasts,
promonocytes, and
monocytes
M6 Erythroleukemia Also known as Di
Guglielmo syndrome;
abnormal proliferation of
both erythroid and
granulocytic precursors;
may include abnormal GENERAL CHARACTERISTICS OF
megakaryocytic and ACUTE MYELOID LEUKEMIAS
monocytic proliferations
M7 Megakaryocytic Large and small
megakaryoblasts with a
high nuclear-
cytoplasmic ratio; pale,
agranular cytoplasm
L1 Homogeneous One population of cells
within the case; small
cells predominant;
nuclear shape is regular
with an occasional cleft;
chromatin pattern is
homogeneous and
nucleoli are rarely
visible; cytoplasm is
moderately basophilic
L2 Heterogeneous Large cells with an
irregular nuclear shape;
clefts in the nucleus are
common; one or more
large nucleoli are
visible; cytoplasm varies Genetic Differences
in color
L3 Burkitt Lymphoma Cells are large and
type homogeneous in size,
nuclear shape is round
or oval; one to three
prominent nucleoli;
cytoplasm is deeply
basophilic with vacuoles
often prominent
WORLD HEALTH ORGANIZATION
(WHO) CLASSIFICATION
• The underlying rationale uses all
available information to define clinically
significant disease entities:
o Morphology
o Cytochemistry
o Immunophenotyped
o Genetics
o Clinical features
• Patients generally have a favorable
prognosis in AML characterized by
recurrent chromosome translocation:
o t(5;17)(q22;q12)
o t(8;21)(q22;q2)
o inv16(p13q22)
• leukemias with complex karyotypes,
partial deletions, or loss of chromosome
are frequently characterized by
multilineage dysplasia and poor therapy
• AML is the most common leukemia
subtype
• AML is a genetically heterogeneous
clonal disorder characterized by a
maturation block and accumulation of
acquired somatic generic alterations in
hematopoietic progenitor cells that alter
normal mechanisms of self-renewal,
proliferation, and differentiation
• Classification of AML subtypes is
clinically relevant because particular
abnormalities are associated with
distinct clinical behavior
o Prognosis is favorable or
unfavorable response to
treatment
• Although AML blasts evolve from
common myeloid precursors, the
subtypes differ in terms of the particular
myeloid lineage involved and the degree
of leukemic cell differentiation
• In the movement beyond morphologic
classification of AML to genetic
classification, two major subgroups of
AML have emerged, patients with
disruptions of:
o FLT3 gene, which encodes a
type III receptor tyrosine kinase.
It is known to be mutated in up
to 30% of patients with AML
o Core-binding factor (CBF)
complex, a transcription factor
complex critical for regulation of
TRIPLE C NOTES 2
 hematopoiesis and normal
myeloid development
• The two major subgroups of genetic
disruptions in AML can demonstrate
interaction in initiating and maintaining
the leukemic clone
Micro-RNAs
• Micro-RNA expression is associated
with cytogenetics, molecular, and
morphological alterations, and clinical
outcomes in AML
• Naturally occurring 19- to 25-nucleotide
RNAs cleaved from 70 to 100 nucleotide
precursors that hybridize to
complementary mRNA targets and leads
to their degradation or inhibit their
translation of the corresponding proteins
Acute Leukemias
• Increase in the number of immature
cells in the bone marrow and arrest in
their maturation
• Frequently results in hematopoietic
insufficiency with or without leukocytosis
• Monoclonal antibodies are helpful in the
classification of acute leukemias
Table 2. Differentiation of AML.
M0 Undifferentiated myeloid blasts
M1-M4 Unique characteristic is the possession
of granulocytic differentiation with
varying degrees of maturation
M4 Represents a combination of myeloid
and monocytic leukemias
M5 Designated category for monocytic
leukemias with additional
subcategories
M6 Reserved for erythroleukemias, an
abnormality of both erythrocytic and
myeloid cell lines
M7 Designation for megakaryocytic
leukemias
Table 3. Examples of Monoclonal Antibody
Classification of Acute Leukemias
Myeloid lineage CD13, CD3, CD15,
MPO, CD117
T-Cell lineage CD2, CD3, CD5, CD7
B-Cell lineage CD19, CD20, CD22,
CD79a
Megakaryoblastic CD41, CD61
FAB M0
• WHO synonym: AML not otherwise
categorized (minimally differentiated)
• Patients are classified in this group
based on:
o Morphological features of the
leukemic cells
o Cytochemical features of the
leukemic cells
o Degree of maturation
• AML, minimally differentiated, has no
evidence of myeloid differentiation
• The myeloid nature of blasts is
demonstrated by immunological markers
and/or ultrastructural studies
o Immunophenotyping is essential
• Evidence of bone marrow failure is
characterized by:
o Anemia
o Neutropenia
o Thrombocytopenia
o Leukocytosis with a significantly
increased number of blasts
Figure 2. M0 (Acute Myeloid Leukemia with Minimal
Differentiation). Typically characterized by decreased
platelets and agranular blasts.
Acute Myeloid Leukemia (FAB M1)
• WHO synonym: Acute Myeloblastic
Leukemia Without Maturation
• Most common type of leukemia in
children younger than 18 months of age,
but it typically occurs in middle-aged
adults with a median age of 46 years
• The typical male:female ratio of FAB M1
is 1:1
TRIPLE C NOTES 3
• The median survival time is 3.5 months
after diagnosis
• Clinical signs and symptoms:
o Rapid or gradual onset that may
resemble an acute infection
 History of fever
 Infections
 Fatigue
 Bleeding episodes
o Physical examination may
reveal:
 Tenderness of the
bones especially ribs
and sternum
 Ulcerated mucus
membranes
 Petechiae
 Purpura
o Additional physical findings:
 Hepatomegaly
 Splenomegaly
 Lymphadenopathy
o Approximately 50% of patients
exhibit no organomegaly or
lymphadenopathy
o Cellular infiltration of organs is
less prominent in AML
compared with ALL
o Occasional localized tumor
masses consisting of
myeloblasts may arise in bone
or soft tissues
o Appearance of chloroma tumor
is an early sign of AML
 Large quantities of MPO
produces a green
appearance if the tissue
is cut
• Laboratory Data
o Anemia and thrombocytopenia
are present in approximately
85% of all AMLs
o Leukocytosis is encountered in
more than one third of patients
o Total leukocyte count is usually
greater than 100 x 109/L
o Peripheral blood smear does
not usually exhibit many
immature erythrocytes (more
common in other forms of acute
leukemia)
o If severe disruption of
erythrocyte development does
occur and many immature forms
are present, the leukemia is
possibly an erythroleukemia
(FAB M6)
o Predominance of myeloblasts in
peripheral blood smear and
bone marrow
 Have regular
cytoplasmic outline
 May contain slender,
red-staining Auer rods
in the cytoplasm
 The nuclear chromatin
is fine and
homogeneous
 3-5 nucleoli are usually
evident
o Some promyelocytes may be
present
o Agranular or hypogranular
segmented neutrophils may be
seen (acquired Pelger-Huët
anomaly)
o Abnormal eosinophils may also
be seen
o Monocytes usually constitute
less than 1% of the nucleated
cells in the peripheral blood
Figure 3. M1 (Acute Myeloid Leukemia Without
Maturation). Typically characterized as having rare
Auer rods with a predominant cell usually of Type I
myeloblast.
TRIPLE C NOTES 4
Acute Myeloid Leukemia (FAB M2)
• WHO synonym: Acute Myeloblastic
Leukemia With Maturation.
• Typically occurs in middle-aged persons
o Median age: 48 years
o 40% of cases occur in
individuals 60 years or older
• Approximate male:female ratio is 1.6:1
• Median survival time is 8.5 months
• Clinical signs and symptoms:
o Hemorrhagic manifestations
 Easy bruising
 Epistaxis
 Gingival bleeding
 Petechiae
o Hepatomegaly, splenomegaly,
and lymphadenopathy
(infrequently seen)
• Laboratory data:
o Anemia and thrombocytopenia
are present in most cases
o Leukocytosis is commonly seen
 Rare patients having
total leukocyte counts
exceeding 300 x 109/L
o Myeloblasts predominate
peripheral blood smear
 Nuclei are usually round
or oval with one or more
prominent nucleoli and
fine reticular chromatin
 Cytoplasm is basophilic
with a variable number
of azurophilic granules
 Auer rods are
commonly seen
 Maturation of the
granulocytic cell line is
also observed
Figure 4. M2 (Acute Myeloid Leukemia with
Maturation). Auer rods are typically present and Type
II myeloblasts may be the predominant cell.
Acute Promyelocytic Leukemia (FAB M3)
• Cottage-loaf nucleus
• Median age of occurrence is 38 years
o Median survival: 16 months
• The approximate male:female ratio is
2:1
• Since the introduction of all-trans
retinoic acid (ATRA) in the treatment
and optimization of the ATRA-based
regimens, the complete remission rate
has reached greater than 90%, and a 5-
year disease-free survival is greater
than 90%
• Clinical signs and symptoms:
o Fatigue
o Symptoms of bleeding
 Bruising
 Hematuria
 Petechiae
o Hepatomegaly, splenomegaly,
and lymphadenopathy are seen
infrequently
• M3 appears to be the most aggressive
of acute leukemia with a severe
bleeding tendency and a fatal course, if
untreated, of only weeks
• Laboratory data:
o Similar to FAB M2 type
o Anemia and thrombocytopenia
are present in most cases
o Total leukocyte counts range
from conditions of leukopenia
(seen infrequently) to
leukocytosis
TRIPLE C NOTES 5
o Promyelocytes are the o Need to identify the PML/RARα
predominating cell type gene fusion product is essential
 Hypergranular, for the clinical management of
microgranular, or these patients and the potential
hypogranular variations to achieve 3-year-disease-free
 Coarsely granular survival rates
promyelocytes with
dumbbell-shaped or
bilobed nuclei may be
seen
 Nuclear chromatin is
finely reticular and the
cells often lack nucleoli
o Myeloblasts and cells at the
myelocyte level of development
may also be present and
contain many small Auer rods
o Increased incidence of
disseminated intravascular
coagulation (DIC)
o Cytogenetically characterized by Figure 5. M3 (Acute Promyelocytic Leukemia-
a balanced reciprocal Hypergranular). Blasts and promyelocytes show
translocation between heavy granulation and multiple Auer rods. WBC count
chromosomes 15 and 17 is usually decreased (usually <5,000) but the range is
 Results in the fusion 3,000-15,000. Auer rods range from 10 to 20 per cell
between PML gene and (faggot cells) and the rods may be intertwined or
single. Fewer Auer rods are possible.
retinoic acid receptor α
(RARα)
o Microgranular (hypogranular)
variant of acute promyelocytic
leukemia (aPML-M3v)
 20-30% of aPML
 Impossible to diagnose
with modern testing
because the leukemic
cells do not show the
hypergranular
cytoplasm and
dysplastic changes of
the nucleus as seen in
typical cases of aPML
o Tumor cells morphologically Figure 6. M3v (Acute Promyelocytic Leukemia-
resemble monoblasts with few, Microgranular Variant). White blood cells are
markedly increased. Promyelocytes are usually
fine cytoplasmic granules and
bilobed and the cytoplasm contains only a few
bilobed and irregular nuclei granules.
o Only rare cells exhibit Auer rods
o Presence of the PML/RARα Acute Myelomonocytic Leukemia (FAB M4)
gene is essential for the • WHO Synonym: Acute Myelomonocytic
diagnosis of aPML because Leukemia
more than 95% of AML-M3
• Referred to as “Naegeli-type” monocytic
cases have the PML/RARα
leukemia
fusion protein

TRIPLE C NOTES 6
• Uncommon in children and young adults
• The highest frequency of occurrence in
adults is older than 50 years of age
• Average male:female ratio is 1.4:1
• Most forms are of the acute form, with
the average length of survival being
approximately 8 months
• Juvenile Myelomonocytic Leukemia
(JMML) is a rare malignant disease in
children
o Accounts for less than 3%
children of all childhood
hematologic malignancies
o No curative therapy except for
stem cell transplantation
• Clinical signs and symptoms:
o Similar to other forms of acute
leukemia
 Fatigue
 Fever
 Bleeding manifestations
o Pharyngitis
o Hepatomegaly and
splenomegaly are seen in about
one-third of patients
o Gingival hyperplasia due to
leukemic infiltration
o Patients with FAB M4 or FAB
M5 leukemia or ALL
(predominantly of the T-cell
type) with hyperleukocytosis are
at risk of leukostasis
development
 Leukostasis –
pathological finding of
slight dilated, thin-
walled vessels filled
with leukemic cells
 Brain and lungs are
most commonly
involved organs
 Symptoms include
headache, visual
impairment, and
shortness of breath
• Laboratory data:
o Proliferation of granulocytes and
monocytes is characteristic
o Anemia and thrombocytopenia
are present
o Total leukocyte count varies
from leukopenia to leukocytosis
 Rarely exceeds 100 x
109/L
 Absolute monocyte
count reaches or
exceeds 5 x 109/L in the
peripheral blood
o On a peripheral blood smear:
 Early myeloid cells
predominate
 Approximately 20% of
the cellular elements
are monocytes
 Blasts may have
indented and
convoluted nuclei as in
monocytes
 The number of nucleoli
averages from 3 to 5
 Auer rods may be
present
 Promyelocytes are often
present but don’t
predominate
 Agranular and
hypogranular
neutrophils may be
seen
 Acquired pseudo-
Pelger-Huët anomaly
may be noted
 Number of platelets is
usually reduced
 Erythrocytic precursors
are not usually seen
 DIC can be observed
o A variant of FAB M4 is FAB M4
Eo
 Bone marrow aspiration
reveals myeloblasts and
monoblasts along with
abnormal eosinophils
TRIPLE C NOTES 7
 o FAB M5b
 WHO Synonym: Acute
Monocytic Leukemia
 Common in the middle
age
 Median age: 49 years
 Male:female ratio is
1.8:1
• Very resistant to therapy
o Life expectancy is short
o Ranges from 5-8 months
depending on the type
• Clinical signs and symptoms:
Figure 7. M4 (Acute Myelomonocytic Leukemia). o Onset is dramatic with:
WBC count is usually increased. Both myelocytic and  Headaches
monocytic differentiation are found. >5 x 109/L  Fevers
monocytes are precursors are found. Auer rods may o Typical symptoms of monocytic
also be present. leukemia include:
 Fatigue
 Weight loss
 Bleeding from the
mouth and nose
o Physical examination reveals:
 Gingival hyperplasia
 Pallor
 Skin lesions
o Enlargement of the lymph nodes
and spleen is uncommon
o Extramedullary masses may be
seen in about one third of
patients
• Laboratory data:
Figure 8. Bone marrow smear of M4 Eo (Acute o Anemia and thrombocytopenia
Myelomonocytic Leukemia with Increased Bone are usually evident
Marrow Eosinophils). >5% and <30% abnormal
o Total leukocyte ranges from 15-
eosinophils. Atypical eosinophils with monocytic or
100 x 109/L
pseudo-Pelger-Huët features in the nuclei and
abnormal basophilic granules. o Peripheral blood smears exhibit
a high proportion of blast forms
Acute Monocytic Leukemia (FAB M5) o 25-75% of the nucleated cells
• Pure monocytic leukemia is uncommon constitute monocytes and
and comprises less than 15% of all promonocytes
leukemias o Blasts are frequently muddy or
• Two forms: smoggy gray-blue cytoplasm
o FAB M5a containing:
 WHO synonym: Acute  Tiny granules
Monoblastic Leukemia  Pseudopods
 Most common in young o Nucleus has a reticular granular
adults chromatin pattern and may
 Median age: 16 years contain 1-5 large nucleoli
 Male:female ratio is o Few immature erythrocytes may
0.7:1 be seen occasionally

TRIPLE C NOTES 8
 • Laboratory Data:
o Blast cells of erythroid and
myeloid origin are found in both
the bone marrow and peripheral
blood
o Erythroblasts on bleed smears
 Have an irregular
outline
 High N:C ratio
o Some of the blasts exhibit the
intense blue color associated
with rubriblasts
o Blasts of myeloid origin may
Figure 9. M5a (Acute Monoblastic Leukemia). WBCs have Auer rods
are usually increased. Blast morphology is variable. o Promyelocytes may also be
Auer rods are usually absent. present as well as
promonocytes and monocytes

Figure 10. M5b (Acute Monocytic Leukemia).

Monocytosis with the promonocyte as the Figure 11. M6a (Erythroleukemia). Usually exhibits
predominant cell is usually characterized. three phases and there is more myeloid involvement
as the disease progresses. Anisocytosis,
poikilocytosis, basophilic stippling, and nucleated red
Erythroleukemia (FAB M6) blood cells are evident.
• WHO Synonym for FAB M6a & M6b:
Acute Erythroid Leukemia
• Also referred to as “erythremic myelosis”
or “Di Guglielmo syndrome”
o Represents a proliferation of
both immature granulocytic and
erythrocytic cell types
• Usually acute
• Median age of occurrence: 54 years
• Male:female ratio is 1.4:1
• Average length of survival: 11 months
• Clinical signs and symptoms:
o Bleeding manifestation
o Hepatomegaly, splenomegaly,
Figure 12. M6b (Pure Erythroid Leukemia). Platelets
and lymphadenopathy are
are decreased. Usually have macrocytic anemia.
infrequently observed
TRIPLE C NOTES 9
Selected Examples of Unusual Forms
1. Eosinophilic Leukemia
• Rare, although it can be
indistinguishable from reactive
eosinophilia or chronic myeloid
leukemia (CML)
• Usually acute when present
• Death generally occurs within 1
year
• Tissue infiltration and cardiac
failure have been described
• Signs and symptoms include:
o Chronic cough
o Pulmonary infiltration by
leukocytes
o CNS involvement
• 20% if patients demonstrate
anemia and thrombocytopenia
• Leukocytosis with a total
leukocyte counts of 50 to 200 x
109/L may exist
o Few patients have
greater than 100 x 109/L
• On peripheral blood smears:
o More than 60% can be
eosinophils
o Granules may not stain
uniformly
o Some of the granules
may appear empty
• In the terminal phase, 80% of
nucleated cells may constitute
blasts
• Abnormal eosinophils are often
present in small numbers in all
leukemias
2. Basophilic Leukemia
• Mast cell leukemia
• Rarest form of all leukemias
• Frequently, an infiltration of
mast cells in large numbers into
affected skin is observed
• Patients exhibit leukocytosis,
with total leukocyte counts
exceeding 30 x 109/L
• Peripheral blood smears can
demonstrate greater than 50%
basophils in this disorder
Figure 13. M7 (Acute Megakaryoblastic Leukemia).
Variable WBC count but usually decreased. Platelets
are variable, bizarre, and typical.
Acute Megakaryoblastic Leukemia (FAB M7)
• WHO synonym: Acute Megakaryoblastic
Leukemia
• 50% or more of the blasts are of
megakaryocyte lineage
• Occurs in children and adults
• It comprises approximately 3% to 5%
cases of AML
• Clinical signs and symptoms:
o Organomegaly (abnormally
enlarged organs) is infrequent
except in children
o Radiographic evidence of bone
lytic lesions has been observed
in children
• Laboratory data:
o Cytopenia is usually present
 Particularly
thrombocytopenia
o Dysplastic features in the
neutrophils and platelets may be
present
o No unique chromosomal
abnormality associated
o Immunophenotyping reveals
that megakaryoblasts express
one or more of the platelet
glycoprotein:
 CD41
 CD61
o Blasts are negative with anti-
MPO antibody
o Prognosis is usually poor,
particularly in infants
TRIPLE C NOTES 10
EPIDEMIOLOGY OF ACUTE • The WHO classification that is
LYMPHOBLASTIC LEUKEMIA (ALL) synonymous with the FAB L1 and L2
classification is:
• Most common cancer in children
o 23% of cancer among children o Precursor B-Cell Acute
younger than 15 years old Lymphoblastic Leukemia (B-
ALL)
• Occurs in 1/29,000 children in the US
 Neoplasm of
each year
lymphoblasts committed
• Has a bimodal age distribution
to the B-cell lineage
o Peaking in children between 3
o Precursor T-Cell Acute
and 5 years of age
Lymphoblastic Leukemia (T-
o Occurs again in older than 65
ALL)
years
 Neoplasm of
• Pediatric ALL occurs slightly more often
lymphoblasts committed
in boys than in girls and in white children
to the T-cell lineage
more often than in black children
Table 4. Morphological classification and
PROGNOSIS OF ALL characteristics of ALL.
• Treatment in children: greatly improved FAB Size of Nuclear Nucleoli Cytoplasm
Type blasts shape
over the past few years
L1 Small Indistinct Scant Invisible
• Treatment in adults: less progressive L2 Large, Indented, Large, Moderat-
heteroge- promi- abundant ely clefted
neous nent
CLASSIFICATIONS OF ALL L3 Large Regular Promi- Prominent,
• ALL is divided into: to oval nent, vacuoles
round basophilic
o FAB L1 (children)
o L2 (older than children and
adults) CHARACTERISTICS OF ACUTE
o L3 (patients with leukemia LYMPHOBLASTIC LEUKEMIA
secondary to Burkitt lymphoma)
• Classification is based on: Clinical Signs & Symptoms
o Morphology • History of symptoms can vary from a
o Cell size few days to a few weeks
o Prominence of nucleoli • Symptoms can include:
o Amount and appearance of o Fatigue
cytoplasm o Fever
• Well-established events in the process o Infection
of neoplastic transformation are from: o Headache
o Altered regulation of the cell o Nausea
cycle o Vomiting
o Apoptosis (programmed cell o Bone and joint pain related to
death) the replacement of normal
• ALL cell lines and circulating leukemic hematopoietic elements is
cells from pediatric patients possess common
different regulatory mechanisms • Pain in the extremities is produced by
• It is generally accepted that T-cell acute infiltration of leukemic cells into the
lymphoblastic leukemia (T-ALL) results tissues
from the malignant transformation of • Physical examination may reveal:
normal developing T-cells in the thymus o Petechiae
(thymocytes) o Hemorrhage
o Pallor

TRIPLE C NOTES 11
 • GIT hemorrhage and hematuria are less
common findings
• Lymphadenopathy and hepatomegaly
are present in 75% of patients
• Leukemic meningitis and cranial nerve
palsies are common
o Caused by nerve infiltration by
leukemic blasts
o Leukemic cells can infiltrate
many areas of the body
• Nephropathy may be present but is
usually precipitated later by therapy that
lyses the abundant leukocytes
• In the FAB L2 variety, the lymphoblasts
may have indented nuclei and frequently
show mature cells of the myeloid type
• Early forms of erythrocytes and
megakaryocytes are absent in all forms
of this type of leukemia
• Patients may develop meningeal
leukemia following prolonged remission
without evidence of abnormalities
• In adults, ALL is differentiated from
lymphosarcoma by the presence of
poorly differentiated lymphocytes, which
may have prominent nucleoli

Laboratory Data
• Total leukocyte count
o Elevated in 60-70% of patients
o Range from 50-100 x 109/L
o Less than 15% of patients have
extreme leukocytosis with a total
leukocyte count of more than
100 x 109/L
o Leukocytopenia is present in
approximately 25% of patients
• Peripheral blood smears
o Show a predominance of blast
cells in 50% of patients
o Composed of close to 100%: Figure 14. L1 (Precursor Lymphoblastic Leukemia).
 Lymphoblasts Mutation of single lymphoid stem cell causing
 Lymphocytes proliferation of malignant lymphoblasts. Characterized
by normocytic/normochromic anemia. Platelets are
 Smudge cells
decreased.
o Blasts
 1-2 nucleoli in the
nucleus
 Auer rods are absent
from the cytoplasm
 High N:C ratio
 Shape of nucleus is
round rather than
indented or twisted
o Reveal granulocytopenia
 Some immature are
often seen in blood as a
response to leukemic
replacement of the bone
marrow
o Presence of anemia owing to Figure 15. L2 (Precursor Lymphoblastic Leukemia).
Slightly similar to L1.
decreased RBC production
o Blood loss
o Severe thrombocytopenia

TRIPLE C NOTES 12
 Table 5. Immunological markers in ALL.
Type FA Td CAL C CD HL SI
B T LA D7 19 A- g
DR
Precur L1 + 0 0 + + 0
sor B- L2
cell
ALL
Comm L1 + + 0 + + 0
on L2
ALL
Pre-B- L1 + + 0 + + 0
cell L2
ALL
Figure 16. L3 (Burkitt Type). The lymphoblasts are B-cell L3 0 0a 0 + + +
ALL
similar in appearance to those found in Burkitt
T-cell L1 + 0a + 0 0 0
Lymphoma. Constitutes about 3-4% of precursor
ALL L2
lymphoblastic leukemias in children and adults.
Null L1 + 0 0 0 + 0
Cell L2
Special Identification Techniques ALL
• Surface markers are proteins on the cell
membranes that can be detected with
immunologic reagents Treatment
o Extremely helpful in • Differ for Philadelpha (Ph) chromosome
differentiating ALL positive and negative ALL
o Different proteins are expressed • Ph chromosome stems from a reciprocal
at different stages of maturation, translocation between chromosomes 9
which allows them to be used as and 22
markers of both cell lineage and o Generally places upstream
maturation domains from the BCR gene
• Terminal deoxynucleotidyl transferase from chromosome 22 in
(TdT) is an intracellular enzyme that juxtaposition with the
catalyzes the non-specific incorporation downstream tyrosine kinase
of nucleotides into DNA domains of Abl, from
o TdT (+) lymphoblasts are found chromosome 9
in the bone marrow and blood of • First and second generation ABL kinase
the majority of patients with ALL inhibitors have become front-line
of T and B-cell lineage therapy in cases of Ph chromosome-
o Present in most cases of positive ALL
lymphoblast lymphoma and in o Incorporation into the hyper-
about one third of patients with Cytoxan (cyclophosphamide),
CML in blast crisis vincristine, Adriamycin
o Predictor of favorable response (doxorubicin), dexamethasone
to treatment (hyper-CVAD) regimen for adult
• The common ALL antigen (cALLA) is ALL with t(9;22) has improved
found on the surface of lymphoblasta in patient outcomes compared to
70% if patients with ALL patient regimens with imanitib
• CD20 expression is associated with mesylate
inferior survival in adults with ALL

TRIPLE C NOTES 13
MIXED LINEAGE LEUKEMIA
• New drug-resistant form of childhood
leukemia
• Established to be a distinct disease
• Not a subtype of the prevalent ALLs
• Gene profiles of more than 12,000
genes established that about 1,000
genes were underexpressed
o About 20 were expressed at
higher levels in MLL when
compared to ALL
o Distinguishes MLL from classic
ALL
CYTOGENETIC ANALYSIS
Cytogenetic Analysis in Acute Myeloid
Leukemia
• Cytogenetic studies are important
because 2/3 of patients with AML or ALL
and 90% of patients with secondary
leukemia will have leukemic blasts
showing clonal chromosomal
abnormalities
• Chromosomal abnormalities
o Differ between AML and ALL
o Differ among various subtypes
• Immunophenotyping and cytogenetic
analyses assist in risk stratification and
provide information that has important
clinical, prognostic, and treatment
implications
• Patient’s response to therapy and
survival are correlated with the
karyotype and gene rearrangement of
malignant cells
• Diagnosed patients can be monitored for
remission and relapse (minimal residual
disease) with the use of chromosomal
and molecular analysis
• Since the initial observation of the Ph
chromosome, a number of recurring
chromosome abnormalities have been
described in human leukemias and
lymphomas, including AML, ALL, and
non-Hodgkin lymphomas
• Such chromosome abnormalities
include:
o Gains and losses of entire
chromosomes
o Deletions
o Translocations
o Inversions
• Analysis of chromosome morphology
(karyotyping) and specific band patterns
of individual chromosomes are useful in
terms of identifying the type of acute
leukemia
• The most consistent and specific
chromosomal abnormalities found in
human leukemia cells are
translocations
• Abnormal gene rearrangements result
from chromosomal abnormalities that
are usually the consequence of
chromosomal translocations
o Involve cellular oncogenes
• Activation of normal cellular oncogenes
by translocation-induced gene
rearrangement are an important part of
the process of malignant transformation
• Cytogenetic analysis of leukemic blasts
has resulted in identification on non-
random clonal chromosomal aberration
in a large number of patients with AML
o Some of these lesions are
correlated with specific FAB
subtypes
o Importance of cytogenetic
lesions as powerful
determinants to the therapeutic
response – suggests that the
mechanisms of transformation
associated with these lesions
are likely to directly influence
the sensitivity of the leukemic
blasts to therapeutic agents
 Example: highly
successful therapeutic
use of the
differentiation-inducing
agent all-trans retinoic
acid
 All-trans retinoic acid
targets the chimeric
protein encoded by the
t(15;17) translocation
associated with acute
promyelocytic leukemia
• BCR/ABL t(9;22) is a translocation
qualitative assay by RT-PCR
o Should be ordered to screen
patients suspected of ALL or
CML for the presence of the
BCR/ABL transcript
TRIPLE C NOTES 14
 o Can be used to distinguish
between the major and minor
transcripts
o Major transcripts are
characterized by the p210
fusion gene product and are
typically detected in CML
o Minor transcripts are
characterized by the p190
fusion gene product and are
typically detected in ALL
o The BCR/ABL quantitative
assay is intended for monitoring
for:
 Genetic recurrence
 Minimal residual
disease
PRINCIPLES OF SPECIAL
CYTOCHEMICAL STAINS
Cytogenetics in Acute Lymphoblastic
Leukemia
• About half of patients with lymphoblastic
leukemia have abnormal karyotypes
• Structural changes in ALL include:
o t(9;22)
o t(4;11)
o t(8;14)
o t(8;22)
o t(2;8)
o Ph1 chromosome
• Gains in chromosome 21 and losses in
chromosome 7, 9, or 20 have all been
cited.
• No consistent markers have been
associated with L1 and L2 types of ALL,
but t(8;14) alteration is commonly seen
in the L3 type of ALL with Burkitt
lymphoma morphology and other
abnormalities of chromosome 14.
• Patients with more than 50
chromosomes in their leukemic cells
have the longest survival
• Patients with an abnormal karyotype
have somewhat shorter survival times
• Patients with a t(4;11), t(9;22), or t(8;14)
alteration do relatively poorly
• Special cytochemical stains can be used
as supplementary sources of information
in the identification and differentiation of
leukemias
• Cytochemistry is the application of
biochemical stains to blood and bone
marrow cells
o Reflect the chemical
composition of cells through the
use of color reactions, without
damaging the cell to the point at
which the cell itself can no
longer be recognized
• Cytochemical stains include:
o Sudan black B
o MPO
o Periodic-acid Schiff (PAS)
o Napthol AS-D chloroacetate
(NASCDA) esterase
o Alpha-napthyl acetate-butyrate
esterase with fluoride inhibition
o Leukocyte alkaline phosphatase
(LAP)
o Acid phosphatase with or
without tartaric acid inhibition
TRIPLE C NOTES 15
Sudan Black B Stain
• The Sudan stains, such as Sudan black
B, are substances belonging to a series
of lipid-soluble pigments that detect
cellular lipids
• Positive-staining reactions are
associated with the granulocytic
leukocytes
o The intensity of staining
becomes more pronounced as
the neutrophilic granulocytes
mature
• This procedure is helpful in
differentiating AML from ALL
Myeloperoxidase Stain
• In humans, peroxidases are found in the
microbodies of liver and kidney cells and
in the granules of myeloid and
monocytoid cells
• Myeloperoxidase is located in the
primary, azurophilic granules
o However, primitive blasts that
are committed to the myeloid
cell line demonstrate
myeloperoxidase activity in
areas such as the endoplasmic
reticulum and the Golgi region
• A positive reaction pattern in early cells
may appear as dots or rod-type
structures, referred to as phi bodies
• This procedure is useful in differentiating
acute myeloid and acute monocytic
leukemias from ALL
Periodic Acid-Schiff Stain
• The periodic acid–Schiff (PAS) reaction
is important in carbohydrate
histochemistry. Positive staining
reactions indicate the presence of
glycogen, a polymer of glucose, and
other 1,2-glycol–containing
carbohydrates
• Mature neutrophils contain high levels of
cytoplasmic glycogen, which is
physiologically related to the high
energy needs of neutrophils in
phagocytosis
• The PAS reaction is strongly positive in
neutrophilic granulocytes except blast
forms, immature and mature platelets,
and erythrocytes in erythroleukemia
(FAB M6)
• The usefulness of this procedure is in
establishing the negative characteristics
of myeloblastic and monoblastic
leukemias from lymphoblastic leukemias
Esterase Stains
• The nonspecific esterase enzymes
alpha-naphthyl acetate and butyrate
esterase are used clinically to recognize
cells of monocytic origin
o If the enzyme is of monocytic
origin, it is inhibited by sodium
fluoride
• Alpha-naphthyl acetate esterase, unlike
NASDCA esterase, is strongly positive
for monocytes, weakly positive or
negative for granulocytes, and positive
for other cell types
Phosphatase Stains
• The phosphatase enzymes are widely
distributed in mammalian tissue
• As a group, these enzymes liberate
orthophosphate from organic
phosphates
• The two major classifications are based
on pH
o Alkaline Phosphatase –
distinguishes leukemia and
leukemoid reaction (see
Chapter 26 – Manual
Procedures of Hematology)
o Acid Phosphatase - monocytes
demonstrate a more intense
positive reaction than do
neutrophils;
 Although lymphocytes
display little activity, T
cells do exhibit intense
positivity in the Golgi
region, whereas B cells
may be positive or
negative
MONOCLONAL ANTIBODIES
Immunophenotyping
• Immunophenotyping by flow cytometry
will confirm the diagnosis of leukemia or
establish a diagnosis in questionable
cases
• In addition to the surface membrane
markers discussed previously, it is
important to realize the role of
monoclonal antibodies in supplementary
TRIPLE C NOTES 16
 differential testing in the various
leukemias and lymphomas
Lymphoid
• The most useful antibodies are those
directed at CD20 (B cells) and CD3 (T
cell) membrane markers
o Precursor B cells express TdT,
CD9, CD 79a positive in early B-
cell precursors, hematogones
o In all, blasts are positive CD34
and TdT
B-Lineage Markers
• CD79a is more frequently present than
is CD20 in precursor B-cell ALL
• CD10 is positive in the majority of cases
of precursor B-cell ALL and is rare in T-
cell ALL. TdT and CD99 are usually
negative in mature B-cell ALL
T-Lineage Markers
• CD3 with TdT identifies cases of T-cell
ALL
• Other T-cell antibodies can be used in
selected cases:
o CD2
o CD5
o CD7
o CD1a
B-Cell Markers
• B-cell maturation is divided into early
pre-B, pre-B, and mature B-cell stages
• The specific surface marker beginning
with pre-B cell is CD19
• The mature B cell is identifiable by the
presence of surface immunoglobulin
(sIg)
Other Surface Membrane Markers
• Other surface membrane markers are
widely distributed among plasma cells,
some bone marrow cells, and some B-
cell malignancies
• The HLA-DR surface marker is found in
various cells
• In addition, some surface markers such
as CD11b, CD11c, CD13, and CD14 are
unique to granulocytes and monocytes
LIFE THREATENING EMERGENCIES
• The most common life-threatening
emergencies confronting patients with
acute leukemia are:
o Infection
o Bleeding
o Leukemic infiltration of organs
o Metabolic abnormalities
o Hyperleukocytosis
Treatment Options
• Refinements in the diagnosis of
subtypes of leukemia and advances in
therapeutic approaches have improved
the outlook for leukemia patients
• Cytotoxic chemotherapy for acute
leukemia patients is very intense
o Most patients with AML and ALL
will achieve remission, but high
relapse rates exist
• Induction therapy consists of
administration of multiple drugs aimed at
inducing a complete remission or
producing an absence of overt leukemia
in the bone marrow or at other sites
Relapse
• When treatment fails in patients with
AML, the available options are
determined by age, duration of the first
remission, and cytogenetic findings,
among other factors
Stem Cell Therapy
• AMLs and ALLs are treatable by stem
cell transplantation
• Progenitor blood cells are considered to
be pluripotent because they have the
ability to evolve into different types of
cells
Future Trends Vaccines
• Vaccines for cancer, including acute
myeloid leukemia, are under
investigation
• Proteins in the peptide vaccines are
produced in large amounts by cells of
MDS, AML, and CML patients
• The peptides are combined with an
“adjuvant” called Montanide to make the
vaccines, and the vaccines are given
with GM-CSF (sargramostim)
• Both Montanide and sargramostim help
the immune system respond to the
vaccines
• The vaccines then activate the immune
system to make specialized cells that
search out and kill the MDS, AML, and
CML cells containing the two proteins
TRIPLE C NOTES 17