Copyright and License Notice for PDF Courses

MediaLab courses are provided in PDF format for the sole use of MediaLab subscribers.

Distribution to non-subscribers is prohibited in every form, including electronic and print. Do not make multiple
copies of this PDF file.

If you are an individual subscriber, you are the only person authorized to use this PDF file. Please do not redistribute
it to others inside or outside your organization. Instead, please contact MediaLab about obtaining an institutional
subscription.

This Copyright and License Notice is part of the Terms of Service for MediaLab. If you have any questions, please
contact us.

MediaLab retains all copyright to this course and all material contained therein.
Acute Leukemia with a Focus on WHO Classification
Author: Margaret Reinhart, MS, MT(ASCP) Reviewer: Rory Huschka, M.Ed., MT(ASCP) and Laurie Bjerklie, M.Ed.,
MT(ASCP)
Course Instructions

Please proceed through the course by clicking on the blue arrows or text links. Use the table of contents to
monitor your progress. Your progress will be saved automatically as you proceed through the course, and
you may later continue where you left off even if you use a different computer. You may encounter practice
questions within the course, which are not graded or recorded.
Course Info

This course carries the following continuing education credits:

P.A.C.E. Contact Hours: 1.50 hour(s)

Florida Board of Clinical Laboratory Science CE - General (Hematology): 1.50 hour(s)
Course Number: 789382

Level of Instruction: Intermediate

Intended Audience: Laboratory personnel who are involved in hematology testing including technologists,
technicians, laboratory managers, supervisors and laboratory directors. Also useful as a review for MLS
students and those who plan to work in hematology after having been away from the department for several
years.

Author Information: Margaret Reinhart, MS, MT(ASCP) is senior lecturer in Biological Sciences at the
University of the Sciences in Philadelphia PA and was the MLS Program Director of Medical Laboratory
Science from 1990 - January 2020. She currently teaches hematology, clinical immunology, parasitology, and
other related courses. She is also adjunct MLS instructor in Hematology at Pennsylvania Hospital (University
of Pennsylvania Hospital System), Philadelphia PA. She holds a Masters Degree in Biology and in Health Care
Administration.

Reviewer Information:

Rory Huschka, M. Ed., MT(ASCP), has over 20 years of combined experience as a Medical Technologist,
Technical Supervisor, Professor, and Manager. He is the former Director of the Clinical Laboratory Science and
Medical Laboratory Technician Programs at Brookline College. He is currently a Program Director at
MediaLab, Inc. Rory holds a BS degree in Medical Technology from North Dakota State University and a
Masters in Educational Leadership.

Laurie Bjerklie, M.Ed., MT(ASCP), is currently a professor at Rasmussen College in Fargo, ND for their MLT
program where she teaches a variety of courses including hematology and microbiology. She is the former
Program Director of the Clinical Laboratory Science program at DeVry University in Phoenix, Arizona. Ms.
Bjerklie has over ten years of experience in higher education and continues to work part time as a bench tech.
 Definition and
Definition and Differentiation of Acute Differentiation of Acute
Leukemias from Chronic Leukemias Leukemia from other
Neoplastic Disorders.

Leukemia, lymphoma and myeloma are neoplastic

proliferative blood cell disorders. These disorders are
considered malignant (cancerous) conditions.
Leukemias usually originate in the bone marrow but can
invade other tissues including the lymphatic system,
whereas lymphomas originate in the lymphoid tissues, but
can invade other tissues including the bone marrow.
Leukemias frequently involve the white blood cells,
although red blood cells and platelets can also be involved.
Myelomas are a more specific type of disease in that the
malignancy involves plasma cells which are specific
lymphocytes that have already recognized and responded
to antigen, and are often located in the bone marrow.
Two major groupings of leukemia are acute leukemias and
chronic leukemias.

Definition and
Definition and Differentiation of Acute and
Differentiation of Acute
Chronic Leukemia continued Leukemia from other
Neoplastic Disorders.

Table 1 below compares and contrasts the characteristics of acute and chronic forms of leukemia.
Table 1. Acute and Chronic Forms of Leukemia.
Acute Leukemia Chronic Leukemia
Onset Sudden Slow and long duration
usually with symptoms sometimes asymptomatic at time of diagnosis

Age Adults and children Mostly found in adults

Hepatosplenomegaly Can be found but varies based on Common
Lymphadenopathy age and cell lineage
Bleeding/Bruising/Other Common Less common
Symptoms Fever common also
Cell Maturity Bone marrow (BM) and Peripheral BM and PB cells are more mature than in Acute
Blood (PB) cells are more immature
>20% blasts in bone marrow or
blood are typically found at
diagnosis
WBC Count Elevated Variable
Can range from very elevated to somewhat
below normal
Prognosis Survival in untreated patients is Survival in untreated patients ranges from
several weeks to several months several months to many years, depending on
type

Ungraded Practice Question

 Definition and
Ungraded Practice Question Differentiation of Acute
Leukemia from other
Neoplastic Disorders.
Characteristics of acute leukemia include all of the following EXCEPT:

Please select the single best answer

< 20% blasts in blood and/or bone marrow

Bleeding and bruising are common presenting symptoms

Survival time if untreated is several weeks to months

Onset of symptoms is fairly sudden

Definition and
Ungraded Practice Question Differentiation of Acute
Leukemia from other
Neoplastic Disorders.
Characteristics of acute leukemia include all of the following EXCEPT:

Please select the single best answer

< 20% blasts in blood and/or bone marrow

Bleeding and bruising are common presenting symptoms

Survival time if untreated is several weeks to months

Onset of symptoms is fairly sudden

Feedback

One of the defining criteria of acute leukemia is the presence of >20% blasts in the blood and/or bone marrow. Less than
20% blasts are seen in chronic leukemias.

Causes and Risk Factors

Causes of Acute Leukemias: Triggers of Genetic
of Acute Leukemia
Changes

Leukemia is a clonal disease that is caused by mutations and altered expression of genes leading to a malignant
transformation of hematopoietic precursors. In most cases, a specific cause of these genetic changes can not be identified.
However, we do know that a number of factors could be responsible for causing these genetic changes including:
Environmental exposures such as exposures to radiation (especially seen in acute and chronic myeloid leukemias)
Chemical and drug exposures such as benzene and organic solvents
Immunosuppression such as in organ transplant patients
Pre-existing genetic abnormalities as in Down Syndrome, Fanconi's Anemia, etc.
 Certain viruses (more often seen in chronic leukemias or lymphomas)
Treatment of previous cancerous conditions with chemotherapy and/or radiation

Causes and Risk Factors

Types of Genetic Changes in Acute Leukemia of Acute Leukemia

Many types of gene mutations, as well as chromosomal abnormalities, are found in acute leukemia. As we will discuss later,
these are in fact a basis of classification, treatment and prognosis.
Typically, it is not just one gene that is mutated, but multiple genetic changes need to occur to cause the leukemia. We
describe the genes affected as:
Oncogenes. These can cause neoplastic conditions through a variety of ways such as changing how procarcinogens
are metabolized, disabling the person's ability to repair DNA damage, altering the person's immune system or affecting
the regulation of cell growth.
Protooncogenes. These genes are the upregulators of cell growth and when mutated, can become oncogenes.
Tumor suppressor genes. These genes are the downregulators of cell growth, and if altered, can result in unchecked
proliferation.

The common chromosomal abnormalities seen in acute leukemia that can be visualized on chromosomal analysis include:
Translocations. A piece of one chromosome can break off and attach to another chromosome. This is one of the most
frequently seen chromosomal abnormalities in acute leukemia. The first translocation that was identified, however, was
in Chronic Myeloid Leukemia - the famous Philadelphia Chromosome (later this same translocation was found in other
leukemias). Translocations can cause altered expression of genes. For instance, they can position a promoter gene to
turn on a gene involved in cell cycle and replication.
Inversions. This occurs when part of the chromosome is turned around, causing altered expression of genes as well.
Deletions. In a deletion, part of the chromosome breaks off and is lost. If the lost gene(s) are tumor suppressor genes,
then replication can continue unchecked.
Additions. In this abnormality part of a chromosome is gained or there is an additional chromosome. This can cause
problems if a number of oncogenes are added.

Causes and Risk Factors

Occurrence and Frequency of Acute Leukemia
of Acute Leukemia

Lymphomas are much more common neoplastic disorders than acute leukemias. Of the acute leukemias, acute myeloid
leukemia (AML) is more common than acute lymphoid leukemia (ALL). Of the two, AML is more common in adults and ALL
is more common in children. A third acute leukemia, mixed phenotype acute leukemia (MPAL) is rare and can be found in
both age groups. As far as gender differences for all leukemias, males in the US have a slightly higher incidence than
females.
Although incidence of leukemia has increased over the years, deaths have decreased due to improved treatments and earlier
detection.
The remainder of this course will be about diagnosis and treatment.

Causes and Risk Factors

Ungraded Practice Question
of Acute Leukemia
Match the genetic or chromosomal abnormality to its definition:

Select the correct match for each item from the drop-down box

Choose Oncogene
 Choose Deletion

Choose Tumor suppressor gene

Choose Inversion

Choose Translocation

Causes and Risk Factors

Ungraded Practice Question of Acute Leukemia
Match the genetic or chromosomal abnormality to its definition:

Select the correct match for each item from the drop-down box

Choose Mutation in these genes can affect how Oncogene

carcinogens are metabolized

Choose Part of a chromosome containing tumor Deletion

suppressor genes is lost

Choose Down regulators of replication Tumor

suppressor
gene

Choose Part of a chromosome is turned around Inversion

Choose Part of a chromosome breaks off and is Translocation

attached to another one

Introduction to the
Classification Systems Classification and
Diagnosis of Acute
Leukemia

Both acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML), along with the other neoplastic blood disorders
were originally classified by the French-American-British (FAB) system. The FAB classification system was based primarily on
cell staging, cell morphology, and cytochemical staining. As genetic and chromosomal changes causing these disorders were
discovered and new techniques for identifying the changes were developed, the World Health Organization (WHO)
established a new classification system based on this new information.
The WHO classification is based on multiple parameters: cell staging, morphology and cytochemistry are part of the initial
diagnosis; chromosomal changes through karyotype analysis, molecular genetic changes, and immunophenotyping of cell
surface markers are then used to more specifically and accurately classify them.
Initial diagnosis also includes relevant clinical data such as medical history, possible toxic exposures, and sometimes physical
exam including imaging studies.
 Introduction to the
Initial Diagnostic Tests and Samples
Classification and
Diagnosis of Acute
Leukemia

The CBC, white blood cell differential, and blood smear examination are what initially alert us to the possibility of a leukemia
diagnosis in the laboratory. The laboratory professional will typically note elevated white counts, immature cells and cell
morphologies, and possibly abnormal red cell or platelet counts. The white cell differential can give a clue as to whether the
leukemia is acute or chronic, and myeloid or lymphocytic.
Cell staging and differentiation between myeloid and lymphocytic is an important first step, as is determining if the leukemia
is acute or chronic.
Further laboratory samples will then be evaluated. Samples will include bone marrow biopsy and aspirations, and less
commonly cerebrospinal fluid or tissue samples. Bone marrow morphologic evaluation is typically done by a pathologist;
thereafter, flow cytometry, cytogenetic analysis, and genetic studies will be performed for the definitive diagnosis.

Introduction to the
Distinguishing between Acute and Chronic Classification and
Leukemia Diagnosis of Acute
Leukemia

As mentioned previously, one of the important first steps

in diagnosis is whether the neoplastic disorder appears to
be acute or chronic. To make this determination, count the
percentage of blasts in the peripheral blood smear and
also in the bone marrow smear. A general guideline is that
if 20% or more blasts are found in one or both of these
specimens, then it is an acute leukemia. This is an
especially important distinction between AML and the
chronic myeloid disorders (e.g. myelodysplastic syndromes
(MDS) and myeloproliferative disorders (MPD), which may
also be referred to as myeloproliferative neoplasms
(MPN)).
The top image to the right shows numerous blasts in a
blood smear of AML. Note the characteristic large nuclei
with loose, lacey chromatin, high nuclear/cytoplasmic ratio, AML
and the presence of nucleoli. Also, there are few
representatives of other stages of maturation. Contrast this
to the smear below which is from a patient with chronic
myeloid leukemia (CML). Note that there are fewer blasts,
and more mature cells. A number of maturation stages can
be seen, such as myelocytes, metamyelocytes. and
segmented neutrophils.

CML
 Introduction to the
Differentiation between Myeloid and Lymphoid Classification and
Lines Diagnosis of Acute
Leukemia

The major WHO categories of acute leukemias are:

Acute Myeloid Leukemia (AML)
Acute Lymphoblastic Leukemia (ALL) - comprised of
Acute B Lymphoblastic and Acute T Lymphoblastic
Acute Leukemia of Ambiguous Lineage
Blastic Plasmacytoid Dendritic Cell Neoplasm

The first two - AML and ALL are by far the most common;
the second two are rare. Therefore, this course will cover
AML and ALL thoroughly, with only mention of the other
two.
Typically, when a white blood cell differential indicates a
substantial percentage of blasts, the first thing necessary is
to determine whether they are myeloblasts or
lymphoblasts. Although it might seem at first like
distinguishing between myeloid or lymphocytic cells is
quite obvious, that is not always the case when it comes to
neoplastic disorders. The first step is to look for obvious
clues. Acute leukemias are characterized by a ''leukemic
gap', which refers to the presence of both blasts and
mature cells, while representative cells in the intermediate
stages are absent. So one way to obtain a clue is to look at
the mature cells. If most of the mature cells seen are
lymphocytes, then it could possibly be ALL. If neutrophils
or other cells in the myeloid line are seen, then it is more
likely to be an AML.
One clue that is almost foolproof is the presence of Auer
Rods. Auer Rods are red staining, elongated needle-like
inclusions that can be seen in the cytoplasm of AML
myeloblasts. They result from the fusion of azurophilic (red
staining) granules. If Auer Rods are seen, you can be
certain that the leukemia is of myeloid origin. However, if
they are not seen, you can not strictly rule out myeloid
leukemias as they aren't seen in everyone or all the time.
(Note: the presence of Auer Rods indicates that the cell is
of myeloid lineage. but doesn't define it as acute. Most
likely it is AML, but Auer Rods can occasionally be seen in
other neoplastic disorders of myeloid lineage.) The image
to the right shows Auer Rods in myeloblasts. Table 2 can
be useful in morphological differentiation.
Table 2. Comparing AML and ALL.
AML ALL
Blast Size Medium to large Small to medium
Cytoplasm Fine granules may be Usually no granules or few
present coarse
Presence Yes No
of Auer
Rods

Nucleus Nuclear chromatin finely 1-3 indistinct nucleoli

dispersed; 2-4 prominent
nucleoli
Image of
Blast

(1) (2)

If Auer Rods are not seen, then cytochemical stains can be

used to distinguish the lineages, or one can go directly to
other diagnostic tests. Though not used as often anymore,
the following page will give a synopsis of the various
cytochemical tests. When not used, the next step for
diagnosing the leukemia may be genetic testing or
immunophenotyping.
1. Najmaldin Saki, PhD; Ahmad Ahmadzadeh, MD; Marziye Bagheri, MSc. "We

present a picture from bone marrow aspiration sample of acute myeloblastic

leukemia (AML-M2) patient." American Society of Hematology, 19 Jan 2019,

http://imagebank.hematology.org/image/62152/acute-myeloblastic-leukemia-

amlm2-3?type=upload

2. VashiDonsk. "A Wright's stained bone marrow aspirate smear of patient w ith

precursor B-cell acute lymphoblastic leukemia. Picture taken by me." Wikimedia

Commons, 15 Jan 2007,

https://commons.w ikimedia.org/w iki/File:Acute_leukemia-ALL.jpg

Introduction to the
Cytochemical Tests Used in Differentiating Classification and
Myeloid Cells from Lymphocytic Cells Diagnosis of Acute
Leukemia

Table 3 lists some of the cytochemical stains used in differentiating lineages (there are additional stains not listed here).
Table 3. Cytochemical Tests Used in Differentiating Myeloid Cells from Lymphocytic Cells
Stain Site of Cells Stained Comments Image
Action
Myeloperoxidase Primary Myeloblasts; Separates AML+ from AML-
Granules Granulocytes;
Auer Rods Monocytes slight
positive

(3)
Sudan Black B Phospholipids Myeloblasts; Separates AML+ from AML-
Granulocytes;
Monocytes slight
positive

(4)

Naphthol AS- Cytoplasm Neutrophilic Separates AML+ from AML-

DChloroacetate/(CAE) - granulocytes;
specific esterase Mast cells

(5)

Periodic acid-Schiff Glycogen Granular pattern with Positive in erythroleukemia,

Stain (PAS) negative background in abnormal erythrocyte
lymphoblasts precursors, and ALL

(6)

3. Giri, Dhurba. "Myeloperoxidase (MPO) Stain: Purpose, Principle, Procedure and Interpretation." LaboratoryTests.org, 6 Nov 2018,

http://laboratorytests.org/myeloperoxidase-mpo-stain/

4. Giri, Dhurba. "Sudan Black B Stain: Purpose, Principle, Procedure and Interpretation." LaboratoryTests.org, 18 Nov 2018, http://laboratorytests.org/sudan-black-b-stain/

5-6. Naphthol AS-D chloroacetate/(CAE) - specific esterase and periodic acid-schiff stain images courtesy of Dr. Jui-Han Huang, MD.

Introduction to the
Differentiation between Myeloid and
Classification and
Lymphoblastic Leukemias Using Cluster of Diagnosis of Acute
Differentiation (CD) Markers Leukemia

As previously mentioned, other tests exist to differentiate between myeloid and lymphoid lines. One way is by the use of
monoclonal antibodies to detect surface markers on the cells. This is done using flow cytometry techniques which will be
described in more detail later. As far as initial differentiation, however, Table 4 is useful to differentiate the major cell lines.
Table 4. Differentiation between Myeloid and Lymphoblastic Leukemias Using CD Markers.
Cell Lineage CD Markers
Myeloid CD13, CD33, CD15, CD117
T-Cell CD2, CD3, CD5, CD7
B-Cell CD19, CD20, CD22, CD79a
Megakaryoblastic CD41, CD61

Introduction to the
Ungraded Practice Question
 Introduction to the
Ungraded Practice Question
Classification and
Diagnosis of Acute
Leukemia
Which of the following cytochemical stains would yield a positive result in AML?

More than one answer is correct. Please select all correct answers

Sudan Black B

Periodic Acid Schiff (PAS)

Myeloperoxidase

Introduction to the
Ungraded Practice Question Classification and
Diagnosis of Acute
Leukemia
Which of the following cytochemical stains would yield a positive result in AML?

More than one answer is correct. Please select all correct answers

Sudan Black B

Periodic Acid Schiff (PAS)

Myeloperoxidase

Feedback

Sudan Black B and Myeloperoxidase stains are positive in AML.

PAS is useful for erythroleukemia and ALL.

Introduction to the
Ungraded Practice Question Classification and
Diagnosis of Acute
Leukemia
The World Health Organization (WHO) classification system relies solely on cell staging, cell morphology and cytochemical
stains.

Select true or false

True

False

Introduction to the
Ungraded Practice Question
 Introduction to the
Ungraded Practice Question
Classification and
Diagnosis of Acute
Leukemia
The World Health Organization (WHO) classification system relies solely on cell staging, cell morphology and cytochemical
stains.

Select true or false

True

False

Feedback

It is the FAB system that is based solely on cell morphology, staging, and cytochemical stains.
The WHO system is based on additional information such as genetic mutations, karyotyping of chromosomes, and
immunophenotyping.

Acute Myeloid Leukemia

Signs and Symptoms of AML (AML) Diagnosis

Unlike chronic neoplastic disorders, symptoms of AML may be present for only a few days or weeks before the patient is
diagnosed. Although they can vary, the most common presenting symptoms of AML are those caused by crowding out of
normal hematopoietic lines such as:
Anemia
Thrombocytopenia
Granulocytopenia

Typical symptoms associated with anemia are fatigue, pallor, dyspnea, and others. Thrombocytopenia manifests as mucosal
bleeding, easy bruising, and heavy periods. Occasionally, patients can have more serious effects such as spontaneous
hemorrhage. Granulocytopenia leads to a greater risk of infections, recurrent infections, and fevers.
Much less common is leukemia cutis, which is the infiltration of the epidermis, dermis, or subcutis by leukemic cells. This
condition causes nodules or papules on the skin. Leukemic cell infiltration of other organs is also possible but less common
or severe than in ALL; however, the liver, spleen, lymph nodes, joints and meninges can occasionally be affected.

Acute Myeloid Leukemia

Complete WHO AML Classification
(AML) Diagnosis: 2016
WHO classification of
AML

The WHO classification of AML includes many more individual diagnostic categories than the old FAB list of M0-M7.
The major categories of AML are:
AML with recurrent genetic abnormalities
AML with myelodysplasia-related changes
AML not otherwise specificied (NOS)- (several of the subcategories of this group are on the FAB M0-M7 list)
Myeloid sarcoma
Myeloid proliferation related to Down Syndrome

Table 5 gives a complete list of the subcategories of each category. This is for your reference, and it is beyond the scope of
this course to cover each. A few of the more common ones will be mentioned in subsequent pages.
Table 5. Complete WHO AML Classification.
AML Category Subcategories

AML with recurrent genetic abnormalities AML with t(8;21)(q22;q22.1);RUNX1-RUNX1T1

AML with myelodysplasia-related changes
Therapy-related myeloid neoplasms
AML NOS (not otherwise specified)
Myeloid sarcoma
Myeloid proliferations related to Down
Syndrome
AML with inv(16)(p13.1q22) or t(16;16)(p13.1;q22);CBFβ-
MYH11
APL with PML-RARA
AML with t(9;11)(p21.3;q23.3);MLLT3-KMT2A
AML with t(6;9)(p23;q34.1);DEK-NUP214
AML with inv(3)(q21.3q26.2) or t(3;3)(q21.3;q26.2); GATA2,
MECOM
AML (megakaryoblastic) with t(1;22)(p13.3;q13.3);RBM15-
MKL1
Provisional entity: AML with BCR-ABL1
AML with mutated NPM1
AML with biallelic mutations of CEBPA
Provisional entity: AML with mutated RUNX1
AML with minimal differentiation
AML without maturation
AML with maturation
Acute myelomonocytic leukemia
Acute monoblastic/monocytic leukemia
Pure erythroid leukemia
Acute megakaryoblastic leukemia
Acute basophilic leukemia
Acute panmyelosis with myelofibrosis
Transient abnormal myelopoiesis (TAM)
Myeloid leukemia associated with Down Syndrome

Acute Myeloid Leukemia

Interpretation of Genetic Designations of the (AML) Diagnosis: 2016
AML Subtypes WHO classification of
AML

As you saw in the previous page, WHO classification is

heavily based on cytogenetics and genetics. Here is how to
interpret the designations, taking the example of the first
one listed - AML with t(8;21)(q22;q22.1);RUNX1-
RUNX1T1:
- t = translocation. The two numbers in parentheses after
t are the numbers of the chromosomes involved in the
translocation.
- The numbers following "q" in the next set of
parenthesis refers to bands on the q or long arm of
the chromosome where that translocation created a
fusion of two genes that initially had not been in proximity
with each other.
- The last part, RUNX1-RUNX1T1 is the name of the actual A smear demonstrating an AML-M2 - t(8;21) abnormality (7).
genes involved. That fusion (RUNX1-RUNX1T1) leads to
the production of a chimeric protein which disrupts the
normal function of a transcription factor complex that
regulates normal hematopoiesis.
So in other words, a part of chromosome #21 broke off
and attached to chromosome #8, resulting in the fusion of
genes which resulted in a protein that interfered with the
regulation of hematopoietic activity - thus, unchecked
proliferation.
Also, just to make the correlation with the FAB
classification, this particular type of AML corresponds
morphologically with the M2 subtype in FAB.
Other designations that you may see include the
designation inv which means that part of the chromosome
was inverted such as in AML with inv(16)(p13.1q22); p
indicates the short arm of the chromosome, whereas q
indicates the long arm.
The image shows a blood smear of the AML with t(8;21)
(q22;q22.1);RUNX1-RUNX1T1: subtype, showing
myeloblasts (note the abundant cytoplasm in these blasts
and azurophilic granules).
7. Patho. "Bone Marrow : Acute Myeloblastic Leukemia w ith Maturation (AML-

M2) Associated w ith a t(8;21) Chromosome Abnormality." Wikimedia Commons, 8

Jul 2008, https://commons.w ikimedia.org/w iki/File:AML-

M2_associated_w ith_a_t(8;21)_chromosome_abnormality.jpg

Acute Myeloid Leukemia

AML FAB Classification (AML) Diagnosis: 2016
WHO classification of
AML

Although the French-American-British (FAB) classification has been replaced, it is still often referred to and provides a point
of reference. Table 6 lists the FAB categories.
Table 6. AML FAB Classification.
FAB Subtype Name
M0 Undifferentiated acute myeloblastic leukemia
M1 Acute myeloblastic leukemia with minimal maturation
M2 Acute myeloblastic leukemia with maturation
M3 Acute promyelocytic leukemia
M4 Acute myelomonocytic leukemia
M4 eos Acute myelomonocytic leukemia with eosinophila
M5 Acute monocytic leukemia
M6 Acute erythroid leukemia
M7 Acute megakaryocytic leukemia
If you look back on the WHO classifications, you will see that a number of these names are listed under the WHO group
"therapy related myeloid neoplasms."

Acute Myeloid Leukemia

Occurrence and Common Types of AML (AML) Diagnosis: 2016
 WHO classification of
AML

AML in general is much more common in adults than children. Average lifetime risk of AML in the US is approximately 0.5%
which translated to approximately 19,520 new cases in 2018.
According to the Merck Manual, some of the more common cytogenetic abmormalities of AML include:
AML with APL-RARA (t15;17)(q24.1). This is a type of promyelocytic leukemia (FAB category M3). Approximately 13%
of AML is this subtype.
AML with inv(16)(p13.1;q22) or t(16;16)/CBFB-MY11. Approximately 5% of AML is this subtype.
AML with t(8;21)/(q22;q22)/RUNX1-RUNX1T1. This was the example shown on a previous page and occurs in
approximately 7% of AML cases.

Prognosis related to these and several other subtypes will be discussed in subsequent pages.

Acute Myeloid Leukemia

Ungraded Practice Question (AML) Diagnosis: 2016
WHO classification of
AML
Match the abbreviation found in the WHO AML designations with the meaning of the designation:

Select the correct match for each item from the drop-down box

Choose p

Choose inv

Choose q

Choose t

Choose uppercase letters at the end of the cytogenetic designation

Acute Myeloid Leukemia

Ungraded Practice Question
(AML) Diagnosis: 2016
WHO classification of
AML
Match the abbreviation found in the WHO AML designations with the meaning of the designation:

Select the correct match for each item from the drop-down box

Choose short arm of p

chromosome

Choose inversion inv

Choose long arm of chromosome q

 Choose translocation t

Choose genes involved uppercase letters at the end of the cytogenetic

designation

Acute Myeloid Leukemia

Ungraded Practice Question
(AML) Diagnosis: 2016
WHO classification of
AML
The WHO classification of AML can be based on all of the following criteria except:

Please select the single best answer

Age of the patient

Cytogenetic analysis of the leukemic cells

Cell type and morphology of the cells

Genetic changes

Acute Myeloid Leukemia

Ungraded Practice Question (AML) Diagnosis: 2016
WHO classification of
AML
The WHO classification of AML can be based on all of the following criteria except:

Please select the single best answer

Age of the patient

Cytogenetic analysis of the leukemic cells

Cell type and morphology of the cells

Genetic changes

Feedback

Multiple criteria can be used to determine the WHO AML subtype including cytogenetics, actual genes involved, and the cell
types and morphology. Although age can figure into the prognosis, it does not determine the diagnosis.

Acute Myeloid Leukemia

Basics of Laboratory Testing (AML) Diagnosis: AML
 laboratory testing

As stated earlier, laboratory diagnosis begins with finding immature cells on the peripheral blood smear. If blasts are seen,
the next step would be to characterize the lineage of the blasts.
Bone marrow biopsies and smears are then obtained. If blasts are 20% or greater in the bone marrow and/or peripheral
blood, presumptive acute leukemia can be determined.
Additional steps that can be taken:
1. Further morphologic evaluation
2. Cytochemical staining
3. Conventional cytogenetic analysis
4. Appropriate molecular genetic and/or FISH studies
5. Obtaining possible CSF sample, skin or other biopsies, depending on patient's clinical situation

Steps 1 and 2 were discussed earlier in the sections on "differentiation between myeloid and lymphoid lines" and
"distinguishing between acute or chronic leukemia".
Steps 3 and 4 will be discussed in the following pages.

Acute Myeloid Leukemia

Cytogenetic Analysis (AML) Diagnosis: AML
laboratory testing

Two-thirds of patients with AML have leukemic blasts

containing chromosomal abnormalities. These
abnormalities differ among the various subtypes, as
indicated in the WHO classification on previous pages.
Cytogenetic analysis helps to inform important clinical,
prognostic, and treatment decisions. Also, the diagnosed
patient can then be monitored for remission and relapse
using chromosomal and molecular analysis.
The analysis of chromosome morphology is known as
karyotyping. Karyotyping is performed by growing the
patient's white blood cells in special media to induce cell
division and replication. Cells are then treated to stop cell
A normal male karyotype (8).
division at the metaphase stage of replication. The cells are
then lysed to release the chromosomes and then stained to
show the banding. A picture of the stained chromosomes
is taken. The chromosomes are counted, sorted by
chromosome number and paired, and then analyzed for
their structure and banding. Table 7 lists some of the
stains used to visualize the chromosomes.
Table 7. Stains Used for Visualizing Chromosomes.
Stain Name What the Stain Yields
Giemsa g banding
Acridine Orange r banding
Treated Giemsa c banding
A karyotype of a male with t(6;9)AML (9).
Quinacrine r banding
Newer methods include spectral karyotyping such as
multiplex fluorescence in situ hybridization (M-FISH),
which is particularly useful in more complex
rearrangements. Various automated methods can also be
used for performing the karyotypes and visualizing the
chromosomes.
The top image shows a normal karyotype (karyogram) of a
male.
The lower right image shows a karyotype (karyogram) of a
male with AML with a translocation of chromosomes 6 and
9 (short arrows) with a further chromosomal change of an
additional chromosome 8 (long arrow).
8. National Human Genome Research Institute. "Karyotype of a human male."

Wikimedia Commons, 1 Mar 2012, https://commons.w ikimedia.org/w /index.php?

curid=583512

9. Song, Yeohan et al. “The Challenge of t (6;9) and FLT3-Positive Acute

Myelogenous Leukemia in a Young Adult.” Journal of leukemia(Los Angeles, Calif.)

vol. 2 (2014): 1000167. doi:10.4172/2329-6917.1000167

Acute Myeloid Leukemia

Molecular Genetics of AML
(AML) Diagnosis: AML
laboratory testing

Although most diagnoses of AML genetic changes are based on whole chromosomal changes seen when doing a
karyogram, smaller scale mutations are becoming increasingly important. This is especially true for approximately 40% of
cases where no chromosomal change has been identified. The following mutations have been found to be important in AML
prognosis and treatment:
FLT-3
NPM1
CEBPA
KIT

Many other mutations are being discovered as well. These gene mutations affect transcription and thus replication either
directly, or through epigenetic regulation. As molecular methods advance, gene testing will become increasingly important.
Next generation sequencing (high-throughput sequencing) is a method used to detect these mutations. Although expensive
and time-consuming, the number of panels are becoming commercially available, and it is likely that this type of testing will
be done more widely in the future. Just to stress how important this information is becoming, it has been shown that in the
AML groups with recurrent genetic abnormalities, finding the NPM1 and CEBPA mutations has been associated with a very
favorable prognosis.
It should be noted, however, that in studying the whole genome of AML patients, many mutations were found that were
actually silent or unrelated to AML.

Acute Myeloid Leukemia

Ungraded Practice Question
(AML) Diagnosis: AML
laboratory testing
Karyotyping used in the diagnosis of AML refers to:

Please select the single best answer

Making images of the patient's chromosomes to detect chromosomal changes or abnormalities

Sequencing genes to detect mutations

Staining cells to determine cell lineage

None of the above

 Acute Myeloid Leukemia
Ungraded Practice Question
(AML) Diagnosis: AML
laboratory testing
Karyotyping used in the diagnosis of AML refers to:

Please select the single best answer

Making images of the patient's chromosomes to detect chromosomal changes or abnormalities

Sequencing genes to detect mutations

Staining cells to determine cell lineage

None of the above

Feedback

Karyotyping to produce a karyogram is done to be able to analyze chromosomes for abnormalities.

Genetic mutations are also important in AML diagnosis, but this process is not known as karyotyping.
Staining cells to determine cell lineage is called cytochemical staining.

Acute Myeloid Leukemia

Ungraded Practice Question (AML) Diagnosis: AML
laboratory testing
Mutations that cause AML generally are to the genes that regulate which of the following?

Please select the single best answer

Cell metabolism

Cell replication

Export of proteins

None of the above

Acute Myeloid Leukemia

Ungraded Practice Question (AML) Diagnosis: AML
laboratory testing
Mutations that cause AML generally are to the genes that regulate which of the following?

Please select the single best answer

Cell metabolism
 Cell replication

Export of proteins

None of the above

Feedback

Harmful mutations leading to AML (as well as other neoplastic disorders) generally are those of genes regulating replication
and inhibition of replication.
Although mutations in genes for cell metabolism and export of proteins can cause other diseases, they are not the causative
mutations of AML.

AML Prognosis and

Prognostic factors of AML
Treatment

Accurate assessment of the prognosis of the patient is important for overall management and treatment of AML. Patients can
be stratified according to their risk of treatment resistance or treatment-related mortality. This will help guide the type and
intensity of treatment.
Important prognostic factors are:
The WHO subtype
Certain cytogentic subtypes have a much better prognosis than others (see subsequent pages).

Clinical Factors
Age at diagnosis. Increased age is associated with a poorer prognosis, although this is also dependent on the WHO
subtype.
Prior hematological malignancies. If the patient had a prior hematological malignancy, this carries a substantially
poorer prognosis.

Other Variables (These variables can be especially important in older patients.)

Platelet count
Serum creatinine
Albumin

AML Prognosis and

Types of AML treatments
Treatment

Treatment of AML is extremely complex, and is based on the genetic changes, the patient's age, and other prognostic factors.
In general, possible treatment regimens consist of:
chemotherapeutic agents
target therapies directed at the specific genetic change
stem cell transplantation

The aim of chemotherapeutic drugs is to stop the cell cycle, and thus halt the proliferation of the leukemic cells, or in some
cases, directly kill rapidly proliferating cells. Table 8 shows a few examples of such drugs. Keep in mind that
chemotherapeutic drugs are used not only to treat AML, but other neoplastic conditions such as chronic leukemias and solid
tumor cancers as well; and, in so doing, these treatments can actually cause AML.
Table 8. Types of AML Treatments.
Group of Drugs Mechanism of Action Example
Alkylating Agents Adding alkyl groups to cause breakage of DNA strands Busulfan
Antimetabolites Prevents DNA replication Methotrexate

Mitotic Inhibitors Plant alkaloids that damage cells and prevent replications Vinsristine
Corticosteroids Induce apoptosis in leukemic cells Dexamethasone
Can also induce differentiation

AML Prognosis and

Newer treatments
Treatment

Although chemotherapeutic drugs are often the mainstay of many forms of AML, newer targeted therapies are constantly
being developed. Targeted therapy refers to a drug that targets or interacts specifically with a protein, enzyme, or receptor
that is being expressed as a result of the mutation that is causing the AML.
An early targeted therapy that was developed was actually to CML in which the translocation known as the Philadelphia
Chromosome caused the fusion of the bcr-abl genes which triggered the overexpression of an enzyme known as a tyrosine
kinase. This enzyme is pivotal in the unchecked replication of the cells. By inhibiting this tyrosine kinase, the cells can be
induced to stop replicating and to differentiate.
Newer research shows that some of the mutations in AML can also involve tyrosine kinases, and thus newer generations of
these inhibitors may be useful in this disease as well.

AML Prognosis and

Method of AML treatment
Treatment

Although it depends on the exact subtype of AML, as well as the patient's age, clinical condition, and other factors, it can
generally be said that treatment for AML is done in two phases. The first is known as induction therapy, in which the
patient is often given a chemotherapeutic drug to achieve complete remission. The definition of complete remission is
revised from time to time, but it can be basically defined as a state of normal hematopoiesis after the induction therapy.
Patients should be evaluated after about two weeks of induction therapy by examination of bone marrow aspirate and
biopsy. If these do not indicate that the patient is in remission, then they typically would receive another round of induction
therapy.
Those patients that do show they are in complete remission are often offered consolidation therapy in order to treat any
residual disease that has not been detected. Options for such treatment include chemotherapy and allogeneic hematopoietic
stem cell transplants.

AML Prognosis and

Prognosis of AML
Treatment

As stated earlier, accurate diagnosis of the specific WHO subtype will help to determine the patient's prognosis, their
response to therapy, their likelihood of remission, and their overall chances of survival.
According the Merck Manual Professional Edition (2018), Table 9 provides some examples of different WHO subtypes and
their prognoses.
Table 9. WHO Subtypes and their Prognoses of AML.
Prognosis Cytogenetic Abnormality/Mutation
Favorable t(15;17)(q24.1q24.1)/PML-RARA
t(8;21)/(q22q22.1)/RUNX1-RUMX1T1
AML with inv(16)(p13.1q22) or t(16;16)(p13.1;q22);CBFB- MYH11
Intermediate Normal karyotype
Poor Trisomy 8
 AML Prognosis and
Ungraded Practice Question Treatment

Important factors which can help determine a patient's prognosis of AML include all of the following EXCEPT:

Please select the single best answer

Patient's family history

The WHO subtype

The patient's platelet count

The patient's age

AML Prognosis and

Ungraded Practice Question
Treatment
Important factors which can help determine a patient's prognosis of AML include all of the following EXCEPT:

Please select the single best answer

Patient's family history

The WHO subtype

The patient's platelet count

The patient's age

Feedback

Although having cancers or leukemias in the patient's family history could possibly make it more likely to develop such a
disease, it does not really bear on the prognosis.
The WHO subtype and patient's age are two very important prognostic factors. Additionally, in certain subtypes, the platelet
count can help inform the prognosis.

AML Prognosis and

Ungraded Practice Question Treatment

What is the mechanism of action of chemotherapeutic drugs known as alkylating agents?

Please select the single best answer

Inducing apoptosis

Targeting the product of a mutated gene

Breaking DNA strands

 AML Prognosis and
Ungraded Practice Question
Treatment
What is the mechanism of action of chemotherapeutic drugs known as alkylating agents?

Please select the single best answer

Inducing apoptosis

Targeting the product of a mutated gene

Breaking DNA strands

Feedback

The mechanism of action of alkylating agents, used commonly in the treatment of AML is that the drug breaks DNA strands
thus preventing replication. This can affect many rapidly proliferating cells, not just the leukemic cells.
The so-called targeted therapies are those developed to interfere with a specific protein that resulted from the genetic
mutation.
Corticosteroids are known to induce apoptosis.

Group of Drugs Mechanism of Action Example

Alkylating Agents Adding alkyl groups to cause breakage of DNA stands Busulfan

Antimetabolites Prevents DNA replication Methotrexate

Mitotic Inhibitors Plant alkaloids that damage cells and prevent replications Vinsristine

Corticosteroids Induce apoptosis in leukemic cells Dexamethasone

Can also induce differentiation

Acute Lymphoblastic
Signs, symptoms, and background of ALL
Leukemia (ALL) Diagnosis

Acute lymphoblastic leukemia is the most common pediatric cancer. 60% of all ALL cases occur in children, with a peak
incidence at age two to five years old. A second peak occurs in adults over 50. As with AML, chromosomal abnormalities and
genetic alterations affect the differentiation and proliferation of precursor cells.
Presenting symptoms can include:
fatigue
pallor
infections
CNS symptoms
easy bruising and bleeding
abdominal pain
swollen lymph nodes
difficulty breathing (especially in T Cell-ALL)

Acute Lymphoblastic
Diagnosing ALL
 Acute Lymphoblastic
Diagnosing ALL Leukemia (ALL) Diagnosis

As with AML, ALL is first suspected when blasts are seen

on the peripheral blood smear. 20% or greater blasts in the
blood or bone marrow constitute a diagnosis of acute
leukemia, and then establishing that they are lymphoblasts
will give an initial diagnosis as ALL. Techniques as
described in the section on cytochemical stains can help
distinguish myeloblasts from lymphoblasts.
In ALL, lymphoblasts can sometimes be very numerous, as
much as 90% of the total white blood count. Bone marrow
smears and biopsies are usually done next; the bone
marrow can typically show between 25-95% lymphoblasts.
To further establish a definitive diagnosis,
immunophenotyping using flow cytometry and
cytogenetic studies can be performed. More information
on this will follow under laboratory diagnosis.
Also, a lumbar puncture to analyze the CSF is usually done
upon initial diagnosis. If the central nervous system is
involved, further testing might be done. Coagulation
studies and baseline chemistries are also done.
The image at the right shows a smear with numerous
lymphoblasts.
10. Patho. "Bone Marrow : Acute Lymphoblastic Leukemia, L1 (ALL-L1)." Wikimedia

Commons, 8 Jul 2008, https://commons.w ikimedia.org/w iki/File:ALL-L1.jpg

A smear with numerous lymphoblasts (ALL L1) (10).

Acute Lymphoblastic
Ungraded Practice Question Leukemia (ALL) Diagnosis

Adults rarely get ALL; it is typically only found in children.

Select true or false

True

False

Acute Lymphoblastic
Ungraded Practice Question
Leukemia (ALL) Diagnosis

Adults rarely get ALL; it is typically only found in children.

Select true or false

True

False
Feedback

Although ALL is the most common pediatric cancer, it can also be seen in adults of all ages, with a peak incidence over 50
years old.

Acute Lymphoblastic
Complete WHO ALL Classification
Leukemia (ALL)
Diagnosis: 2016 WHO
classification of ALL

In the old FAB classification systems, leukemia and

lymphoma were a point of initial differentiation. A disease
was called a lymphoma if it was initially or mostly affecting
the lymphatic system (lymph nodes, spleen, etc.), although
it could spread to the bone marrow and blood. The
disorder was termed leukemia if the major site was the
bone marrow and blood, although it could also affect the
lymphatic system.
However, in the WHO classification, the initial
differentiation is based on whether the lineage is that of a
B cell or T cell which can be determined by cell surface
markers using flow cytometry.
The WHO classification of acute lymphoblastic leukemia
was revised in 2016, and includes the following types A Wright's stained bone marrow aspirate smear of patient with
described in Table 10. precursor B-cell acute lymphoblastic leukemia (2).
Table 10. WHO Classification of Acute Lymphoblastic
Leukemia.
Category Subcategory
B-cell lymphoblastic leukemia/lymphoma,
not otherwise specified
B-cell lymphoblastic leukemia/lymphoma B-cell lymphoblastic leukemia/lymphoma with hypodiploidy
with recurrent genetic abnormalities
B-cell lymphoblastic leukemia/lymphoma with hyperdiploidy
B-cell lymphoblastic leukemia/lymphoma with t(9;22)(q34;q11.2)[BCR-
ABL-1]
B-cell lymphoblastic leukemia/lymphoma with t(v;11q23)[MLL
rearranged]
B-cell lymphoblastic leukemia/lymphoma with t(12;21)(p13;q22)[ETV6-
RUNX-1]
B-cell lymphoblastic leukemia/lymphoma with t(1;19)(q23;p13.3)
[ZTCZX3-PBX1]
B-cell lymphoblastic leukemia/lymphoma with t(5;14)(q31;q32)[IL-
3IGH]
B-cell lymphoblastic leukemia/lymphoma with intrachromosomal
amplification of chromosome 21 (iAMP21)
B-cell lymphoblastic leukemia/lymphoma with translocations involving
tyriosine kinases or cytokine receptors (BCR-ABL1-like ALL)
T-cell lymphoblastic Early T-cell precursor lymphoblastic leukemia
leukemias/lymphomas
Notice that some of the chromosomal and genetic changes
are similar to those seen in AML.
The image on the right is of a B Cell lymphoblastic
leukemia.
2. VashiDonsk. "A Wright's stained bone marrow aspirate smear of patient w ith

precursor B-cell acute lymphoblastic leukemia. Picture taken by me." Wikimedia

Commons, 15 Jan 2007,

https://commons.w ikimedia.org/w iki/File:Acute_leukemia-ALL.jpg

Acute Lymphoblastic
FAB Classification
Leukemia (ALL)
Diagnosis: 2016 WHO
classification of ALL

In the former FAB classification, ALL was divided into three

types based on cell size, prominence of nucleoli, and
amount and appearance of cytoplasm:
L1 (found mostly in children)
L2 (found mostly in older children and adults)
L3 (associated with Burkitt's Lymphoma)

The image to the right is a smear from a bone marrow that

was diagnosed as ALL L1, often seen in children. Notice the
consistently sized lymphoblasts with scant cytoplasm.
11. Hanggara, Dian Sukma. "Leukemia Limfoblastik Akut L1 (ALL-L1)."

PatologiKlinik.com, 4 Aug 2017, https://patologiklinik.com/2017/08/04/leukemia-

limfoblastik-akut-l1-all-l1/

A smear from a bone marrow that was diagnosed as ALL L1 (11).

Acute Lymphoblastic
Ungraded Practice Question
Leukemia (ALL)
Diagnosis: 2016 WHO
classification of ALL
All of the following are classification criteria in the WHO classification system of ALL EXCEPT:

Please select the single best answer

Surface markers to determine lymphoblast lineage

The amount and appearance of the cytoplasm of the lymphoblasts

Chromosomal abnormalities

Genetic changes
 Acute Lymphoblastic
Ungraded Practice Question Leukemia (ALL)
Diagnosis: 2016 WHO
classification of ALL
All of the following are classification criteria in the WHO classification system of ALL EXCEPT:

Please select the single best answer

Surface markers to determine lymphoblast lineage

The amount and appearance of the cytoplasm of the lymphoblasts

Chromosomal abnormalities

Genetic changes

Feedback

The amount and appearance of the cytoplasm of the lymphoblasts was an important diagnostic criteria in the former FAB
system; however, surface markers, chromosomal changes, and genetic mutations are now essential diagnostic criteria of the
WHO system.

Acute Lymphoblastic
Ungraded Practice Question Leukemia (ALL)
Diagnosis: 2016 WHO
classification of ALL
An essential point of differentiation in the WHO classification system of ALL is whether the disorder is a leukemia or
lymphoma.

Select true or false

True

False

Acute Lymphoblastic
Ungraded Practice Question
Leukemia (ALL)
Diagnosis: 2016 WHO
classification of ALL
An essential point of differentiation in the WHO classification system of ALL is whether the disorder is a leukemia or
lymphoma.

Select true or false

True

False
Feedback

The WHO classification is based on chromosomal and genetic changes which can be found in disorders that can present as
either a leukemia or lymphoma.

Acute Lymphoblastic
Determination of ALL Lineage Leukemia (ALL)
Diagnosis: ALL laboratory
testing

As previously mentioned, once it is determined that the patient has 20% or greater blasts, the next step is to establish
whether they are lymphoblasts or myeloblasts. Pages 11 and 12 deal with morphological differences as well as using
cytochemical stains. Also, histochemical staining for terminal deoxynucleotidyl transferase (TdT) can be useful in determining
that the blast is of lymphoid lineage. TdT is an enzyme found in immature (developing) lymphocytes which functions to help
synthesize their specific antigen receptors.
If it is found to be Acute Lymphoblastic Leukemia, the next important step is to discover whether the lymphocytes involved
are T cells or B cells. This can be accomplished by immunophenotyping for "markers" - that is , cell surface proteins that are
specific for either B cells or T cells. Flow cytometry uses fluorescently stained antibodies specific for these markers. Using this
technique, we can not only determine B or T cell lineages, but we can also count the numbers in a given quantity of blood or
bone marrow.
These membrane proteins or markers usually have numbers prefaced by "CD" for Cluster of Differentiation. Among the
many markers that can be detected, finding CD3 can identify the cell as a T cell and CD19, CD20, and CD22 will indicate B cell
lineage. Table 11 shows some examples of CD markers for different cell lines.
Table 11. CD Markers for ALL Cell Lines.
Cell Type CD Marker
Leukocyte CD45
T cells (all) CD3
T helper cells CD4
Cytotoxic T cells CD8
B cells CD19 and CD20

Acute Lymphoblastic
Chromosomal Analysis Leukemia (ALL)
Diagnosis: ALL laboratory
testing

Chromosomal analysis can detect a specific translocation

or other chromosomal abnormality such as hyperdiploidy
or hypodiploidy, and thus help establish the WHO subtype.
Hyperdiploidy refers to excess numbers of chromosomes;
in ALL we can often see as many as 51-67 chromosomes!
Interestingly, this hyperdiploidy is found commonly in
childhood ALL, is not associated with substantial genetic
abnormality except for the excess chromosomes, and
actually renders a good prognosis. Hypodiploidy refers to
fewer than 46 chromosomes
The image to the right shows hyperdiploidy in a childhood A karyotype showing B cell ALL hyperdiploidy (12).
case of ALL (B cell).
12. Woo, Jennifer S et al. “Childhood B-acute lymphoblastic leukemia: a genetic
update.” Experimental hematology & oncology vol. 3 16. 13 Jun. 2014,

doi:10.1186/2162-3619-3-16

Acute Lymphoblastic
Genetic Analysis
Leukemia (ALL)
Diagnosis: ALL laboratory
testing

Although chromosomal aberrations are the hallmark of ALL, they are not sufficient to generate leukemia. It is usually the
genetic changes that are linked to overexpression or underexpression of various genes which regulate proliferation or
maturation. It is interesting that a Philadelphia Chromosome (seen in CML) can be found in some cases of ALL. This involves
the BCR/ABL1 genes and the tyrosine kinases which are targeted by some CML treatments.
Some commonly seen genetic changes include:
ETV6-RUNX1 (caused by a translocation of 12;21)
IKZF1 (involves a deletion of a key transcription factor important in B cell development)
BCR/ABL1 (involves a kinase activating factor)

Predisposing factors that can lead to these changes include certain diseases such as Down Syndrome or Fanconi anemia,
exposure to radiation, pesticides, or other biological or chemical toxins. However, most cases of ALL appear de novo in
previously health individuals.

Acute Lymphoblastic
Ungraded Practice Question Leukemia (ALL)
Diagnosis: ALL laboratory
testing

A three year old has been diagnosed with ALL. Chromosomal analysis performed on this child's lymphocytes showed 53
chromosomes. A true statement about this case is:

Please select the single best answer

This child has hypodiploidy

This finding is associated with a good prognosis

This child has Down's Syndrome

This most likely was a mistake

Acute Lymphoblastic
Ungraded Practice Question
Leukemia (ALL)
Diagnosis: ALL laboratory
testing

A three year old has been diagnosed with ALL. Chromosomal analysis performed on this child's lymphocytes showed 53
chromosomes. A true statement about this case is:

Please select the single best answer

 This child has hypodiploidy

This finding is associated with a good prognosis

This child has Down's Syndrome

This most likely was a mistake

Feedback

Although it seems quite abnormal to have duplications of chromosomes, cases of ALL with excess chromosomes often have
a good prognosis.
Excess numbers of chromosomes (greater than 46) is called hyperdiploidy, not hypodiploidy.
Although Down Syndrome is associated with a chromosomal aberration, and people with Down's syndrome are more
susceptible to ALL, having 53 chromosomes is not specific to Down's syndrome.
It is unlikely that seeing extra chromosomes in a chromosomal analysis is a mistake.

ALL Prognosis and

Prognostic Factors for ALL
Treatment

As with AML, an accurate assessment of prognosis is important for the treatment and management of ALL. Both clinical
factors and cytogenetic changes impact prognosis and treatment.
The two most important clinical factors are age and white blood count. The younger the age and the lower the white blood
count at the time of initial diagnosis, the better the prognosis. Even including cases with less favorable cytogenetic profiles,
the prognosis for children of disease-free survival for five years is >80%. Someone with disease-free survival for five years
is considered to be cured.
Among cytogenetic findings, hyperdiploidy (~51-65 chromosomes) and the absence of Philadelphia Chromosome are
associated with a good prognosis.
The bullets below summarizes some of the clinical and genetic prognostic factors.
Favorable prognostic factors
Age 3-9 years
WBC count <25,000/ μL in adults or <50,000 in children
Hyperdiploidy (51-65 chromosomes)
T(1/19) and t(12/21)
No CNS involvement at diagnosis

Unfavorable prognositc factors

Karyotype with hypodiploidy (<46 chromosomes) at diagnosis
Very high hyperdiploidy - 66-88 chromosomes at diagnosis
Presence of Philadelphia Chromosome BCR-ABL1
Other specific translocations associated with an unfavorable outcome
Increased age in adults
More frequent and severe comorbidities

ALL Prognosis and

Common Cytogenetic Abnormalities and their Treatment
Prognosis
The following are among the more common cytogenetic abnormalities:
Those with a favorable prognosis:
hyperdiploidy (51-65 chromosomes in leukemia cells)
t(12;21)/TEL-AML1 (ETV6-RUNX1)

Those with an unfavorable prognosis:

t(9;22)/BCR-ABL1; (Philadelphia Chromosome positive)
hypodiploidy (<46 chromosomes in leukemia cells)

ALL Prognosis and

Treatment of ALL Treatment

Types of treatment for ALL include systemic chemotherapy, prophylactic CNS chemotherapy (sometimes CNS radiation), and
supportive care. For some patients, immunotherapy, targeted therapy, stem cell transplantation and/or radiation therapy
might be used. For Philadelphia Chromosome (+) patients, a targeted therapy of a tyrosine kinase inhibitor can be used.

The method of treatment is similar to AML treatment. It begins with induction therapy to induce complete remission (defined
as <5% blast cells in the bone marrow, an absolute neutrophil count of >1000/uL, a platelet count of 100,000 and no need
for blood transfusion ). The success of initial induction treatment is often indicative of positive overall survival. Typical
chemotherapy drugs to induce remission are vincristine, corticosteroids, and an anthracycline.
After induction, some patients may go on to receive an allo-stem cell transplant. Others will go on to receive more
chemotherapy in an intensification/consolidation phase.
Following the intensification/consolidation phase, patients may then be put on a maintenance chemotherapy for up to 3
years.
ALL treatment is very complex; it depends on the patient's initial diagnosis, age, general health and initial induction therapy
response. There are numerous drugs that are used, as well as radiation, and on the horizon are immunotherapies and
targeted therapies.

ALL Prognosis and

The Future of ALL Therapy Treatment

Many different targeted therapies are in various phases of research, trials and approval. Some examples include:
Monoclonal antibodies. These are antibodies which are synthesized to recognize a membrane target expressed by the
leukemic cells, thus lessening the side effects often seen with standard chemotherapies. A common target for ALL is
the CD22 membrane marker. The monoclonal antibody is conjugated with some type of toxin or enzyme inhibitor, thus
delivering it directly to the leukemic cells.
Proteosome inhibitors. Proteosomes are the large protein complexes in cells which degrade unneeded or excess
proteins. By inhibiting this action, proteins build up in the cell and will eventually kill it.
JAK inhibitors. A signaling pathway in cells known as the JAK/STAT pathway is a way that leukemic cells can bypass
normal growth and proliferation restrictions. By inhibiting this bypass, the cells will stop proliferating and may go on to
mature.

ALL Prognosis and

Ungraded Practice Question
Treatment
A three year old is diagnosed with ALL. Choose the characteristics which would yield the most favorable prognosis for this
child. Check all that apply:

More than one answer is correct. Please select all correct answers

A white blood count of 65,000/ μL

 62 chromosomes

Philadelphia Chromosome

No CNS involvement

ALL Prognosis and

Ungraded Practice Question Treatment

A three year old is diagnosed with ALL. Choose the characteristics which would yield the most favorable prognosis for this
child. Check all that apply:

More than one answer is correct. Please select all correct answers

A white blood count of 65,000/ μL

62 chromosomes

Philadelphia Chromosome

No CNS involvement

Feedback

Hyperdiploidy (51-65 chromosomes) and no CNS involvement are both very favorable prognostic factors, as well as the
patient's age.
Finding the Philadelphia Chromosome, or leukemic involvement in the central nervous system are very unfavorable
prognostic factors.

Acute Leukemia of
Acute Leukemia of Ambiguous Lineage Ambiguous Lineage and
Blastic Plasmacytoid
Dendritic Cell Neoplasm

Acute leukemia of ambiguous lineage (ALAL) was a

subtype established in the 2016 WHO classification
revision. It is a very rare leukemia, that is further
subdivided into five possible subgroups, partially based on
chromosomal analysis.
ALAL patients present with an acute leukemia, but a
specific lineage can not be assigned.
Some types of ALAL are immature hematopoietic
neoplasms that show no differentiation into lymphoid or
myeloid lines (AUL), whereas other types exhibit markers
of both AML and ALL. The latter are referred to as mixed
phenotype acute leukemia (MPAL).
The image to the right shows a case of ALAL. The arrow
points to hypolobate neutrophils, emphasizing how
unusual some of these cases can be. A smear showing ALAL with hypolobate neutrophils (13).
When immunophenotyping these leukemias, AUL lacks T-
cell specific, B-cell specific, and myeloid specific markers.
MPAL, on the other hand, shows various
immunophenotypes.
There are no clinical features that are specific to ALAL. but
rather have signs and symptoms similar to AML and/or
ALL. ALAL does generally carry a poor prognosis.
13. Moraveji, S et al. “Acute leukemia of ambiguous lineage w ith trisomy 4 as

the sole cytogenetic abnormality: A case report and literature review.” Leukemia

research reports vol. 3,2 33-5. 2 May. 2014, doi:10.1016/j.lrr.2014.04.003

Acute Leukemia of
Blastic Plasmacytoid Dendritic Cell Neoplasm Ambiguous Lineage and
Blastic Plasmacytoid
Dendritic Cell Neoplasm

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a

rare but very aggressive leukemia. Patients show
characteristics similar to leukemia and lymphoma; it is
found in the bone marrow and blood, but can spread to
lymph nodes, spleen, central nervous system, and skin. The
skin lesions are characteristic and appear deep purple and
numerous. Most patients have been elderly males.
This leukemia derives from plasmacytoid dendritic cells
and like ALAL has phenotypic ambiguity. It has also been
known as blastic NK cell leukemia/lymphoma.
The leukemic cells are negative for many of the
cytochemical stains such as myeloperoxidase, α-
naphthylbutyrate esterase, ND naphthol AS-D
A bone marrow aspirate of BPDCN (14).
chloroacetate esterase. They show complex karyotypes and
genetic mutations.
Treatment requires high levels of chemotherapy. Because
this disease is usually found in very elderly patients who
typically can not tolerate such regimens, targeted therapies
are being pursued.
The image to the right is a bone marrow aspirate of
BPDCN which shows medium to large cells with scant
cytoplasm, immature chromatin, irregular nuclear contours
and prominent nucleoli.
14. Sullivan, Jill M, and David A Rizzieri. “Treatment of blastic plasmacytoid

dendritic cell neoplasm.” Hematology. American Society of Hematology. Education

Program vol. 2016,1 (2016): 16-23. doi:10.1182/asheducation-2016.1.16

Acute Leukemia of
Ungraded Practice Question Ambiguous Lineage and
Blastic Plasmacytoid
Dendritic Cell Neoplasm
Acute leukemia of ambiguous lineage (ALAL) is a common leukemia in which the lineage can not be specified or which
shows features of both lymphoid and myeloid lines.
Select true or false

True

False

Acute Leukemia of
Ungraded Practice Question Ambiguous Lineage and
Blastic Plasmacytoid
Dendritic Cell Neoplasm
Acute leukemia of ambiguous lineage (ALAL) is a common leukemia in which the lineage can not be specified or which
shows features of both lymphoid and myeloid lines.

Select true or false

True

False

Feedback

Although it is true that ALAL has either non-specific or mixed lineage, it is quite a rare leukemia.

References
References

Arber, Daniel A et al. “The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute
leukemia.” Blood vol. 127,20 (2016): 2391-405. doi:10.1182/blood-2016-03-643544
Ballesteros, Enrique, MD, Sohani, Allyah R., MD, Fedoriw, Yuri, MD. "Pathology of Acute Leukemia of Ambiguous Lineage."
Medscape, 8 Dec 2020, https://emedicine.medscape.com/article/1977857-overview
Cessna, Melissa H., MD, Wang Sa A., MD. Initial Diagnostic Workup of Acute Leukemia: A Pocket Guide for the Clinician.
College of American Pathologists, Mar 2017, https://documents.cap.org/documents/acute-leukemia-pocket-guide.pdf
Dastugue, Nicole et al. “Hyperdiploidy with 58-66 chromosomes in childhood B-acute lymphoblastic leukemia is highly
curable: 58951 CLG-EORTC results.” Blood vol. 121,13 (2013): 2415-23. doi:10.1182/blood-2012-06-437681
Davis, Amanda S., MD, Viera, Anthony J., MD, MPH, Mead, Monica D., MD. "Leukemia: An Overview for Primary Care." Am
Fam Physician. 89(9):731-738. 1 May 2014, https://www.aafp.org/afp/2014/0501/p731.html
Emadi, Ashkan, MD, PhD, York Law, Jennie, MD. "Acute Lymphoblastic Leukemia (ALL)." Merck Manual Professional Version,
May 2020, https://www.merckmanuals.com/professional/hematology-and-oncology/leukemias/acute-lymphoblastic-
leukemia-all
Emadi, Ashkan, MD, PhD, York Law, Jennie, MD. "Acute Meyloid Leukemia (AML)." Merck Manual Professional Version, May
2020, https://www.merckmanuals.com/professional/hematology-and-oncology/leukemias/acute-myeloid-leukemia-aml
Lagunas-Rangel, Francisco Alejandro et al. “Acute Myeloid Leukemia-Genetic Alterations and Their Clinical Prognosis.”
International journal of hematology-oncology and stem cell research vol. 11,4 (2017): 328-339. PMID: 29340131
Kouchkovsky, De, Abdul-Hay, M. “'Acute myeloid leukemia: a comprehensive review and 2016 update'.” Blood cancer journal
vol. 6,7 e441. 1 Jul. 2016, doi:10.1038/bcj.2016.50
McPherson R, Pincus M, eds. Henry's Clinical Diagnosis and Management by Laboratory Methods. St. Louis MO: Elsevier;
2017.
Seiter, Karen, MD et. al. "Acute Myeloid Leukemia (AML) Staging." Medscape, 30 Dec 2020,
https://emedicine.medscape.com/article/2006750-overview
Sullivan, Jill M, and David A Rizzieri. “Treatment of blastic plasmacytoid dendritic cell neoplasm.” Hematology. American
Society of Hematology. Education Program vol. 2016,1 (2016): 16-23. doi:10.1182/asheducation-2016.1.16
Terwilliger, T, and M Abdul-Hay. “Acute lymphoblastic leukemia: a comprehensive review and 2017 update.” Blood cancer
journal vol. 7,6 e577. 30 Jun. 2017, doi:10.1038/bcj.2017.53
Turgeon, Mary Louise. Clinical Hematology Theory and Procedures 5th ed. Philadelphia: Wolters Kluwer/Lippincott William
& Wilkins; 2012.