Tabuk University

Faculty of Applied Medical SciencesDepartment Of Medical Lab.

Technology

3rd Year – Level 5 – AY 1434-1435

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HEMATOLOGY – 2, MLT 307

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ACUTE MYELOID LEUKEMIA (AML)

By/Mr. Waqqas Elaas;

M.Sc; MLT

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Objectives• Define AML and recognize the different subtypes.• Recognize the common clinical presentation of a patient

with AML.• Identify various prognostic factors in patients with AML.• Understand the different complications that are a result of

the disease.• Classify AML.• Cite lab. methods for diagnosing AML.

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What are Leukemias?• Leuko = Leuco = white cells• Aemia = related to blood• The leukaemias are a group of disorders characterized by the

accumulation of malignant white cells in the bone marrow and blood. These abnormal cells cause symptoms because of:

(i) bone marrow failure (i.e. anaemia, neutropenia, thrombocytopenia) and

(ii) infiltration of organs (e.g. liver, spleen, lymph nodes, meninges, brain, skin or testes).

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Classification• According to cell type with regard to both cell

maturity and cell lineage.– Chronic

• More mature cells• Usually occurs in adults• Clinical onset is gradual• Anemia and thrombocytopenia are mild• WBC is increased• Organomegaly is prominent

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Classification– Acute

• Immature cells• Occurs in all ages• Clinical onset is sudden• Anemia and thrombocytopenia are mild to severe• WBC is variable• Organomegaly is mild

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Cell lineage

• Lymphoid• Myeloid

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Leukemias

Acute Chronic

AML ALL CML CLL

Classification

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Acute leukaemia• Acute leukaemia is defined as the presence of over 20% of

blast cells in the blood or bone marrow at clinical presentation. It can be diagnosed with even less than 20% blasts if specific leukaemia-associated cytogenetic or molecular genetic abnormalities are present.

• It is further subdivided into acute myeloid leukaemia (AML) and acute lymphoblastic leukaemia (ALL) on the basis of whether the blasts are shown to be myeloblasts or lymphoblasts.

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AML• AML is a cancer of the bone marrow characterized by the

accumulation of Myeloblasts in the bone marrow and blood, usually with more than 20% of nucleated cells.

• AML causes bone marrow to produce too many immature white blood cells (blast cells). This Suppresses normal blood cell production.(Anemia, leucopenia, thrombocytopenia)

• AML occurs in all age groups.• It is the common form of acute leukemia in adults.• Generally is a disease of older people and is uncommon before

the age of 40.• Slightly more common among men than women.

Myeloid maturation

myeloblast promyelocyte myelocyte metamyelocyte band neutrophil

MATURATION

Adapted and modified from U Va website

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Aetiology (causes)• Idiopathic (most).• Several genotoxic agents have been linked to AML (1-2% of causes)

– Ionizing radiation, benzene, atomic bomb exposure, cigarette smoking.

• AML secondary to prior chemotherapy (5-10%) : e.g.:– Alkylating agents– Topoisomerase II inhibitors

• Secondary to congenital defects (rare) : e.g.:– Down syndrome - trisomy 21– Fanconi anemia - defect in DNA repair

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Clinical features Clinical features are a result of the following:• Bone marrow failure : anaemia (pallor = شحوب, lethargy = كسل); neutropenia (fever, malaise=تعب, infections of mouth, throat,

skin, respiratory or others ); and thrombocytopenia (spontaneous bruises=رضوض,

pupura= الجلد تحت bleeding gums and ,نزفmenorrhagia= كثيفة شهرية .(دورة

• Organ infiltration : Tender bones=غضة, lymphadenopathy= اللمفاوية الغدد moderate ,تضخمsplenomegaly, hepatomegaly and meningeal syndrome (headache, nausea and vomiting, blurring of vision=تشويش and diplopia= مزدوجة .(رؤية

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Classification• FAB (French-American-British) scheme, which divides AML

into eight variants; according to morphology & cytochemistry. This is the old classification.

• The World Health Organization (WHO) classification attempts to be more clinically useful and to produce more meaningful prognostic information than the FAB criteria. Each of the WHO categories contains numerous descriptive sub-categories.

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FAB classification of AMLM0— undifferentiated blasts 3 - 30% blasts, – No azurophilic granules

M1—blasts Without maturation More than 90% cells are myeloblasts -MPO+, SBB++8 frequently seen

M2—blasts With maturation 30 - 89% are myeloblastsMore than 10% maturing myeloids (promyelocyte and beyond)

M3—Promyelocytic More than 30% hypergranular promyelocytesAuer rods – DIC - t(15:17) is diagnostic

M4—Myelomonocytic More than 30% blast (myeloblasts, monoblasts and promonocytes)More than 20% monocytic elements (NSE+)More than 20% myeloid elements (MPO+, SBB+)

M5M5a—Monoblastic(more than 80% monoblasts)M5b—Monocytic (less than 80% monoblasts)

More than 30% blast (myeloblasts, monoblasts and promonocytes)More than 80% NSE+ : monocytic elementsLess than 20% MPO+, SBB+ : myeloid elements

M6—Erythroid M6a—ErythroleukemiaM6b—Pure erythroid leukemia

More than 50% of nucleated marrow cells are erythroid precursorsMore than 30% of non erythroid elements are myeloid blasts PAS+

M7—Megakaryocytic More than 30% blasts of megakaryocytic lineage – assoc. with fibrosis

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1. CBC2. Blood film3. Bone marrow4. Biochemical tests (not diagnostic)5. Cytochemistry6. Immunophenotype/Flow cytometry7. Cytogenetic analysis

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Laboratory diagnosis1. CBC :• TWBCs : may be decreased, normal or increased to 200 x 109/L

or more.• Normocytic normochromic anemia• Thrombocytopenia.2. Blood film examination shows a variable numbers of blast cells. These are immature precursors, with large size, and primitive nuclei (ie the nuclei contain nucleoli), sometimes (in M3); Auer Rods are seen. (it is difficult to differentiate under the microscope between cells of AML & ALL) 3. Bone marrow examination : hypercellular with >20% leukaemic

blasts.4. Biochemical tests may reveal a raised serum uric acid, serum

lactate dehydrogenase (LDH) or, less commonly, hypercalcaemia.

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AML

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Auer rods are clumps of azurophilic granular material that form elongated needles seen in the cytoplasm of leukemic blasts. Auer rods result from the coalescence اندماج/ of primary التحامgranules

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Auer rods in AML

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Laboratory diagnosis (cont.)5. Cytochemistry:

Non-specific esterase stain is used to identify a monocytic component in AMLs.

TdT :Terminal Deoxynucleotidyl Transferase, is an enzyme marker for primitive lymphoid cells

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AML – cytochemistry

Reaction

M0

M1 M2 M3 M4 M5 M6 M7

Peroxidase (POX)

- + + + +/- - +/- -

Sudan Black B - + + + +/- - +/- -

Unspecific esterases

- - - - + + - -

PAS - - - - - - + -

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Laboratory diagnosis (cont.)6. Immunophenotype/Flow cytometry

Immunophenotyping is a technique used to study an Ag expressed by cells. It involves the labeling of white blood cells with antibodies directed against surface proteins on their membrane. The labelled cells are processed in a flow cytometer, a laser-based instrument capable of analyzing thousands of cells per second.

Cluster of differentiation (CD): is a protocol used for the identification and investigation of cell surface molecules present on white blood cells, providing targets for immunophenotyping of cells. (also called cell markers).

Common positive cell markers in AML are : CD13, CD33, CD34

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Immunophenotype (cont.)• Flow cytometry

– Fast and accurate way to identify, quantify and determine lineage

– Physical properties• Forward scatter--cell size• Side scatter--cytoplasmic

granularity– Cells can be stained with

fluorescently labeled antibodies that recognize cell markers

• CD34– Stem cell marker

• CD117, CD33, CD13, MPO– Myeloid markers

• CD14, CD64– Monocytic marker

• Glycophorin A– Erythroid marker

• CD41, CD61– Megakaryocytic markers

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Laboratory diagnosis (cont.)• 7. Cytogenetic analysis:(analysis of chromosomal abnormalities)

• t(15;17)(q22,q12)• t(8;21)(q22,q22)• inv(16)(p13,q12)• 11q23

** Acute promyelocytic leukemia (M3) is frequently associated with disseminated intravascular coagulation (DIC = pathological activation of coagulation); and so laboratory findings of DIC may seen (including : Thrombocytopenia, prolonged PT & PTT, low fibrinogen level, high FDPs, D-dimer)

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Homework

• What can you do to reduce the risk of leukemia?

• A 15 year old male had flu-like symptoms with a severe sore throat for two weeks prior to admission. On physical Exam; he showed cervical and axillary adenopathy. No other organomegaly. His CBC showed :

RBCs: 3.2 x 1012/L, HB: 9.9 g/dL, MCV: 90.5 fL (normal), WBC :42.6 x 109/L, Lymphocytes :12 %, Blast cells : 88%, PLT 22 x 109/L.

1. Is it possible that the patient had an acute infection? Why?2. If the patient had leukemia, is it ALL or AML? Why?