MLAB 1415- Hematology
Keri Brophy-Martinez
Introduction to Hematopoietic Neoplasms
Terms

Neoplasm or tumor: “New growth”
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Results from a dysregular prolieferation of a single transformed cell
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Can be malignant or benign
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Malignant
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“Deadly”
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Clone of abnormal, proliferating cells, without function or
differentiation
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Have the potential to metastasize or get bigger
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“Cancer”
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Benign
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Premalignant
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Originate from highly organized, differentiated cells
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Do not spread or invade surrounding tissues
Classification of Neoplasms in Bone Marrow
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Lymphoid
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Myeloid
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Only lymphocytic cells affected
Granulocytes, monocytes, megakaryocytes,
erythrocytes affected
Both Lymphoid and Myeloid lines can
include benign and malignant neoplasms
More Terms
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Leukemia
 A malignant disease of hematopoietic tissue characterized
by replacement of normal bone marrow elements with
abnormal (neoplastic) blood cells. Abnormal cells are also
seen in peripheral blood
Lymphoma
 Abnormal proliferation of lymphoid cells within the
lymphatic tissue or lymph nodes, results in a solid tumor
Leukemias
General Classification
Acute
Chronic
Age
All ages
Adults
Clinical onset
Sudden
Gradual
Course of disease
Weeks to months
Months to years
Predominant cell
Blasts
Some mature forms
Mature forms
Anemia
Mild-severe
Mild
Thrombocytopenia
Mild-severe
Mild
WBC
Variable
Increased
Organomegaly
Mild
Prominent
Blood Picture
Acute
Chronic
How Do Leukemias Arise?
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
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Somatic mutation of a single
hematopoietic stem or progenitor
cell
Unlimited self-renewal of the
cancer-initiating cell
As the mutant cell line
predominates, normal
hematopoiesis is inhibited
causing leukemic cells to spill
into peripheral blood
Proto-oncogenes and Oncogenes



Proto-Oncogene—normal unaltered gene that has the
potential to become an oncogene
Oncogene–Altered cell genes that cause tumors
Located at breakpoints of chromosomal aberrations, such as
translocations
Oncogene Activation

Factors

Genetic susceptibility

Fanconi’s anemia

Down’s syndrome (18-20 fold increased incidence)

Somatic mutation
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Ionizing radiation, nuclear weapons
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Chemicals and drugs

Benzene,Chloramphenicol, phenylbutazone

Certain chemotherapy drugs that are cytotoxic, especially when used in
conjuction with therapeutic radiation

Viral infection
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Retrovirus-HIV-1, HTLVI, II

EBV

Immunologic dysfunction

Ataxia-telangiectasia - lymphoid leukemia or lymphoma

Sex-linked agammaglobulinemia
Epidemiology

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Most new cases found in older adults ( > 67 yrs old)
50% of leukemias are acute
More common in whites
More common in males
Age groups
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ALL: children 2-5 years old: lymphoid
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CLL: Adults> 50: lymphoid
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AML: adults: myeloid
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CML: adults: myeloid
Evaluation of Leukemia


Note onset of symptoms
Analyzing CBC

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Observe cell lineage

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Thrombocytopenia?
RBC level/ anemia?
Lymphoid or Myeloid?
Assess maturity of predominating cells
Lab Features and Patient Symptoms

Normochromic,normocytic anemia
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Thrombocytopenia
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If count decreased- results in infections
Immature leukocyte precursors seen
Bone marrow hypercellular


Results in bleeding episodes
Platelet morphology and function can be abnormal
Leukocyte count can be increased, decreased or normal


Due to erythropenia
Bone pain due to marrow expansion
Maturation abnormalities in all cell lines
Uric acid increased

Due to cellular breakdown
Official Hematopoietic Neoplasm Classification


Two systems
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French-American British(FAB)
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Historical

World Health Organization (WHO)**

Widely accepted
Important because..

Allows clinicians a way to compare therapeutic regimens
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System for ID and comparison of clinical features & lab findings

Permit associations of cytogenetic abnormalities with disease
FAB classification
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Consists of three groups
 Myeloproliferative Disorders(MPD)
 Myelodysplastic Syndromes(MDS)
 Acute leukemia (AL)
Based on morphological characteristic of Wrightstained cells in peripheral blood or bone marrow
with supplementary cytochemical stains
WHO Classification
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
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Classification system uses morphology, cytochemistry and
immunophenotyping to determine cell lineage and degree of
maturation(similar to FAB)
Additionally uses genetic and clinical features prior to therapy and history
of MDS to define subgroups
Consists of three groups
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Myeloid
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Lymphoid
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Further classified as MPD, MDS/MPD, MDS, AML
Further classified as B/T cell, T/NK cell, Hodgkin’s Disease, CLL
Histiocytic
Lab Techniques for Diagnosis and classification of
neoplasms

Cytochemical analysis
 In vitro staining of cells to look at cells’ chemical
composition
 Evaluation of positivity in these stains must be determined
on the leukemic blast stage of the cell
 Usually performed on bone marrow slides
 Helpful in differentiating lymphoid or myeloid lineage of
blasts in AL
 Reactions are either enzymatic or nonenzymatic
Cytochemical stains
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Types
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Myeloperoxidase (MPO)
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Activity is present in the primary granules and Auer rods of
myeloid cells
Separates myeloid and lymphoid blasts
Stains late myeloblasts, granulocytes, monocytes less intensely
Differentiates AML from ALL
Granules stain black to reddish-brown
Cytochemical stains: con’t
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Sudan Black B

Activity is present in phospholipids in the membrane of 1̊ (nonspecific)
and 2̊ granules (specific)

Differentiates AML from ALL
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Granules stain black
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Periodic Acid Schiff (PAS)
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Activity is in glycogen and related substances

Stains lymphocytes, granulocytes, megakaryocytes

Helpful in diagnosing erythroleukemia where there is strong reactivity
in normoblasts
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Stains red-purple in blocks in cytoplasm
Cytochemical stains: con’t
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Esterases
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Specific Esterase (Naphthol AS-D Chloroacetate)

Activity is in cytoplasm

Stains neutrophilic granulocytes, differentiates monoblasts and
myeloblasts
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Granules of myeloblasts stain blue-black

Nonspecific Esterase (Alpha-Naphthyl Acetate)

Activity is in cytoplasm
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Stains monocytes and also megakaryocytes
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Differentiates myeloblasts from monoblasts
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Granules stain orange red
Cytochemical stains: con’t


Leukocyte Alkaline Phosphatase (LAP)
 Enzyme within the 2O or specific granules of maturing
granulocytes
 Distinguishes leukemoid reactions ( ) from chronic
myelogenous leukemia ( )
Acid Phosphatase (Tartrate Resistant)
 Present in lysosomes in normal leukocytes
 Helpful in diagnosing hairy cell leukemia because they
are NOT inhibited by TRAP
Cytochemical stains: con’t
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Terminal Deoxynucleotidy transferase (TdT)
 Primitive cell marker found in cell nuclei
 Distinguishes ALL from malignant lymphoma
Toluidine Blue
 Positive marker for basophils and mast cells
Lab Techniques for Diagnosis and classification of
neoplasms
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Immunologic marker studies
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Cell surface markers


Monoclonal or polyclonal antisera is added to cell
suspensions of fresh peripheral blood or bone marrow
and an immunofluorescent method is used in a flow
cytometry instrument to analyze the markers which
are expressed as cluster designations (CD).
CD’s identify antibodies that are specific for certain
cells and allow for a positive identification.
Lab Techniques for Diagnosis and classification of
neoplasms

Molecular Genetics

This newer method of diagnosing leukemia consists of DNA probes and
polymerase chain reaction (PCR)-based studies. They are rapid and precise
and are used to confirm chromosomal abnormalities that are not detected by
conventional studies. They are also used to monitor residual disease
following therapy.

FISH (Fluorescence in situ hybridization)
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Cytogenetics (Chromosome studies)

Identifies chromosome translocations which are specific for
certain leukemias

Philadelphia chromosome (t[9:22]) is associated with CML
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t[15:17] is associated with acute promyelocytic leukemia
Treatment
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Cures are not common except in childhood
leukemia. The best hope for a cure in adults
lies in bone marrow transplantation.
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Cytoreductive chemotherapy
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Reduces the leukemic cell mass
Block DNA synthesis
Block RNA synthesis
Complications arise from marrow hypoplasia and
resulting cytopenia
Treatment

Radiotherapy (radiation)

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Kills focalized leukemic cells
Usually used in addition to chemotherapy and for CNS
prophylaxis
Bone marrow transplantation
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Bone marrow is eradicated with chemo and radiation.
Compatible donor cells are transfused and they travel to the
empty marrow where they engraft and repopulate the marrow
with healthy cells.
Complications include graft vs host (GVH) disease which can be
fatal.
References


McKenzie, Shirlyn B., and J. Lynne. Williams. "Chapter 21."
Introduction. Clinical Laboratory Hematology. Boston:
Pearson, 2010. Print.
http://www.lls.org/content/nationalcontent/resourcecenter/fre
eeducationmaterials/generalcancer/pdf/facts.pdf