Leukemia/Lymphoma

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Leukemia/Lymphoma
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Components and General Properties of Blood

• Seven kinds of formed elements
• Erythrocytes red blood cells (RBCs)
• Platelets (thrombocytes) (clotting cells)
• Cell fragments from special cell in bone marrow
• Leukocytes white blood cells (WBCs)-5 kinds

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Components and General Properties of Blood

• Five leukocyte types divided into two categories
• Granulocytes (with granules)
• 3)Neutrophils
• 4)Eosinophils
• 5)Basophils
• Agranulocytes (without granules)
• 6)Lymphocytes
• 7)Monocytes

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Components and General Properties of Blood
Monocyyte
Small lymphocyte
Neutrophil
Platelets
Eosinophil
Small lymphocyte
Erythrocyte
Young (band) neutrophil
Neutrophil
Monocyte
Large lymphocyte
Neutrophil
Basophil
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Leukocytes

• Least abundant formed element
• 5,000 to 10,000 WBCs/?L
• Protect against infectious microorganisms and
  other pathogens
• Conspicuous nucleus
• Spend only a few hours in the bloodstream before
  migrating to connective tissue
• Retain their organelles for protein synthesis
• Granules
• All WBCs have lysosomes called nonspecific
  (azurophilic) granules inconspicuous so
  cytoplasm looks clear
• Granulocytes have specific granules that contain
  enzymes and other chemicals employed in defense
  against pathogens

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Types of Leukocytes

• Granulocytes
• Neutrophils (60 to 70) polymorphonuclear
  leukocytes
• Barely visible granules in cytoplasm three- to
  five-lobed nucleus
• Eosinophils (2 to 4)
• Large red-orange granules
• Basophils (less than 1)
• Large, abundant, violet granules

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• Agranulocytes
• Lymphocytes (25 to 33)
• Variable amounts of bluish cytoplasm (scanty to
  abundant) ovoid/round, uniform dark violet
  nucleus
• Monocytes (3 to 8)
• Largest WBC generally ovoid, kidney-, or
  horseshoe-shaped nucleus

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Granulocytes

• Neutrophilsincreased numbers in bacterial
  infections
• Phagocytosis of bacteria
• Release antimicrobial chemicals
• Eosinophilsincreased numbers in parasitic
  infections, collagen diseases, allergies,
  diseases of spleen and CNS
• Phagocytosis of antigenantibody complexes,
  allergens, and inflammatory chemicals
• Release enzymes to destroy large parasites

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• Basophilsincreased numbers in chickenpox,
  sinusitis, diabetes
• Secrete histamine (vasodilator) speeds flow of
  blood to an injured area
• Secrete heparin (anticoagulant) promotes the
  mobility of other WBCs in the area

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Agranulocytes

• Lymphocytesincreased numbers in diverse
  infections and immune responses
• Destroy cells (cancer, foreign, and virally
  infected cells)
• Present antigens to activate other immune cells
• Coordinate actions of other immune cells
• Secrete antibodies and provide immune memory

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Agranulocytes

• Monocytesincreased numbers in viral infections
  and inflammation
• Leave bloodstream and transform into macrophages
• Phagocytize pathogens and debris
• Present antigens to activate other immune
  cellsantigen-presenting cells (APCs)

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The Leukocyte Life Cycle

• Leukopoiesisproduction of white blood cells
• Pluripotent stem cells (PPSCs)
• Myeloblastsform neutrophils, eosinophils,
  basophils
• Monoblastsform monocytes
• Lymphoblasts give rise to all forms of
  lymphocytes

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Leukopoiesis
Pluripotent stem cell
Colony-forming units (CFUs)
Precursor cells
Mature cells
leaves
Eosinophilic myelocyte
Eosinophilic promyelocyte
Eosinophilic myeloblast
Eosinophilic CFU
Eosinophil
Basophilic CFU
Basophilic myeloblast
Basophilic promyelocyte
Basophilic myelocyte
Basophil
Neutrophilic CFU
Neutrophilic myeloblast
Neutrophilic promyelocyte
Neutrophilic myelocyte
Neutrophil
Monoblast
Promonocyte
Monocyte
Monocytic CFU
B lymphocyte
B prolymphocyte
T lymphocyte
T prolymphocyte
Lymphoblast
Lymphocytic CFU
NK cell
NK prolymphocyte
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What Is Leukemia?

• Cancer of the white blood cells
• Acute or Chronic
• Affects ability to produce normal blood cells
• Bone marrow makes abnormally large number of
  immature white blood cells called blasts

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History

• Means white blood in Greek
• Discovered by Dr. Alfred Velpeau in France, 1827
• Named by pathologist Rudolf Virchow in Germany,
  1845

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Leukocyte Disorders

• Leukemiacancer of hemopoietic tissue that
  usually produces an extraordinary high number of
  circulating leukocytes and their precursors
• Myeloid leukemia uncontrolled granulocyte
  production
• Lymphoid leukemia uncontrolled lymphocyte or
  monocyte production

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Acute vs Chronic Leukemia

• Acute leukemia appears suddenly, progresses
  rapidly, death within months blasts found in
  peripheral blood
• Chronic leukemia undetected for months, survival
  time average of 3 years
• Effects normal cell percentages disrupted
  impaired clotting opportunistic infections

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Main Types

• Acute Lymphocytic Leukemia (ALL)
• Acute Myelogenous Leukemia (AML)
• Chronic Lymphocytic Leukemia (CLL)
• Chronic Myelogenous Leukemia (CML)

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Demographics of Leukemia Patients (2001 Data)
CLLChronic Lymphocytic ALLAcute
Lymphocytic CMLChronic Mylogenous AMLAcute
Mylogenous
Total Reported Cases 31,500
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Pictures Of Blood
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Development of Leukemia in the Bloodstream
Legend
White Cell
Red Cell
Platelet
Blast
Germ
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Causes

• High level radiation/toxin exposure
• Viruses
• Genes
• Chemicals
• Mostly unknown

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Signs and Symptoms of AML

• Insidious nonspecific onset
• Pallor due to anemia
• Febrile (fever) due to ineffective WBC
• Petechiae (skin bruising) due to thrombocytopenia
• Bone pain
• Petechiae

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Typical Labs of AML

• Leukocytosis
• Blastemia
• Leukemic hiatus
• Auer rods only found in myelocytic blasts
• Thrombocytopenia
• Anemia
• gt20 blasts in Bone Marrow

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Auer Rods
Auer Rod
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CD Markers

• The cluster of differentiation (cluster of
  designation) (often abbreviated as CD) is a
  protocol used for the identification and
  investigation of cell surface molecules providing
  targets for immunophenotyping of cells.
• The CD markers can be used to identify the type
  of cell.

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Other Findings

• CD 13 and CD 33 in flowcytometry
• Cytochemistries-stains that can be used to
  differentiate leukemias
• Myeloperoxidase
• Sudan black B
• Choloroacetate esterase (specific)
• Nonspecific esterase

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Flow Cytometry
Large clustering of CD 33s shows presence of
blasts
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FAB (1976) Classification

• M0 -- Undifferentiated AML
• M1 -- AML without maturation
• M2 -- AML with maturation
• M3 -- Acute Promyelocytic Leukemia
• M4 -- Acute Myelomonocytic Leukemia
• M5 -- Acute Monocytic Leukemia
• M6 -- Erythroleukemia (DiGuglielmos)
• M7 -- Megakaryoblastic Leukemia

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Myeloperoxidase (MPO)
p-Phenylene diamine Catecol H2O2
MPO gt Brown black deposits Brown deposits
considered to be a positive test-differentiates
AML from other leukemias
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M1 and M2
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M3
M4
M5
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Chloracetate (Specific) Esterase Myeloid Cell
Line
Naphthol-ASD-chloracetate CAE gt Free
naphthol compounds Stable diazonium salt (eg,
Fast Corinth) gt Red deposit
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Non-Specific Esterase Monocytic Line
? Naphthyl acetate ANAE gt Free naphthyl
compounds Stable diazonium salt (eg,
Fast blue RR) gt Brown deposits
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FAB vs WHO Classifications of Hematologic
Neoplasm

• FAB criteria
• Morphology
• Cytochemistry

• WHO criteria
• Morphology
• Immunophenotyping
• Genetic features
• Karyotyping
• Molecular testing
• Clinical features

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WHO Classification of AML

• AML with recurrent cytogenic translocations
• AML with multi-lineage dysplasia
• AML and myelodysplasia, therapy related
• AML, not otherwise categorized

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AML with Recurrent Cytogenetic Translocations
(WHO 1995)

• t(821) -- some maturation of neutrophilic line
  rare in older patients AML1/ETO fusion protein
  gt90 FAB M2
• t(1517) -- APL (granular and microgranular
  variants) retinoic acid receptor (RAR)
  leukemias middle aged adults DIC
• inv(16) or t(1616) -- monocytic and
  granulocytic abnormal eosinophilic component
• 11q23 -- monocytic children most common is
  t(911)

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Lymphocytic Leukemias

• Can involve T or B lymphocytes
• B lymphs mature in bone marrow
• Responsible for making antibodies
• T lymphs go to thymus to mature
• Are cytotoxic cells of immune system

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FAB Classification of ALL

• L1 Small homogeneous blasts mostly in children
• L2 Large heterogeneous blasts mostly in adults
• L3 Burkitt large basophilic B-cell blasts with
  vacuoles

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L3
L2
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Periodic Acid Schiff
Periodic acid Glycogen oxidation gt Aldehyde
Schiff reagent (para-rosaniline, Na
metabisulfite) gt Red deposit
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ALL Cytochemistries

• Oil Red O stains L3 vacuoles
• Terminal deoxynucleotidyl transferase (Tdt) DNA
  polymerase in early lymphoblasts
• Cell surface markers (CDs)
• Cytoplasmic and surface immunoglobulins B-cell
  line
• T-cell receptor (TCR)

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WHO Classification of Lymphoproliferative
Syndromes

• Precursor B Lymphoblastic Leukemia/Lymphoma
  (ALL/LBL) -- ALL in children (80-85 of childhood
  ALL) LBL in young adults and rare FAB L1 or L2
  blast morphology
• Precursor T ALL/LBL -- 15 of childhood ALL and
  25 of adult ALL
• Burkitt Leukemia/Lymphoma (FAB L3)

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Prognosis
Indicators Favorable Poor
WBC lt 50,000/?L ? 50,000/?L
Age 1 - 10 lt 1 or ?10
Gender Female Male
Blast B-cell T-cell and mixed
Karyotype Hyperploidy Trisomy 4, 10, 17 t(1221) (TEL/AML1) Hypoploidy Trisomy 5 t(119 (E2A/PBX1) Mixed lineage leukemia T(922) (Ph)
BM blast count during induction Mkd reduction at day 7 Mild reduction at day 7
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Burkitts Lymphoma
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CML
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Typical Labs in CML

• Leukocytosis with blastemia
• Thrombocytosis
• Basophilia
• Micro-megakaryocytes
• Low LAP score (intermediate if infected)
• About 10 blasts in BM
• Philadelphia chromosome

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Bone marrow aspirate and biopsy
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• Conventionally, a leukocytosis exceeding 50,000
  WBC/mm3 with a significant increase in early
  neutrophil precursors is referred to as a
  leukemoid reaction.

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• Serum leukocyte alkaline phosphatase is normal or
  elevated in leukemoid reaction, but is depressed
  in chronic myelogenous leukemia.

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• Leukemoid reactions are generally benign and are
  not dangerous in and of themselves, although they
  are often a response to a significant disease
  state

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• Historically, various clues including the
  leukocyte alkaline phosphatase score and the
  presence of basophilia were used to distinguish
  CML from a leukemoid reaction. However, at
  present the test of choice in adults to
  distinguish CML is an assay for the presence of
  the Philadelphia chromosome, either via
  cytogenetics and FISH, or via PCR for the BCR/ABL
  fusion gene.

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Leukocyte Alkaline Phosphatase (LAP)
Naphthol AS-MX phosphate LAP at
pH8.6 gt Naphthol AS-MX Diazonium
salt (eg, Fast blue RR) gt
Insoluble pigment
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LAP Score

• Count 100 consecutive segmented neutrophils and
  bands
• Score
• 0 no granules
• 1 occasional diffuse granules
• 2 moderate number of granules
• 3 many strongly positive granules
• 4 confluent strongly positive granules

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0
1
2
3
4
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LAP Score
Example 0 x 35 cells 0 1 x 30 cells
30 2 x 20 cells 40 3 x 10 cells 30 4 x 5
cells 20 120 LAP Score
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Philadelphia Chromosome

• 9 22 translocation almost specific to CML
• Karyotype to visualize Ph chromosome
• Produces BCR/c-abl fusion oncogene
• Gene product p190 is a hyperactive tyrosine
  kinase
• Ph chromosome seen in ALL produces p210 and
  chronic neutrophilic leukemia produces p230

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Karyotype 46,XX,t(922)(q34q11.2) -- Ph
chromosome
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FISH showing the BCR (green), ABL (orange), and
BCR-ABL fusion signals (arrow) Apositive
(contains a residual ABL signal), Bnormal
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Chronic Lymphocytic Leukemia

• Exclusive in elderly
• Lymphocytosis unrelated to viral infection
• Hyper-mature lymphocytes with highly condensed
  nuclei
• Smudge cells

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CLL PB and BM
Smudge cell
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Bone Marrow