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acute leukemias for undergraduates

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  2. 2. Leukemoid Reaction• A leukemoid reaction describes a highWBC count with neutrophilia,usually inresponse to infection.• The WBC count may be as high as 50,000/microL and can easily mimic CML orAML.
  3. 3. • Serum leukocyte alkaline phosphatase isnormal or elevated in leukemoid reaction,but is depressed in chronic myelogenousleukemia.• The bone marrow in a leukemoid reaction,if examined, may be hypercellular but isotherwise typically unremarkable.
  4. 4. Features Suggesting LeukemoidReaction• Toxic granulation.• High LAP score.• Presence of an obvious cause for theneutrophilia.
  5. 5. • As noted above, a leukemoid reaction is typically a response to an underlying medical issue. Causes of leukemoid reactions include:• Hemorrhage• Drugs– Use of Sulfa drugs– Use of Dapsone– Use of glucocorticoids– Use of G-CSF or related growth factors– All-trans retinoic acid (ATRA)• Infections– Clostridium difficile– Tuberculosis– Pertussis– Infectious mononucleosis (lymphocyte predominant)– Visceral Larva Migrans (eosinophil predominant)• Asplenia• Diabetic ketoacidosis• Organ necrosis– Hepatic necrosis– Ischemic colitis• As a feature of Trisomy 21 in infancy (incidence of ~10%)• As a paraneoplastic phenomenon (rare)
  6. 6. Leukemoid reaction
  7. 7. LAP score
  8. 8. What is leukemia?Leukemia is cancer of the white blood cells. Whiteblood cells help your body fight infection. Yourblood cells form in your bone marrow. In leukemia,however, the bone marrow produces abnormalwhite blood cells. These cells crowd out the healthyblood cells, making it hard for blood to do its work.
  9. 9. Hematopoieticstem cellNeutrophilsEosinophilsBasophilsMonocytesPlateletsRed cellsMyeloidprogenitorLymphoidprogenitorB-lymphocytesT-lymphocytesPlasmacellsnaïveALLAML
  10. 10. Myeloid maturationmyeloblastpromyelocytemyelocytemetamyelocyteband neutrophilMATURATIONAdapted and modified from U Va website
  11. 11. Definition• Leukemia: is a cancer of the blood or bone marrowcharacterized by abnormal proliferation of bloodcells,usually WBCs(Leukocytes).• Acute leukemia: rapid increase of immature bloodcells.• Chronic leukemia: excessive build up of relativelymature,but still abnormal blood cells.
  12. 12. About the Disease• Leukemia, lymphoma and myeloma arecancers that originate in the bone marrow(leukemia & myeloma) or in lymphatictissues (lymphoma).
  13. 13. Different Types of BloodCancers• Leukemia• Non-Hodgkin Lymphoma• Hodgkin Lymphoma• Myeloma• Myelodysplastic Syndromes
  14. 14. What is Leukemia• Greek word which means “white blood”• Leukemia is when cells spread rapidly and destroy livingtissue.• It grows/invades the bone marrow which is the factory ofblood and replaces normal blood elements with cancercells.• Cancer cells replace all bone marrow cells which causesinfection and bleeding problems.• Leukemia is basically white blood cells that don‟t workwell and cause trouble.
  15. 15. Signs and Symptoms• Most of the signs and symptoms are due to:1-Anemia.2-Leukopenia.3-Thrombocytopenia.• Bicytopenia,Pancytopenia.• All symptoms associated with leukemiacan be attributed to other diseases,consequently,leukemia is alwaysdiagnosed by laboratory investigations.
  16. 16. Causes• Leukemia,like other malignancies, resultsfrom somatic mutations in the DNA.• Certain mutations produce leukemia byactivating oncogenes or deactivatingtumor suppressor genes.• These mutations may occur spontaneouslyor as a result of exposure to radiationor carcinogenic substances,and likelyto be influenced by genetic factors.
  17. 17. Causes-cont‟d• Ionizing radiation• Viruses: Human T-lymphotropic virus (HTLV-1)• Chemicals: Benzene,chemotherapy.• Smoking: slight increase in leukemiaincidence.• Genetic predisposition toward developingleukemia: Down syn.,Fanconi anemia
  18. 18. Classification• Multiple classification systems.• FAB classification:French-American-British Classification.• FAB Classification relies on morphologic,cytochemical,and immunophenotypingcriteria to define 8 major subtypes(M0-M7)
  19. 19. FAB vs WHO Classification• French-American-British (FAB) Cx– Cellular morphology and cytochemical stain– Acute leukemia as > 30% bone marrow blasts– Widely used• World Health Organization Cx– Cellular morphology and cytochemical stain– Immunologic probes of cell markers, cytogenetics,molecular abnormalities & clinical syndrome– Acute leukemia as > 20% bone marrow blasts– Standard for diagnosis
  20. 20. Acute myeloid leukemias (AML)Classification - FAB1. M0: minimally differentiated2. M1: myeloblastic leukemia without maturation3. M2: myeloblastic leukemia with maturation4. M3: hypergranular promyelocytic leukemia5. M4: myelomonocytic leukemia6. M4Eo: variant, increase in marrow eosinophils7. M5: monocytic leukemia8. M6: erythroleukemia (DiGuglielmos disease)9. M7: megakaryoblastic leukemia
  21. 21. AML classification - WHOAML not otherwise categorized1. AML minimally differentiated2. AML without maturation3. AML with maturation4. Acute myelomonocytic leukemia5. Acute monocytic leukemia6. Acute erythroid leukemia7. Acute megakaryocytic leukemia8. Acute basophilic leukemia9. Acute panmyelosis with myelofibrosis
  22. 22. Acute vs Chronic LeukemiaAcute ChronicAge Children & youngadultsMiddle age andelderlyOnset Sudden insidiousDuration weeks to months yearsWBC count Variable High
  23. 23. Acute vs Chronic LeukemiaAcute ChronicPlatelets Low Early: Normal/ HighLate: LowAnemia High (>90%) None/ mildPredomi-nant cellsBlast cells Mature cellsAML = myeloblastALL= lymphoblastCML=granulocytesCLL=lymphocytes
  24. 24. Acute vs Chronic LeukemiaAcute ChronicMarrowcellularity>20% marrowblasts (WHO)> 30% marrowblasts (FAB)>70% marrowcellularity(hypercellular);No dysplasiaDiagnosis PBS, BM exam,cytochemicalstains, surfacemarkers,EM,chromosomePBS (peripheralblood smear)
  25. 25. Acute Myeloid Leukemia
  26. 26. Definition• Acute myeloid leukemia (AML): acutemyelogenous leukemia,acute non-lymphocytic leukemia.• AML consists of a group of relatively well-defined hematopoietic neoplasmsinvolving precursor cells commitedto the myeloid line(WBCs,RBCs,PLTs)
  27. 27. Chracteristics• AML is characterized by a clonal proli-feration of myeloid precursors with areduced capacity to differentiate intomature cellular elements.• As a result,there is an accumulation ofleukemic blasts or other immatureforms in the BM,peripheral blood,andother tissues with a variable reduction inthe production of normal RBCs,platelets,and mature granulocytes.
  28. 28. Specific:• M2 : Chloroma:-presents as a mass lesion „tumorof leukemic cells‟• M3 : DIC• M4/M5 : Infiltration of soft tissues,gum infiltration, skin deposits ,Meningealinvolvement-headache, vomiting, eye symptoms
  29. 29. ABCChloromasNEJM 1998
  30. 30. Gum hypertrophy
  31. 31. Leukostasis• accumulation of blasts in microcirculationwith impaired perfusion• lungs: hypoxemia, pulmonary infiltrates• CNS: stroke• only seen with WBC >> 50 x 109/L
  32. 32. Pathological Features• CBC and differential.• Blood film (smear).• Bone marrow examination: BM aspirateand trephine biopsy.1-Morphology.2-Immunephenotyping.3-Cytogenetics and molecular biology.
  33. 33. Jemshidi trephine &Salah aspiration needle
  34. 34. Bone marrow in acute leukemia• necessary for diagnosis• useful for determining type• useful for prognosis• Acute leukemias are defined by thepresence of > 20% blasts in bone marrow(% of nucleated marrow cells)
  35. 35. • Bone marrow aspirate & trephine:Hypercellular,– blast cells ( > 20%),– presence of Auer rods - AML type• Cytochemistry :Special stains to differentiate AML fromALL ;Positivity with Sudan black &Myeloperoxidase (MPO) in AML
  36. 36. WBC Count in AML• WBC count in AML can be high,normal,orlow.• Median WBC count in AML is 15 000/uL.• 20% of patients have > 100 000/uL• 25-40% of patients have <5000/uL• 95% of patients have blast cells on bloodfilm.
  37. 37. Distinguishing AML from ALL• light microscopy– AML: Auer rods, cytoplasmic granules– ALL: no Auer rods or granules.• flow cytometry• special stains (cytochemistry)
  38. 38. Cytochemical Stains• Since the early 20th century, cytochemical staining ofcells has been a useful tool in differentiatinghematopoietic diseases.• Smears and imprints made from bone marrow, lymphnodes, spleen, or peripheral blood are preferred.– In enzymatic techniques, fresh smears are used toensure optimal enzyme activity• Certain elements may be inhibited during the fixationof smears and imprints
  39. 39. Myeloperoxidase (MPX/MPO)• The proxidase enzyme reacts with H2O2 & release O2,which oxidizes the indicator dye and produce orange-brown granules in the cells (3-amino-9-erythrocarbazol)• Enzyme MPX is found in the 1o granules ofgranulocytes, neutrophils and precursors (from thepromyelocyte stage on) & eosinophils• Monocytes may be weakly pos• Leukemic myeloblasts are usually pos and Auer rodsstain very strongly• Used for differentiating AML (+) from ALL (-)• Normal bone marrow smear <5 days old used for controlslides (promyelocyte - neutrophils)
  40. 40. MPO (right) & Sudan black (left)showing intense localised positivity inblasts
  41. 41. Myeloperoxidase(MPO)p-Phenylene diamine + Catecol + H2O2MPO > Brown black deposits
  42. 42. Chloracetate (Specific) EsteraseMyeloid Cell LineNaphthol-ASD-chloracetateCAE > Free naphthol compounds+ Stable diazonium salt (eg, Fast Corinth)> Red deposit
  43. 43. Non-Specific Esterase (NSE)• Nonspecific esterase liberates alpha-naphthyl fromthe substrate alpha-naphthyl acetate. Alpha-naphthyl is couples with the dye molecule to formdark reddish-brown granules• Monocytes, monblasts, macrophages, histiocytes,megakaryocytes and some carcinomas are NSE pos• Abnormal erythroblasts are strongly pos• Lymphocytes are neg or may show dot positivity
  44. 44. Non-Specific EsteraseMonocytic LineNaphthyl acetateANAE > Free naphthyl compounds+Stable diazonium salt (eg, Fast blue RR)> Brown deposits
  45. 45. NSE continued• Used for differentiating myelomonocytic andmonocytic leukemia (+) from granulocytic leukemia (-)• Monocyte NSE are fluoride sensitive• Peripheral smear with appreciable # of monocytes or anormal BM smear used for control slideshttp://www.healthsystem.virginia.edu/internet/hematology/hessedd/malignanthematologicdisorders/leukemias/aml-m4.cfm
  46. 46. Double Esterase in M4NSE with Fl inhibitionHistiocyte
  47. 47. Periodic Acid Schiff (PAS)• Periodic acid oxidizes glycogen, mucoproteins, andother high-molecular weight carbohydrates toaldehydes.• Aldehydes react with colorless Schiff reagent,staining them a bright red-pink• Megakaryocytes and platelets stain strongly pos• Normoblasts will stain Pos• Lymphoblasts in ALL show course and granular(block) positivity
  48. 48. PAS Continued• Myeloblasts are Neg• Aids in diagnosis of ALL, erythroleukemia, andmegakaryoblastic leukemia• Normal bone marrow smear used for control slideshttp://www.pathologyoutlines.com/leukemia.html
  49. 49. Periodic Acid SchiffPeriodic acid + Glycogenoxidation > Aldehyde + Schiff reagent(para-rosaniline, Na metabisulfite)> Red deposit
  50. 50. AML
  51. 51. AML
  52. 52. Auer rods in AML
  53. 53. ALL
  54. 54. P. Smear AML
  55. 55. MO: Minimally differentiated• Undifferentiated Blasts(No maturation)• Myeloid phenotype -CD13, CD33, CD34• (-) SBB, MPO• Negative: Auer rods,Esterase
  56. 56. M1 AML without maturation > 30% myeloblasts Large cells, round nucleus Nucleoli (+) scanty cytoplasm >3% MPO, SBB (+) <20% NSE (+) CD 13, 33, 117
  57. 57. M1 AML without maturation
  58. 58. M2 AML with maturation• Common type• >30% myeloblasts• >10% granulocyte• Kidney shape nucleus• Nucleoli (+)• (+) Auer rods• Eosinophilic granules• >50% MPO, SBB (+)• CD 13, 33
  59. 59. M2 AML with maturation
  60. 60. Auer Rods
  61. 61. M3 (hypergranular promyelocytic)• Promyelocyte-predominant• Large, kidney shape• (+) Auer rods (faggot cells)• basophilic, bilobed nuclei• CD 13,33• High incidence of DIC
  62. 62. Acute myeloid leukemia with veryabnormal cells (AML M3/ t15;17)
  63. 63. M4 Acute myelomonocytic• >30% myeloblast (FAB)• >20% granulocyte• >20% promonocytes andmonocytes• CD 11, 13, 33,14• (+) Auer rods common• High serum lysozyme level– M4Eo = w/ eosinophilia
  64. 64. M3M5M4
  65. 65. M5: acute monocytic leukemia1. M5a – without maturation– Monoblasts , few promonocytes2. M5b – with maturation– Blast, Promonocytes (BM), Monocytes(Blood)
  66. 66. M5a• Monoblast ameboid withround to oval nuclei,• prominent nucleoli,• <20% promonocytes/mono• Vacuolated cytoplasm
  67. 67. AML M5a
  68. 68. M5b• > 20% promonocytes,monocytes• Promonocytes folded,convulated nucleus• Azurophilic granules
  69. 69. AML M5b
  70. 70. M6 - erythroleukemia Large, bizarre,round-to-oval cells (+) nucleoli > 50% Erythroblasts > 30 % Myeloblasts CD 45,71 Glycophorin A CD 13, 15,33 myeloblast PAS (+)
  71. 71. M6 (erythroblast)
  72. 72. M6 (erythroblast)
  73. 73. M7 – acute megakaryoblastic• >30% megakaryoblasts• platelet like granules onPAS stain• NSE (but not BE) (+)• Myeloid blasts may showSBB or MPO (+)• CD 41,42,61
  74. 74. M7 (Megakaryoblast)
  75. 75. Megakaryoblast
  76. 76. Acute Nonlymphocytic Leukemias
  77. 77. • Acute lymphoblastic leukemias
  78. 78. FAB Classification of ALLL1: Small homogeneous blasts; mostly inchildrenL2: Large heterogeneous blasts; mostly inadultsL3: “Burkitt” large basophilic B-cell blastswith vacuoles
  79. 79. WHO Classification ofLymphoproliferative Syndromes Precursor B Lymphoblastic Leukemia/Lymphoma(ALL/LBL) -- ALL in children (80-85% ofchildhood ALL); LBL in young adults and rare;FAB L1 or L2 blast morphology Precursor T ALL/LBL -- 15% of childhood ALLand 25% of adult ALL Burkitt Leukemia/Lymphoma (FAB L3)
  80. 80. • Confirmation:– Immunophenotyping– Molecular genetics– Cytogenetics: Chromosomal abnormalities
  81. 81. ALL L1
  82. 82. L3L2
  83. 83. Burkits / ALL L3
  84. 84. Prognostic factors• High WBC – relapse in testis /cns• Infants <1 yr, children >10 yr poor outcome• L1- good prognosis• L2,3- bad prognosis
  85. 85. Prognosis• The response to treatment and overallsurvival of patients with AML areheterogenous.• Prognostic factors are related to patientand tumor characteristics:1-Age2-Performance status3- Karyotype
  86. 86. Adverse Clinical Predictors• Advanced age.• Poor performance status.• History of exposure to cytostatic agents orradiotherapy.(Therapy-related AML).• History of MDS or other hematologicaldiseases
  87. 87. THANK YOU