Leukemia – Classification, clinical manifestation, pathology & diagnosis – AML+CML

1. 7/5/15 Leukemia – Classification, clinical manifestation, pathology & diagnosis – AML+CML Dr. Ksheera Cariappa Assistant professor

2. CONTENT • GENERAL – Definition, classification , pathogenesis and diagnosis • AML • CML

3. ACUTE MYELOID LEUKEMIA

4. Acute Myeloid Leukemia • What is it? - Clonal expansion of myeloid precursor cells with reduced capacity to differentiate - As opposed to ALL/CLL, it is limited to the myeloid cell line • differentiated from ALL based on morphology, cytogenetics, cytochemical analysis, cell surface markers

5. Epidemiology • Incidence -3.5 per 100,000 people per year • Median age at diagnosis- 67 years • AML incidence increases with age

6. ALL naïve germinal center B-lymphocytes Plasma cells Lymphoid progenitor T-lymphocytes Neutrophils AML Myeloid progenitor Eosinophils Hematopoietic stem cell Basophils Monocytes Platelets Red cells

7. myeloblast promyelocyte myelocyte metamyelocyte band neutrophil Myeloid maturation MATURATION Adapted and modified from U Va website

8. Acute Leukemia • Blasts in the marrow

9. Two-hit model of leukemogenesis Loss of function of transcription factors needed for differentiation eg. AML1-ETO CBFb-SMMHC PML-RARa Gain of function mutations of tyrosine kinases eg. FLT3, c-KIT mutations N- and K-RAS mutations BCR-ABL TEL-PDGFbR differentiation block enhanced proliferation Acute Leukemia +

10. Etiology • Heredity • Down syndrome • Fanconi anemia • Bloom syndrome • ataxia-telangiectasia • Congenital neutropenia (Kostmann syndrome) • Radiation • High-dose radiation (atomic bombs survivors)

11. chemical and occupational exposures • Benzene • petroleum products • Paint • embalming fluids • ethylene oxide • Herbicides • smoking

12. Prospective data suggested an elevated risk of myeloid leukemia associated with cigarette smoking (relative risk, 1.4; 95% confidence interval, 1.2 to 1.6). •  Population-attributable risk calculations suggested that approximately 14% of all US leukemia cases (including 17% of myeloid may be due to cigarette smoking.

13. Drugs • Alkylating agent • Topoisomerase II inhibitor • Chloramphenicol • Phenylbutazone • Chloroquine • methoxypsoralen

14. FAB CLASSIFICATION • FAB classification for AML: M₀ - minimally differentiated leukaemia M₁ - myeloblastic leukaemia without maturation M₂ - myeloblastic leukaemia with maturation M₃ -hypergranular promyelocytic leukaemia -hypo or microgranular promyelocytic leukaemia (M₃ variant)

15. M₄ - myelomonocytic leukaemia -myelomonocytic leukaemia with bone marrow eosinophilia (M₄E₀) M₅ - monocytic leukaemia - Undifferentiated (monoblastic)(M5a) - well-differentiated (promonocytic- monocytic)(M5b) M₆ - Erythroleukaemia M₇ - megakaryocytic leukaemia

16. WHO classification of AML 1. AML with recurrent genetic abnormalities: • AML with t(8;21)(q22;q22);(AML1/ETO) • AML with abnormal bone marrow eosinophils inv(16)(p13q22) or t(16;16)(p13;q22);(CBFβ/MYH11) • Acute promyelocytic leukaemia (AML with t(15;17)(q22;q12)(PML/RARα) and variants • AML with 11q23(MLL)abnormalities.

17. 2. AML with multilineage dysplasia • Following a myelodysplastic syndrome • Without antecedent myelodysplatic syndrome 3. AML and MDS ,therapy related • Alkylating agent-related • TopoisomerasetypeII inhibitor-related • Other types

18. 4. AML not otherwise specified • AML minimally differentiated • AML without maturation • AML with maturation • Acute myelomonocytic leukaemia • Acute monoblastic and monocytic leukaemia • Acute erythroid leukaemia • Acute megakaryoblastic leukaemia • Acute basophilic leukaemia • Acute panmyelosis with myelofibrosis • Myeloid sarcoma

19. 5. Myeloid proliferations related to Down’s Syndrome • Transient abnormal myelopoiesis - TAM –DS newborns Blasts Megakaryocytic lineage • Myeloid leukaemia associated with down syndrome - DS have 50 fold increase incidence during 5 yrs 6. Blastic plasmacytoid dendritic cell neoplasm • Blastic NK cell lymphoma , agranular CD4+ natural killer leukaemia 7. Acute leukemia of ambiguous lineage

20. Diagnosis of acute leukaemias • >20% blasts in the bone marrow • Morphology • Cytochemistry • Immunophenotyping • Cytogenetics • Molecular genetics

21. Morphology Blast cells are large cells with : • High N:C ratio • Nucleus is large with open chromatin • Nucleoli- 1- 5 • Thin rim to moderate amount of cytoplasm

24. Acute myeloblastic leukaemia, minimally differentiated (FAB synonym-AML M0): Appear undifferentiated by light microscopy. AML-M0

25. AML-M1 • Acute myeloblasticleukaemia without maturation (FAB synonym-AML M1): • There is no or minimal maturation along granulocytic pathway. • Cell population is predominantly composed of myeloblasts.

26. There are two types of blasts: one type does not show any cytoplasmicazurophil granules, while other show a few granules. • Cytochemically >3% of blasts are peroxidase positive. • Immunologic cell marker studies reveal expression of at least two myeloid antigens ( CD13, CD33, CD117, MPO)

28. AML-M2 • Acute myeloblasticleukaemia with maturation (FAB synonym-AML M2): • This is the most frequent subtype (30-45%). • There is clear evidence of maturation to promyelocyte stage and beyond. • Auer rods, which represent aggregates of azurophilic granules in lysosomes, are commonly seen.

29. Mature cells are >10%. • Monocytic cells should be less than <20%. Some cases of AML M2 show increased basophils. • Myeloperoxidase reaction is strongly positive.

32. AML-M3 • Acute promyelocytic leukaemia (FAB-AML-M3): AML- M3 constitutes 5-10% of all cases of AML • Morphology: Leukaemic cells are hypergranular; granules obscure the nuclei of cells; with some cells showing bundles of Auer rods(faggot cells). M3 variant is the microgranular/ hypogranular form. Both types are peroxidase positive.

33. Cytogenetics: Majority of the M3 cases demonstrate a reciprocal translocation involving chromosomes 15 and 17, t(15;17) • Bleeding diathesis: Granules of the promyelocytes contain thromboplastin like material resulting in widespread disseminated intravascular coagulation, DIC with profound bleeding manifestations

35. AML-M4 • Acute myelomonocyticleukaemia (FAB synonym-AML M4): • Along with granulocytic series a significant proportion of cells of monocytic series are also present. • In bone marrow blasts are >20% and monocytic cells and their precursors and neutrophils and their precursors each are >20%.

36. Leukaemic cells express myeloid associated antigens (CD13, CD33) and markers of monocytic differentiation (CD14, lysozyme)

38. Acute myelomonocyticleukaemia with eosinophils (AML-M4E0) • Morphology: This subtype of M4 is characterised by the presence of eosinophil precursors which demonstrate eosinophilic and basophilic granulation and their nuclei may be monocytoid.

39. Cytogenetics: Majority of these cases are associated with inversion of chromosome 16. • Clinical features: there is skin and CNS infiltration of leukaemic cells.

41. AML-M5 • Acute monoblasticleukaemia and acute monocyticleukaemia(FAB synonym-AML M5): • In this type monocytic differentiation is readily apparent with 80% or more of leukaemic cells in bone marrow being monocytic.

42. In acute monoblasticleukaemia 80% or more of all monocytic cells are monoblasts. • In monocyticleukaemia predominant monocytic cells are promonocytes.

44. AML-M6 • Acute erythroidleukaemia( FAB synonym AML M6): • Characterised by predominant population of erythroblasts. • Two subtypes: A)Erythroleukaemia (>20% of non erythroid cells are myeloblasts and >50% of all nucleated cells are erythroblasts.B)Pure erythroidleukaemia (>80% of marrow cells are erythroblasts with no significant myeloid component)

45. In erythroleukaemia the erythrocytes are bizzare looking with bi and tri nucleate forms and megaloblastic nuclear features. • Cytoplasmic vacuoles are commonly seen. Erythroblasts are commonly observed in peripheral blood. • PAS stain is positive in erythroblasts.

47. AML-M7 • Acute megakaryoblasticleukaemia( FAB synonym AML M7): • Megakaryoblasts are more than 20% in bone marrow. • Characterised by marked bone marrow fibrosis. • In peripheral blood fragments of megakaryoblasts, micromegakaryocytes or dysplastic large platelets are seen.

48. Cytochemically blast cells are negative for MPO. • Immunological markers for platelet glycoprotein IIb/IIIa and factor VIII related antigen are positive. • The megakaryoblasts express CD41(glycoprotein IIb/IIIa) and/or CD61(glycoprotein IIIa)

50. Acute basophilic leukaemia: Very rare type with basophilic differentiation. Blasts contain basophilic granules and characteristically show positive metachromatic staining with toluidine blue.