2Leukaemia is a disease resulting from the neoplastic proliferation of haemopoietic or lymphoid cells.Leukaemias are broadly divided into:Acute leukaemias, which, if untreated, lead to death in weeks or months.chronic leukaemias, which, if untreated, lead to death in months or years.
3They are further divided into lymphoid, myeloid and biphenotypic leukaemias, the latter showing both lymphoid and myeloid differentiation.Acute leukaemias are characterized by a defect in maturation, leading to an imbalance between proliferation and maturation; since cells of the leukaemic clone continue to proliferate without maturing to end cells.
4Diagnosis of leukaemia. The diagnosis of leukaemia and categorizationrequired the following parameters.1- Morphology.2-Cytochimestry3-Immunophenotyping.4-Cytogenetic.5- Molecular study.
6BackgroundAcute lymphoblastic leukemia (ALL) is the most common malignancy diagnosed in children, representing nearly one third of all pediatric cancers.The annual incidence rate for acute lymphoblastic leukemia is 30.9 cases per million population. The peak incidence occurs in children aged 2-5 years.
7Pathophysiology In acute lymphoblastic leukemia, a lymphoid progenitor cell becomes genetically alteredand subsequently undergoes dysregulatedproliferation, survival, and clonal expansion.In most cases, the pathophysiology of transformedlymphoid cells reflects the altered expression ofgenes whose products contribute to the normaldevelopment of B cells and T cells
8Clinical feature.Children with acute lymphoblastic leukemia (ALL) generallyPresent with signs and symptoms that reflect bone marrowinfiltration and extramedullary disease.Because leukemic blasts replace the bone marrow, patientsPresent with signs of bone marrow failure, including anemia,thrombocytopenia, and neutropenia.Clinical manifestations include fatigue and pallor, petechiae andbleeding, and fever.In addition, leukemic spread may manifest as lymphadenopathyAnd hepatosplenomegaly. Other signs and symptoms of leukemiaincluding weight loss, bone pain, and dyspnea.
9The classification of ALL FAB classification.L1 ALL L2 ALL L3 ALLCell size Mainly small Large, heterogeneous Large, homogeneousNuclear chromatin Fairly homogeneous Heterogeneous Finely stippled,Nuclear shape Mainly regular Irregular; clefting RegularNucleolus Not visible Usually visible Usually prominentAmount of cytoplasm Scanty Variable abundant Moderately abundantCytoplasmic basophilia Slight to moderate Variable StrongCytoplasmic vacuolation Variable Variable Often prominent
14Clinical correlates of FAB categories of ALL Many cases of L3 ALL represent a distinct entity thatrequires specific management. However, the categorizationof a case as L1 or L2 ALL is of little importance.The FAB L1 category includes more childhoodcases with a relatively good prognosis.The incidence of ALL L1 falls with increasing age whereasthe incidence of ALL L2 does not vary much with age.ALL L2 has generally been found to have a worseprognosis, although the difference is not major.
15WHO proposed classification of acute lymphoblastic leukemia The recent WHO International panel on ALL recommends thatthe FAB classification be abandoned, since the morphologicalclassification has no clinical or prognostic relevance.1- Acute lymphoblastic leukemia/lymphoma Synonyms:Former Fab L1/L2i. Precursor B acute lymphoblastic leukemia/lymphoma.Cytogenetic subtypes:t(12;21)(p12,q22) TEL/AML-1t(1;19)(q23;p13) PBX/E2At(9;22)(q34;q11) ABL/BCRT(V,11)(V;q23) V/MLLii. Precursor T acute lymphoblastic leukemia/lymphoma2- Burkett's leukemia/lymphoma Synonyms: Former FAB L33- Biphenotypic acute leukemia
17Characterization of the Immunophenotyping is referred to as Immunophenotyping and is achieved by means of labeled antibodies that recognize specific epitopes of cellular antigens.In general, the most useful antibodies are monoclonal antibodies (McAb) produced by hybridoma technology but, for some antigens, polyclonal antibodies (PcAb) (antisera) are better.The technique employed for Immunophenotyping may be immunocytochemistry or, much more often, flow cytometry.Immunophenotyping is essential for the diagnosis of B- or T-lineage acute lymphoblastic leukaemia (ALL).
18First panel Second panel B lymphoid CD19, CD22, CD79a, CD10 T lymphoid CD3, CD2, CD7Second panelIf B lineage cm, k, l, CD20, CD24If T lineage CD1a, SmCD3, CD4, CD5, CD8, anti-TCR ab, anti-TCR gd
20Cytogenetic study.With techniques now available, 70–90% of cases of ALLhave a demonstrable cytogenetic abnormality.In ALL, chromosomal abnormalities correlatewith other clinical and hematological factorsof prognostic importance but they also havea considerable independent prognosticsignificance.
21B-lineage ALL L1 high/ hyperdiploidy. L1 or L2/t(9;22)/BCR-ABL fusion L1 or L2/t(4;11)(q21;q23)L1 or L2/t(12;21)(p12;q22)/early precursor or common ALLL1 or L2/t(1;19)(q23;p13)/pre-B ALL
22T-lineage ALL. L1 or L2/t(10;14)(q24;q11) Burkett's-lineage L3/t(8;14)(q24;q32) or t(8;22)(q24;q11) ort(2;8)(p12;q24).
24Distinguishing between AML and ALL Correct assignment of patients to the categorizeAML and ALL is very important for prognosisand choice of therapy.The FAB group recommended the use ofMPO,SBB and non-specific esterase (NSE)stains.If Cytochemical reactions for myeloid cells arenegative, presumptive diagnosis of ALL mustbe confirmed Immunophenotyping.
25Background.AML is the most common acute leukaemia affecting adults, and its incidence increases with age.Although AML is a relatively rare disease, accounting for approximately 1.2% of cancer deaths in the United States, its incidence is expected to increase as the population ages.
26The malignant cell in AML is the myeloblast. Pathophysiology.The malignant cell in AML is the myeloblast.In normal haematopoiesis, the myeloblast is an immature precursor of myeloid white blood cells; a normal myeloblast will gradually mature into a mature white blood cell.However, in AML, a single myeloblast accumulates genetic changes which "freeze" the cell in its immature state and prevent differentiation Such a mutation alone does not cause leukemia; however, when such a "different combined with other maturation which disrupt genes controlling proliferation, the result is the uncontrolled growth of an immature clone of cells, leading to the clinical entity of AML.
27Clinical feature.The symptoms of AML are caused by replacement of normal bone marrow with leukemic cells, which causes a drop in red blood cells, platelets, and normal white blood cells.These symptoms include fatigue, shortness of breath, easy bruising and bleeding, and increased risk of infection
28M0 Undifferentiated acute myeloblastic leukemia. The classification of AMLFAB classification.M0 Undifferentiated acute myeloblastic leukemia.M1 Acute myeloblastic leukemia with minimal maturation.M2 Acute myeloblastic leukemia with maturation.M3 Acute promyelocytic leukemia.M4 Acute myelomonocytic leukemia.M4 eosAcute myelomonocytic leukemia with eosinophilia.Acute monocytic leukemia.M6 Acute erythroid leukemia.M7 Acute megakaryoblastic leukemia.
29Criteria for the diagnosis of acute myeloid leukaemia of M0 Blasts .30% of bone marrow nucleated cellsBlasts .30% of bone marrow non-erythroid cells <3%of blasts positive for Sudan black B or for myeloperoxidaseby light microscopy.Blasts demonstrated to be myeloblasts by immunologicalmarkers or by ultrastructural cytochemistry.
31Criteria for the diagnosis of acute myeloid leukaemia of M1. Blasts 30% of bone marrow cells .Blasts .90% of bone marrownon-erythroid cells .3% of blasts positive for peroxidase orSudan black BBone marrow maturing monocytic component (promonocytes tomonocytes) .10% of non-erythroid cellsBone marrow maturing granulocytic component (promyelocytes topolymorphonuclear leucocytes) .10% of non-erythroid cells
33Criteria for the diagnosis of acute myeloid leukaemia of M2. Blasts 30% of bone marrow cells. Blasts 30–89% of bone marrownon-erythroid cellsBone marrow maturing granulocytic component (promyelocytes topolymorphonuclear leucocytes) >10% of non-erythroid cellsBone marrow monocytic component (monoblasts to monocytes)<20% of non-erythroid cells and other criteria for M4 not met
35Acute hypergranular promyelocytic Leukaemia M3 AML In acute hypergranular promyelocyticleukaemia the predominant cell is a highlyabnormal promyelocyte.In the majority of cases, blasts are fewer than30% of bone marrow nucleated cells. Thedistinctive cytological features are sufficient to permit a diagnosis and
36In some cases there are giant granules or multiple Auer rods, which are often presentin sheaves or ‘faggots’. Most cases have a minority of cells that are agranular.M3 AML has been found to be very sensitiveto the differentiating capacity of all-trans-retinoic acid (ATRA). Following such therapyan increasing proportion of cells beyond thepromyelocyte stage are apparent.
39Criteria for the diagnosis of acute myeloid leukaemia of M4. Blasts .30% of bone marrow cellsBlasts .30% of bone marrow non-erythroid cellsBone marrow granulocytic component 20% of non-erythroid cellsSignificant monocytic component as shown by one of the following:Bone marrow monocytic component 20% of non-erythroid cells and peripheral blood monocytic.Bone marrow resembling M2 but peripheral blood monocytic component .5000/cumm.
41Criteria for the diagnosis of acute myeloid leukaemia of M5 Blasts .30% of bone marrow cellsBlasts .30% of bone marrow non-erythroid cellsBone marrow monocytic component .80% of non-erythroid cellsAcute monoblastic leukaemia (M5a)Monoblasts .80% of bone marrow monocytic componentAcute monocytic leukaemia (M5b)Monoblasts <80% bone marrow monocytic component
45Criteria for the diagnosis of acute myeloid leukaemia of M7 Blasts 30% of bone marrow nucleated cells.Blasts demonstrated to be megakaryoblasts byimmunological markers, ultrastructural examinationor ultrastructural cytochemistry
48The WHO classification of AML. Therapy-related AML and MDS. Alkylating agent-related Topoisomerase II-inhibitor-related Other typesAML with recurrent cytogenetic abnormalities*AML with t(8;21)(q22;q22)AML with abnormal bone marrow eosinophils withinv(16)(p13q22) or t(16;16)(p13;q22)Acute promyelocytic leukemia witht(15;17)(q22;q12)AML with 11q23 (MLL) abnormalities.AML with multilineage dysplasia following MDS.AML not otherwise categorized. This group is nearly similar to FAB group, but blast cells are 20% in stead of 30%
50The World Health Organization (WHO) classification assigns some chronic myeloidleukaemias to a myeloproliferative category andothers, in which there are also dysplasticfeatures, to a myeloproliferative/myelodysplasticcategory
51Classification of the chronic myeloid leukaemias, based on the WHO classification. Myeloproliferative disordersChronic myelogenous leukaemia Chronic neutrophilicleukaemiaChronic eosinophilic leukaemiaBasophilic leukaemiaMast cell leukaemiaMyelodysplastic/myeloproliferative disordersChronic myelomonocytic leukaemiaChronic myelomonocytic leukaemia with eosinophiliaMyelodysplastic/myeloproliferative disorder associated witht(5;12)(q33;p13)*Atypical chronic myeloid leukaemiaJuvenile myelomonocytic leukaemia
52Chronic granulocytic leukaemia Chronic granulocytic leukaemia (CGL) is a diseaseentity with specific haematological, cytogeneticand molecular genetic features.Alternative designations are chronic myelogenousleukaemia, chronic myeloid leukaemia and chronicmyelocytic leukaemia.
53CGL is a disease ofbi- or triphasic with a chronic and an acutephase and, sometimes, an interveningaccelerated phase
54The chronic phase of chronic granulocytic leukaemia Clinical and haematological features.CGL is predominantly a disease of adults. The usual clinicalpresentation is with splenomegaly, hepatomegaly, symptomsof anaemia, and systemic symptoms such as sweating andweight loss.Occasionally this is an incidental diagnosis when a bloodcount is performed for another reason.
55The peripheral blood usually shows anemia and leucocytosis with a very characteristicdifferential count.The two predominant cell types are the myelocyte and the mature neutrophil .
56Almost all patients have an absolute basophilia and more than 90% haveeosinophilia.The platelet count is most often normal orsomewhat elevated but is low in about 5%of cases.
57BM film of a patient with CGL showing neutrophil leucocytosis with left shift.
58The bone marrow is intensely hypercellular with marked granulocytic hyperplasia andwith the myeloid/erythroid (M:E) ratiobeing greater than 10:1.There is hyperplasia of neutrophil,eosinophil and basophil lineages.
60CGL in accelerated phase and blast Transformation After a variable period in chronic phase, usuallyseveral years, CGL undergoes further evolution.There may be an abrupt transformation to anAcute leukaemia, designated blast transformation,or there may be an intervening phase ofaccelerated disease.
61The WHO group have suggested the following criteria for accelerated phase: (i) Myeloblasts constitute 10–19% of peripheral bloodwhite cells or bone marrow nucleated cells.(ii) peripheral blood basophiles are 20% or more ofnucleated cells.(iii) there is persistent thrombocytopenia or persistentthrombocytosis that does not respond to treatment.(iv) there is an increasing white cell count and increasingspleen size that does not respond to treatment.(v) cytogenetic evolution .(vi) there is marked granulocyte dysplasia or prominentproliferation of small dysplastic megakaryocytes inlarge clusters or sheets.
62Transformation may be myeloid or lymphoid. Blast transformation phase.Transformation may be myeloid or lymphoid.It is important to make the distinction sincethere lymphoblastic transformation. Lymphoidblast crisis is more likely to emerge suddenlywithout a preceding accelerated phase
63Cytogenetic and molecular genetic features CGL was the first malignant disease for which aconsistent association with an acquired non-random cytogenetic abnormality was recognized.In 1960 Nowell and Hungerford reported itsAssociation with an abnormal chromosome designated the Philadelphia (Ph) chromosome after the city of its discovery.
66Chronic lymphocytic leukaemia (CLL) is a chronic B-lineage lymphoproliferative disorder defined by characteristic morphology and immunophenotype.Small lymphocytic lymphoma is an equivalent lymphoma without circulating neoplastic cells
67CLL is the most common leukaemia in western Europe and North America with an incidence indifferent surveys varying between 1 and morethan 10/ /year.The incidence is lower in Chinese, Japanese andSouth American Indians.It is typically a disease of the elderly with ahigher incidence in males
68Clinical feature. In the later stages, CLL is characterized by lymphadenopathy, hepatomegaly,splenomegaly and eventually by impairmentof bone marrow function.In the early stages of the disease there areno symptoms or abnormal physical findingsand the diagnosis is made incidentally
69Various arbitrary levels of absolute lymphocyte count have been suggested for the diagnosis ofCLL (for example greater than /cumm).But the demonstration of a monoclonal populationof B lymphocytes with a characteristicimmunophenotype permits diagnosis at an earlierstage when the lymphocyte count is less elevated.
70A scoring system for the immunophenotypic diagnosis of chronic lymphocytic leukaemia (CLL) Score 1 for each of the following:• Weak expression of SmIg• Expression of CD5• Expression of CD23• No expression of FMC7• No expression of CD22A score of ≥4 points is confirmatory of CLL
71Peripheral blood chronic lymphocytic leukaemia showing two mature lymphocytes and one smear cell
72Peripheral blood findings In the early stages of the disease the Peripheralblood abnormality is confined to the lymphocytes.Later in the disease course there is a normocytic,normochromic anaemia and thrombocytopenia.Neutropenia is uncommon unless cytotoxic therapyhas been administered
73Bone marrow findings. The bone marrow aspirate is hypercellular as a consequence of infiltration bylymphocytes with similar features to thosein the peripheral blood.Lymphocytes percentage in the bonemarrow is 40% of all nucleated marrow cells total.
74Rai staging system for chronic lymphocytic leukaemia 0 Peripheral blood and bone marrow lymphocytosis only.I Intermediate Lymphocytosis and lymphadenopathy.II Intermediate Lymphocytosis plus hepatomegaly,splenomegaly or both.III Lymphocytosis and anaemia (haemoglobinconcentration less than 11 g/dl).IV Lymphocytosis and thrombocytopenia (platelet countless than /cmm)
75CLL Transformation. Chronic lymphocytic leukaemia may undergo two types of transformation.1-Prolymphocytoid transformation.2-large cell transformation, referred to asRichter’s syndrome.