acute leukaemia

ACUTE LEUKAEMIA

by

DR. KAMAL E. HIGGYCONSULTANT HAEMATOLOGIST

ACUTE MYELOID LEUKAEMIAETIOLOGICAL FACTORS

GENETIC DISORDERS

Congenital Defects

Down syndrome

Bloom Syndrome

Monosomy 7 syndrome

Klinefelter Syndrome

Turner Syndrome

Neurofibromatosis

Congenital Dysmorphic Syndrome

Marrow Failure Syndromes

Fanconi Anaemia

Dyskeratosis Congenita

Schwachman – Diamond Syndrome

Amegakaryocytic Thrombocytopenia

Blackfan – Diamond Syndrome

Kostmann Agranulocytosis

Familial Aplastic Anaemia

ACUTE MYELOID LEUKAEMIAETIOLOGICAL FACTORS

Environmental Factors Solvents (benzene) Smoking Ionizing radiation Atomic bomb exposure Nuclear power exposure Medical radiation Non ionizing radiation Chemotherapy Alkylating agents Topoisomerase II inhibitors Other drugs Chloramphenicol Phenylbutazone

Acute Myeloid Leukaemia DEFINITION

Acute Myeloid Leukaemia (AML) is currently defined by the FAB criteria.

The percentage of blasts, the presence of cytochemical myeloperoxidase, the major cell types present defined by morphology and esterase cytochemistry & the immunophenotype define the 8 FAB subtypes

Acute Myeloid LeukaemiaCLINICAL FEATURES

Increasing incidence with age. Median age at diagnosis is 50 years.

Symptoms are due to marrow failure (anaemia, infection, haemorrhage) or hyperleucocytosis.

Rarely presenting symptoms are due to chloromas or CNS involvement.

Acute Myeloid LeukaemiaLABORATORY DIAGNOSIS

MORPHOLOGY

Myeloblasts are usually medium sized with an eccentric nucleus and open chromatin. Cytoplasm shows variable basophilia. Few granules or Auer rods may be present.

Maturing granulocytes may be normal or show the dysplastic features e.g. Psaudo Pelger (hypogranular bilobed neutrophils).

Monoblasts are generally very large with abundant grey-blue or basophilic cytoplasm. Nuclei are round or lobulated, and are central in the cell. Fine azurophil granules may be present but Auer rods are rarely, if ever, present.

Promonocytes and monocytes show abnormal nuclear maturation, granulation and loss of basophilia.


MORPHOLOGY (Cont…) Erythroid precursors may be normal or show varying

degrees of dyserythropoiesis. Eosinophils may be present in varying numbers, with

normal or abnormal morphology. Specific abnormal appearances are linked with cytogenetic abnormalities

(e.g. inv 16) Basophils are rarely increased in AML, and if present show

abnormal or poor granule formation and nuclear maturation. Megakaryocytes may be reduced or increased. Dysplastic

hyperlobated, hypolobated, multinuclear, small and blastic forms may be present

Acute Myeloid LeukaemiaFAB Subtypes

M0-M5: >20% myelo/monoblasts by morphology or immunophenotype.

M6: >50% erythroid precursors, >20% blasts in non-erythroid cells.

M7: >20% megakaryoblasts present.

Note: Cytogenetics, molecular genetics, previous MDS or MPD, previous

chemo- or radiotherapy and the presence of trilineage myelodysplasia do not have any place in the FAB system. All these features which contribute to the definition of prognostically important sub-groups is included in the new classification system.

Acute Myeloid LeukaemiaFAB Subtypes

>20%

>20%

>20%

>20%

>20%

>20%

M0 M1 M2

M3 M4

M5b

M5a

M6 M7

ACUTE MYELOID LEUKAEMIAS WHO CLASSIFICATION

Acute myeloid leukaemia with reccurent genetic abnormalities

Acute myeloid leukaemia with t(8;21)(q22;q22); (AML1(CBFa)/ETO)

Acute myeloid leukaemia with abnormal bone marrow eosinophils inv (16)(p13q22) or t(16; 16)(p13;q22); (CBFb/MYH11)

Acute promyelocytic leukaemia (AML with t(15; 17)(q22;q12)(PML/RARα) & variants

Acute myeloid leukaemia with 11q23 (MLL) abnormalities

Acute myeloid leukaemia with multilineage dysplasia Acute myeloid leukaemia and myelodysplastic syndromes,

therapy related

ACUTE MYELOID LEUKAEMIAS

WHO CLASSIFICATION Acute myeloid leukaemia not otherwise categorized Acute myeloid leukaemia minimally differentiated Acute myeloid leukaemia without maturation Acute myeloid leukaemia with maturation Acute myelomonocytic leukaemia Acute monoblastic and monocytic leukaemia Acute erythroid leukaemias Acute megakaryoblastic leukaemia Acute basophilic leukaemia Acute panmyelosis with myelofibrosis Myeloid sarcoma Acute leukaemia of ambiguous lineage Undifferentiated acute leukaemia Bilineal acute leukaemia

Biphenotypic leukaemia


CYTOCHEMISTRY

Myeloperoxidase (MPO): identify blast cells as myeloid. Auer rods are detected twice as frequently as on Romanowsky stains. Dysplastic neutrophils may be negative. Eosinophil granules are always positive. Monoblasts and promonocytes may be negative. Sudan Black B gives identical results.

Myeloperoxidase (MPO)


CYTOCHEMISTRY

Chloroacetate Esterase (CAE) specifically identifies cells of the granulocyte lineage, from the early promyelocyte stage to mature neutrophils.

Chloroacetate Esterase


CYTOCHEMISTRY

α -Naphthyl Acetate (Nonspecific) Esterase (ANAE) stains monocytes and megakaryocytes at all stages of maturation.

Nonspecific Esterase


CYTOCHEMISTRY

Periodic Acid Schiff (PAS) stains many cell types. It is not lineage specific but the pattern of staining may be helpful, e.g. positive monoblasts, NRBC, megakaryoblasts…

Acute Myeloid Leukaemia LABORATORY DIAGNOSIS

TREPHINE BIOPSY HISTOLOGY

Helpful when there is severe cytopenia and a dry tap, e.g. in megakaryoblastic AML.

Useful for identifying megakaryocyte dysplasia and dyserythropoiesis.

Immunophenotyping is possible when the diagnosis is in doubt.

Not suitable for fine classification which is based on percentages of cell types and cytological detail.


FLOW CYTOMETRY

Myeloid blasts express combinations of CD34, CD13, CD33, CD117, HLA-DR, CD14, CD15. Glycophorin A and CD42b identify erythroid and megakaryocyte lineage blasts respectively.

Essential to separate undifferentiated AML (M0) from ALL. May need supplementation with APAAP staining for cytoplasmic immunoreactive MPO.


FLOW CYTOMETRY (Cont…)

Essential for diagnosing biphenotypic leukaemias and detecting aberrant (promiscuous) antigen expression.

Can identify patient specific unique blast cell phenotypes.

Can identify blasts of megakaryocyte, erythroid, monocyte and granulocyte lineages.

Can identify leukaemic contamination of 'remission' marrow harvests.

Can identify early relapse.


MOLECULAR GENETICS

The 3 major good prognosis translocations are all detectable by RT-PCR, which can be used to detect up to 40% more cases than are found by metaphase cytogenetics.

All translocations with cloned breakpoint genes are theoretically detectable by RT-PCR.

Real time multiplex PCR may become available for routine diagnosis.

RT-PCR is used for monitoring speed of response to treatment and may be predictive for relapse


CYTOGENETICS

An abnormal karyotype is found in 50-60% of AML cases at presentation.

t(15;17), t(8;21) and inv/del/t(16) are associated with specific morphology, younger patients and a good prognosis.

Complex karyotypes and abnormalities of chromosomes 5 and 7 are associated with older patients, trilineage dysplasia and a poor response to treatment.

Other karyotypic abnormalities are prognostically neutral.


IMMUNOHISTOCHEMISTRY / IMMUNOFLUORESCENCE

APAAP staining for cMPO, cCD3 and cCD79 should be done in all cases of undifferentiated acute leukaemia.

Immunofluorescent staining for nuclear PML protein should be done in all cases of suspected acute promyelocytic leukaemia. A microparticulate pattern is diagnostic of APML.

Acute Lymphoblastic Leukaemia (ALL) Clinical Features

ALL can occur at any age but has a peak incidence between 2 - 10 years. It is characterized by:

bone marrow failure lymphadenopathy thymic enlargement in T-lineage ALL bone pains which may be associated with

radiological changes and fractures tendency to relapse in the CNS and testis.

Acute Lymphoblastic LeukaemiaLaboratory Diagnosis

FAB Classification


MorphologyBlast morphology is variable some are small with high nuclear / cytoplasmic

ratios and indistinct nucleoli (so-called L1 blasts) while others are larger with more prominent

nucleoli and more abundant cytoplasm (L2). A third type (L3) shows large blasts with

moderately abundant highly vacuolated basophilic cytoplasm

Blast morphology does not correlate with cell lineage and cytochemistry is of little value.

ALL (L2)

ALL (L3)

Acute Lymphoblastic LeukaemiaLaboratory Diagnosis - FAB Classification

ALL (L1)

PAS Ac. Phos (T-ALL)


CYTOCHEMISTRY


Flowcytometry ALL is derived from precursor lymphocytes that are undergoing

antigen receptor gene (Ig and TCR) rearrangement.

B-lineage ALL

The precursor nature of the cells is established by demonstrating lack of surface Ig, the presence of nuclear TDT and sometimes the expression of CD34.

Sub classification is as follows:

Pre-pre B-ALL: CD19+ CD10- cytoplasmic mu heavy chain negative. Common ALL: CD19+ CD10+ cytoplasmic mu present in <20% of cells Pre B-ALL: CD19+ CD10+ cytoplasmic mu present in >20% of cells Blasts of all subgroups will express cytoplasmic CD22 and CD79b.


Flowcytometry

T-lineage ALL

The precursor nature of the cells is established by demonstrating TDT and sometimes CD34 positivity and the lack of surface TCR/CD3.

T-cell lineage is demonstrated by the expression of CD7 and/or CD1a.

Expression of the other pan-T cell markers is variable.


CYTOGENETICS Hyperdiploidy is common. A number of balanced translocations

have been identified in ALL:

t(12;21) - this is the commonest translocation in ALL (30% of cases). It results in the TEL-AML fusion gene and is primarily associated with the common phenotype.

t(9;22) - this is commoner in adults and is associated with a very poor prognosis.

t(4;11) - this translocation results in the MLL-AF4 fusion gene. It is associated with pre-pre B-ALL and is associated with a poor prognosis.

t(1;19) - associated with pre-B ALL and results in the formation of the E2A-PBX fusion gene.

These translocations are demonstrable by RT-PCR techniques.

TAL-1 deregulation is the commonest genetic abnormality in T-ALL. This may occur as the result of the t(1;14) or more commonly due to chromosome 1p32 deletions.

Acute Lymphoblastic LeukaemiaPROGNOSTIC FACTORS

The following are poor prognostic factors in ALL:

age <1 year and >10 years male sex CNS disease at presentation high white cell count t(9;22) t(4;11) hypodiploidy

Acute Lymphoblastic LeukaemiaOUTCOME AND THERAPY

The treatment of ALL consists of the following "phases":

Remission induction - vincristine, prednisolone, daunorubicin, asparaginase.

Consolidation - various combinations of chemotherapeutic agents.

CNS directed therapy - high dose systemic and intrathecal methotrexate.

Maintenance therapy - vincristine, prednisolone, mercaptopurine and methotrexate for 2 years.

Childhood ALL is associated with 75% long term survival. Minimal residual disease assessment using PCR based strategies appear to be able to predict relapse although they are not yet in routine clinical use. Allogeneic transplantation is the treatment of choice at relapse.

The outlook in adult ALL is poor with approximately 20% long-term survivors. Allogeneic transplantation is advisable in first remission.

acute leukaemia

Documents