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Acute Myeloid Leukaemia— phenotype and genotype Barbara J. Bain Department of Haematology St Mary’s Hospital, London.

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Presentation on theme: "Acute Myeloid Leukaemia— phenotype and genotype Barbara J. Bain Department of Haematology St Mary’s Hospital, London."— Presentation transcript:

1 Acute Myeloid Leukaemia— phenotype and genotype Barbara J. Bain Department of Haematology St Mary’s Hospital, London

2 The phenotypic diagnosis A case of leukaemia was first clearly described in 1845 by John Hughes Bennett and another, 6 weeks later, by Rudolf Virchow These were phenotypic diagnoses at autopsy, the blood being examined microscopically A few years later Virchow was the first to use the term “Leukhemia”

3 Things moved on The next one and a half centuries saw advances in the phenotypic diagnosis of leukaemia based on Better stains and microscopes Examination of the blood and, later, the bone marrow during life

4 Things moved on further Further advances improved the phenotypic diagnosis. These were Cytochemical stains Immunophenotyping Integration and codification of phenotypic diagnoses

5 And further still The FAB classifications from the last quarter of the 20th century can be seen as the culmination of phenotypic diagnosis The FAB group defined, and categorized and gave haematologists and, later, cytogeneticists throughout the world a common language Phenotypic diagnosis had become quite sophisticated

6 But meanwhile — Cytogenetic techniques were improving and with the introduction of chromosome banding there was a quantum leap in what could be recognized The incorporation of cytogenetic information into leukaemia classification started, e.g. MIC Group 1986.

7 And furthermore — Molecular genetics was invented Now we have FISH, SKY, spectral karyotyping, CGH PCR and RT-PCR Microarray analysis

8 What does this mean for leukaemia diagnosis and classification? Can we move from a MIC to a MIC-M approach Should we do so?

9 The WHO classification The WHO classification has taken an approach which is partly based on cytogenetic and molecular genetic analysis First, antecedent factors are considered Secondly, genetic features are considered Thirdly, phenotype is used to categorize the remaining patients

10 The WHO classification When appropriate, patients are first assigned to the category of therapy-related AML, regardless of whether they have recurring cytogenetic abnormalities These cases are further categorized as –Alkylating agent-related –Topoisomerase II-interactive drug-related –Other

11 WHO and therapy-related AML This is, in part, a genetic classification Alkylating agent-related AML is characterized by abnormalities of chromosomes 5 and 7, inv(3), t(3;3), t(6;9), t(8;16) and complex rearrangements

12 WHO and therapy-related AML Topoisomerase II-inhibitor-related AML is characterized by balanced translocations with 21q22 (AML1) and 11q23 (MLL) breakpoints including t(3;21), t(8;21), t(4;11), t(9;11) and t(11;19)(q23;p13.1) [but not t(11;19)(q23;p13.3)]

13 WHO and therapy-related AML Topoisomerase II-inhibitor-related AML is associated not only with t(8;21)(q22;q22) but also with other translocations usually found in de novo AML, e.g. t(15;17)(q22;q12) and inv(16)(p13q22)

14 Where does therapy-related AML with recurrent cytogenetic abnormality belong? It may be important to classify such cases as therapy-related in order to accurately judge the magnitude of the problem of therapy- induced AML (and ALL) These cases might also be prognostically worse than de novo cases

15 Where does therapy-related AML with recurrent cytogenetic abnormality belong? However—t-AML and de novo AML with the same cytogenetic abnormality have a lot of features in common and maybe the prognosis is not so very different

16 Therapy-related AML with t(8;21)(q22;q22) Slovak et al. (2002) reported a median survival of < 3 years and a 5 year survival of < 30% However the treatment was not standardized and in the larger group of AML patients with a 21q22 bp a quarter of patients were untreated or undertreated Slovak et al. (2002) Genes Chromosomes Cancer, 33, 379.

17 Therapy-related AML with t(15;17)(q22;q21) Andersen et al. (2002) reported, for intensively treated patients, that there was: a 69% CR rate a median survival of 29 months a 5-year survival of 45% Andersen et al. (2002) Genes Chromosomes Cancer, 33, 395.

18 Therapy-related AML with t(15;17)(q22;q21) nCR rate4-yr EFS4-yr OS 3497%65%85% ‘secondary’ 64293%68%78% de novo ‘Secondary’ = 15 surgery, 17 RT, 10 Chemo, 10 Chemo + RT Pulsoni et al. (2002) Blood, 100, 1972

19 Therapy-related AML with inv(16)(p13q22) Andersen et al (2002) reported, with intensive treatment: 85% CR 29 months median survival 45% 5-year survival Andersen et al. (2002) Genes Chromosomes Cancer, 33, 395.

20 What should we do? We should follow the WHO classification but should note the presence of relevant cytogenetic abnormalities We should recognize that topoisomerase II- inhibitor-related secondary AML is quite a different matter from t-AML related to alkylating agents

21 WHO classification AML — with recurrent cytogenetic abnormalities Advantages Certain cytogenetic/molecular genetic categories are recognized AML, e.g. with t(8;21) and inv(16) with less than 30% blasts is recognized

22 WHO classification AML — with recurrent cytogenetic abnormalities Disadvantages Acute promyelocytic leukaemia with classical and variant translocations are lumped together All cases of AML with all 11q23 breakpoint and MLL rearrangement are lumped together

23 Acute promyelocytic leukaemia and “variants” (i) SubtypeGeneATRAPML response distrib M3/M3v/t(15;17)PML-RARAYes Abnormal M3-like/t(11;17)PLZF-RARANoNormal (q23;q21) M3-like/t(11;17)NuMA-RARAProbablyNormal (q13;q21) M3-like/t(1;17)NPM-RARAProbablyNormal

24 Acute promyelocytic leukaemia and “variants” (ii) RARA and the normal cell RAR  binds to RXR. In the absence of RA, RAR  -RXR binds to RARE in the promoter of target genes, interacts with co-repressors, attracts HDACs, leading to chromatin condensation and repression of transcription In the presence of RA, RAR  -RXR is converted from a repressor to an activator since it now binds to co-activators and fails to bind to co-repressors

25 Acute promyelocytic leukaemia and “variants” (iii) RARA and the promyelocytic leukaemia cell PML-RAR  binds more strongly to co-repressors and does not dissociate in the presence of physiological levels of RA However, pharmacological levels of ATRA correct the situation HDAC inhibitors should enhance the action of ATRA and appear to do so Jones and Saha (2002) Br J Haematol, 118, 714

26 Acute promyelocytic leukaemia and “variants” (iv) RARA and the leukaemia cell of M3-like/t(15;17)/PLZF- RARA AML PLZF is a transfer factor that binds to co-repressors PLZF-RAR  therefore binds to corepressors through two binding sites —dependent on RARA and independent of it Pharmacological doses of ATRA don’t break the second bond and therefore cases are refractory to this therapy However HDAC inhibitors should enhance and appear to do so

27 Acute promyelocytic leukaemia and “variants” (v) Conclusion Not all acute leukaemias involving the RARA gene represent the same disease –Some respond to ATRA and some do not –Some respond to arsenic trioxide and some do not –A molecular classification can help us to predict or understand response to therapy

28 Acute myeloid leukaemia with 11q23 breakpoints and MLL rearrangement Is this one disease or many? The results of the European 11q23 workshop and other published data suggest that this is many diseases

29 Acute myeloid leukaemia with 11q23/MLL rearrangement n< 1 yALL t(4;11)18334 %95 % t(6;11) 30 7 %10 % t(9;11)12517 % 7 % t(10;11) 2032 %20 % t(11;19)/MLL-ELL 2114 %nil t(11;19)/MLL-ENL 3241 %66 %

30 Acute myeloid leukaemia with 11q23/MLL rearrangement FABt-AML/MDS t(4;11)M45.5 % t(6;11)M4/M5anil t(9;11)M5a 10 % t(10;11)M5a 1 % t(11;19)/MLL-ELLM4 7 % t(11;19)/MLL-ENLM4/M5anil

31 Acute myeloid leukaemia with 11q23/MLL rearrangement Conclusion Acute leukaemia with 11q23/MLL involvement is many disease not one This does not yet have much therapeutic significance but maybe it will have in the future

32 Are there other cytogenetic/ molecular genetic categories of AML we should recognize? M1/t(9;22)/BCR-ABL fusion FAB: M1 (occasionally M0, M2, M4) Usually no Auer rods Mainly de novo but occasionally after topoisomerase-II-interactive drugs Poor prognosis

33 Are there other cytogenetic/ molecular genetic categories of AML we should recognize? M2 or M4Baso/t(6;9)/DEK-CAN fusion FAB: M2 (occasionally M4 or M1) May show basophilic differentiation and Auer rods Mainly de novo but occasionally after topoisomerase-II-interactive drugs or alkylating agents Poor prognosis

34 Are there other cytogenetic/ molecular genetic categories of AML we should recognize? M5/t(8;16)/MOZ-CBP fusion FAB: M5 or M4 with haemophagocytosis Abnormal coagulation de novo or secondary (topoisomerase-II- interactive drugs or alkylating agents) Poor prognosis

35 Are there other cytogenetic/ molecular genetic categories of AML we should recognize? M7/t(1;22)/OTT-MAL fusion FAB: M7 Mainly infants

36 Are there other cytogenetic/ molecular genetic categories of AML we should recognize? Various/inv(3) or t(3;3)/EVI1 dysregulation Any category except M3; M7 over-represented Platelet count often normal or even increased Trilineage myelodysplasia De novo, secondary to alkylating agents or transformation of a MPD

37 Can we recognize any purely molecular categories of AML?

38 Can we recognize any purely molecular categories of AML? Yes

39 AML with CEBPA mutations (i) Mutations in the CEBPA gene, encoding the CCAAT/enhancer binding protein  have now been identified in 7-10% of AML CEBP  is exclusively expressed in myelomonocytic cells and is essential for neutrophilic differentiation

40 AML with CEBPA mutations (ii) CEBP  negatively regulates MYC and positively regulates the genes encoding the receptors for M-CSF, G-CSF and GM-CSF CEBP  deficient mice have granulopoiesis arrested at the myeloblast stage

41 AML with CEBPA mutations (iii) CEBP  mutations were found in 15 of 134 patients studied prospectively and were an independent good prognostic feature Such mutations, in the absence of FLT3 ITD, could lead to a patient being classified and treated as ‘good prognosis’ Preudhomme et al (2002) Blood, 100, 2717

42 What does the future hold? Is the future micro-arrays?

43 What does the future hold? Clinical assessment and morphology will remain crucial, particularly for diagnosis Cytochemistry will continue to decline in importance, but should not be neglected Immunophenotyping will hang on Molecular diagnosis will lead to more accurate classification and scientifically- based more effective treatment


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