Myelodysplastic Syndromes: WHO 2008
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1 Myelodysplastic Syndromes: WHO 2008 Attilio Orazi, M.D., FRCPath. (Engl.) Weill Medical College of Cornell University New York, NY Congresso Nazionale SIE - Società Italiana di Ematologia - MIC Milano Convention Centre. Milano Ottobre, 2009
2 MDS are clinically heterogeneous: Classification/Scoring Systems Based on morphologic differences including % blasts FAB 1976, 1982 FAB plus dysplasia (10% rule; uni vs. multi), and cytogenetics (5q-) IPSS 1997 WHO 2001 WHO 2008 WPSS 2007 Based on % blasts, karyotype, # of cytopenias Based on WHO classification and Transfusion Table courtesy of Dr. John Bennett
3 Myelodysplastic syndromes Key diagnostic points and updates
4 Guidelines: recommended levels for defining cytopenias: Hemoglobin <10 g/dl Neutrophil count <1.8 x10 9 /L Platelet count <100 x10 9 /L Values higher than these are not exclusionary of a diagnosis of MDS, if definitive morphologic and/or cytogenetic findings consistent with MDS are present
5 Morphologic Guidelines for the Diagnosis and Classification of MDS: Dyserythropoiesis, dysgranulopoiesis: >10% of cells on the BM aspirate smear Dysmegakaryopoiesis >10% of cells evaluated on 30 megakaryocytes at least (BM aspirate smear or biopsy) Ring sideroblasts (iron stain of aspirate) Blast percentage Correlate the findings with marrow biopsy
6 Erythroid precursors - multinucleation/asymmetrical nuclei and/or nuclear bridging
7 Granulocytes Pelger-Huet like nuclei, hypogranular cytoplasms Megakaryocytes Dwarf, non/ hypolobated forms including micromegakaryos
8 Definition of Ring Sideroblasts (updated from 3 rd Edition) > 15% erythroblasts with 1/3 or more of the nucleus encircled by >5 iron granules (was >10)
9 Blast Count PB 200 / BM 500 cell differential Cells counted as blasts include: - Myeloblasts - Monoblasts - Mk-blasts But not erythroblasts or promyelocytes! Mufti et al. Haematologica. 2008; 93:1712-7
10 The role of BM biopsy in MDS is better described in the 4 th edition Quality check for the adequacy of BM aspirate Provides information on stroma and cellularity (e.g. fibrosis, fatty marrow) Yields tissue for a number of studies: - immunohistochemistry - in-situ hybridization - molecular procedures (e.g. microdissection)
11 Blast count is poorly reproducible particularly in thick paraffin sections. Clusters of blasts more specific.
12 Blasts and blast clusters identification made easier by immunohistology (e.g. CD34, CD117). Micromegakaryocytes by CD61 or CD42b CD34 pos. cell clusters: 3-5 cells CD42b pos. dwarf- / micro- megs.
13 Flow cytometry to demonstrate an abnormal myeloid maturation (abnormalities in light scatter and antigenic expression) Cases with inconclusive morphologic and cytogenetic findings and >3 or more aberrant features by flow cytometry should be reevaluated over several months for definitive morphologic or cytogenetic evidence of MDS Utility of CD14/CD56 in CMML Substitution of blast % determined by flow cytometry for a visual blast count is discouraged
14 Interlaboratory variability issues: still work in progress Ogata K, et al. Diagnostic utility of flow cytometry in low-grade myelodysplastic syndromes: a prospective validation study. Haematologica Aug;94(8): Epub 2009 Jun 22. van de Loosdrecht AA et al. Standardization of flow cytometry in myelodysplastic syndromes: report from the first European LeukemiaNet working conference on flow cytometry in myelodysplastic syndromes. Haematologica Aug;94(8):
15 MDS: WHO 2008 Subtypes and changes
16 2008 WHO Classification of MDS Refractory Cytopenia with Unilineage Dysplasia (RCUD) Refractory Anemia (RA) Refractory Neutropenia (RN) Refractory Thrombocytopenia (RT) Refractory Anemia with Ring Sideroblasts (RARS) Refractory Cytopenia with Multilineage Dysplasia (RCMD) Refractory Anemia with Excess of Blasts (RAEB) Subtypes: RAEB - 1, RAEB 2 Myelodysplastic Syndrome with isolated del(5q) chrom. abnormality Myelodysplastic syndrome, Unclassifiable (MDS,U)
17 Refractory cytopenia with Mono-/bi-cytopenia 1 Dysplasia ( 10%) unilineage unilineage dysplasia No or <1% blasts 2,3 <5% blasts (RA, RN, RT) PB BM <15% ring sideroblasts Refractory anemia with Anemia Erythroid dysplasia only ring sideroblasts No blasts <5% blasts 15% ring sideroblasts Refractory cytopenia with Cytopenia(s) Dysplasia in 10% of the multilineage dysplasia No or <1% blasts 2,3 cells of two or more lineages <5% blasts < or >15% ring sideroblasts 1 Pancytopenia= MDS,u 2 If 1% blasts in PB (<5% blasts in BM)= MDS,u 3 If 2-4% blasts in PB (< 5% blasts in BM)= RAEB-1 [Note: Monocytes < /L; No Auer rods]
18 RAEB-1 PB Cytopenias <5% blasts* No Auer rods BM Uni- or multi-lineage dysplasia 5 9% blasts* No Auer rods *Also RAEB-1 if 2-4% PB blasts in cases of RCUD or RCMD with < 5% blasts in BM RAEB-2 Cytopenias 5 19% blasts Auer rods +/- Auer rods +/- Uni- or multi-lineage dysplasia 10 19% blasts MDS with isolated del(5q) Anemia, usually normal or mildly increased platelets Normal to increased megakaryocytes with hypolobated nuclei [Note: Monocytes < /L] No or <1% blasts <5% blasts No Auer rods No Auer rods
19 Diagnostic P.B. diagnostic algorithm algorithm for MDS: for P.B. MDS smear with BM (200 <5% cell blasts differential) (P.B. smear 200 cell differential) Step 1:Type of Cytopenia Mono-bicytopenia Pancytopenia Step 2: P.B. Blast Percentage Step 2: P.B. Blast Percentage No or <1% UD = RCUD UD = RARS * MD = RCMD * no blasts, > 15% RS 1%: MDS,u 2-4%: RAEB %: RAEB-2 No or <1% UD = MDS,u MD = RCMD
20 MDS with isolated 5q- Frequent erythroid hypoplasia Lenalidomide efficacy; potential leukemogenic effect A subset of patient with JAK2 mutation is mentioned (i.e. the so called proliferative variant)
21 Isolated 5q- associated with JAK2 mutation (6.2% of isolated 5q- cases) Thrombocytosis plus granulocytic proliferation in the BM and/or PB JAK2 positive cases had a trend towards: Higher platelet count (475 vs 250 x 10 9 /L; P=0.15) Higher WBC (5.21 vs 4.45x10 9 /L; P=0.012) No difference in median HB (9.0 vs 8.7 g/dl; P=0.272) Ingram et al: Leukemia 2006;20:
22 MDS, unclassifiable (3 settings) 1. Patients with refractory cytopenia with unilineage dysplasia (RCUD) or refractory cytopenia with multilineage dysplasia (RCMD) but with 1% blasts in the peripheral blood 2. Cases of MDS with unilineage dysplasia which are associated with pancytopenia. RCUD (in contrast with RCMD) only allows for a single cytopenia or bicytopenia
23 3. Persistent (6 mo.) cytopenia(s) lacking morphologic features of MDS, but with cytogenetic abnormalities presumptive for MDS* +8* -7 or del(7q) t(11;16)(q23;p13.3) -5 or del(5q) t(3;21)(q26.2;q22.1) del(20q)* -Y* i(17q) or t(17p) -13 or del(13q) del(11q) del(12p) or t(12p) del(9q) idic(x)(q13)) t(1;3)(p36.3;q21.2) t(2;11)(p21;q23) inv(3)(q21q26.2) t(6;9)(p23;q34) * As single cytogenetic abnormality, in the absence of morphologic findings, is insufficient evidence for MDS
24 Subtypes of MDS difficult to distinguish from other myeloid neoplasms (discussed in the 4 th edition) Hypoplastic MDS from acquired aplastic anemia MDS with fibrosis from other fibrotic myeloid neoplasms (e.g. PMF) MDS with > 50% erythroblasts from acute erythroid leukemia
25 MDS, hypoplastic (Cellularity <30% in patients <70 years old, and <20% in >70 years old) AA CD34: h-mds vs. AA: p=.001 hypo-mds hypo-mds Maschek H,et al. Ann Hematol 1993; 66: Orazi et al. Am J Clin Pathol 1997; 107: Bennet & Orazi. Haematologica 2009; 94:264-8
26 Hypocellularity favors response to immunosuppressive therapy? Lim ZY, et al. Low IPSS score and bone marrow hypocellularity in MDS patients predict hematological responses to antithymocyte globulin. Leukemia. 2007;21: Sloand EM, et al. Factors* affecting response and survival in patients with myelodysplasia treated with immunosuppressive therapy. J Clin Oncol. 2008;26: Sloand EM. Hypocellular myelodysplasia. Hematol Oncol Clin North Am. 2009;23: Review. *Younger age & HLA-DR15 were the most significant factors favoring response to therapy
27 MDS with fibrosis (>2+ fibrosis)
28 100 II 80 % MDS fibrosis <2 Survival 60 I II I 40 p < MDS fibrosis > mo Lambertenghi-Deliliers G, Orazi A, Lucks R, et al.: Br J Haematol, 1991
29 Scott BL, et al. Marrow fibrosis as a risk factor for posttransplantation outcome in patients with advanced myelodysplastic syndrome or acute myeloid leukemia with multilineage dysplasia. Biol Blood Marrow Transplant 2007;13: Buesche G, et al. Marrow fibrosis predicts early fatal marrow failure in patients with myelodysplastic syndromes. Leukemia 2008; 22: Della Porta MG, et al. Clinical relevance of bone marrow fibrosis and CD34-positive cell clusters in primary myelodysplastic syndromes. J Clin Oncol 2009;27:754-62
30 Erythroid-rich MDS: when the erythroid cells are > 50% of all the nucleated cells A second count for the non-erythroid cells: AML, if the blasts are > 20% of the nonerythroid cells (erythroleukemia) MDS, if the blasts are less than 20% of the non-erythroid cells If MDS, then use the original blast % for further MDS subtyping
31 Myelodysplastic/Myeloproliferative Neoplasms (MDS/MPN) (Hematopoietic malignancies with hybrid features both of MDS and MPN) Chronic myelomonocytic leukemia (CMML) Atypical chronic myeloid leukemia* (acml) MDS/MPN, unclassified (MDS/MPN,u) Refractory anemia with ring sideroblasts and thrombocytosis (RARS-T) *BCR-ABL1 negative
32 Diagnosis of MDS/MPN: WHO requires to exclude In all cases the presence of Ph chromosome/bcr/abl1 fusion gene plus, in cases with eosinophilia PDGFR Alpha abnormalities PDGFR Beta abnormalities* *May resemble CMML or acml. Include cases formerly termed CMML with t(5;12)
33 CMML: definition Monocytes >1 x10 9 /L >10% of WBC Dysplasia (mono or multilineage) is not required if all other criteria are met and there is a cytogenetic abnormality or reactive causes are excluded and monocytosis has lasted for at least 3 months ANBE
34 Monoblasts Promonocytes Monoblasts Large cells with abundant cytoplasm minimally vacuolated round nuclei with fine chromatin and nucleoli Promonocytes Irregular or folded nuclei, small indistinct nucleoli, fine chromatin and a more vacuolated and/or granular cytoplasm Abnormal monocytes (immature) Abnormal monocytes More condensed chromatin, convoluted nuclei and more granulated Adapted from WHO Classification of Tumors of Haematopoietic and Lymphoid Tissues. IARC: Lyon Fig.1-04 (p. 21).
35 RARS-T Its incidence is rare (underdiagnosed?) JAK2 positive in approx. 60% of cases MPL W515K/L (very rare) RARS-T has a worse outcome than ET Is controversial Wardrop D, Steensma DP. Is RARS-T a necessary or useful diagnostic category? Br J Haematol. 2009;144:809-17
36
37 BM Biopsy
38 RARS-T study (44 cases) Massachusetts General Hospital, Boston, MA 1 U. Mass Memorial Med. Center, Worcester, MA 2 Brigham and Women s Hospital, Boston, MA 3 Weill Cornell College of Medicine, New York, NY 4 Genzyme, Inc, New York, NY 5 Baystate Med. Center, Springfield, MA 5 Hasserjian RP 1, Soupir CP 1, Wang Sa A 2, Vergilio JoA 3, Orazi A 4, Dong H 5
39 Karyotype (40 patients) 31 (78%) had a normal karyotype 9 cases (22%) with abnormal karyotype which included: isolated +8 (4 cases) isolated del(20)(q11.2) (1 case) isolated Y (1 case) isolated del(11)(q23) (1 case) +1p,-6 (1 case) del(7)(q11.2)(1 case)
40 Morphology: comparative analysis of RARS-T, RARS and MPN RARS-T & RARS: have similar Erythroid hyperplasia Left shift erythrop. Dyserythropoiesis RARS-T & MPN both have: Megs. equally numerous Similarly large and pleomorphic Similarly variable fibrosis
41 Fibrosis in RARS-T 5 of 32 cases (16%) in our series 12 of 28 cases* (43%) had fibrosis: MF-1: 7 cases MF-2: 5 cases *Schmitt-Graeff AH, et al. Haematologica 2008
42 Morphology of JAK2 (-) vs. JAK2 (+) JAK2(+) cases had a significantly higher PLT and WBC counts, and a trend towards a higher HB level JAK2(+) cases had more prevalent large Megs often with staghorn or bulbous nuclear morphology while small and normal-sized Megs. were predominant in the JAK2(-) cases No differences in reticulin fibrosis and morphologic dysplasia in other lineages
43 Additional issues Prognosis Molecular pathogenesis
44 100 Overall survival of RARS-Tpatients Percent survival cases Time (months) The median follow-up for all patients was 59 months; at the time of study analysis (9/20/09), 26 patients had died..median survival for all pts. 74 months
45 Transformation to AML Our series: 4/27 (15%) cases with follow up Pich A, Godio L. RARS- T associated with JAK2V617F mutation transformed to AML with chromosomal evolution including monosomy 7. Leuk Res (2009). In press.
46 Prognostic role for JAK2 23 cases Schmitt-Graeff A et al. Haematologica 2008
47 Prognostic role for JAK2? Suggested by: Atallah E, et al. Leukemia. 2008; 22: Not confirmed by: Wang SA, et al. Leukemia. 2006;2 0: Raya JM, et al. Int J Hematol. 2008; 88: Hasserjian RP, et al. our current study
48 Molecular pathogenesis: Malcovati L, et al. Blood Aug 19. [Epub ahead of print] 19 RARS-T defined according to WHO 2008 Like in RARS, up-regulation of ALAS2 and downregulation of ABCB7 in CD34+ cells, i.e. RARS-T has distinctive sideroblastic features at the molecular level. X-chromosome inactivation shows that JAK2 pos. cells are only a fraction of clonal granulocytes 3 RARS pts. with low or normal plts. progressed to RARS-T (2/3 acquired JAK2). It may develop from RARS through the acquisition of somatic mutations of JAK2, MPL or other as-yet-unknown genes
49 Summary Morphology is still very important for diagnosis Perform always iron and reticulin stains Look out for monocytes in PB and BM and use cytochemistry and flow to confirm monocytosis Bone marrow biopsy supplemented by immunohistology (particularly useful in cases with fibrosis or fatty BM) Karyotype is mandatory. FISH may supplement BCR-ABL1 and, in cases with eosinophilia, PDGFR rearrangements Test for JAK2 when appropriate (e.g. RARS-T, 5q-)
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