Abstract
Flow cytometry (FC) is increasingly recognized as an important tool in the diagnosis and prognosis of myelodysplastic syndromes (MDS). However, validation of current assays and agreement upon the techniques are prerequisites for its widespread acceptance and application in clinical practice. Therefore, a working group was initiated (Amsterdam, 2008) to discuss and propose standards for FC in MDS. In 2009 and 2010, representatives from 23, mainly European, institutes participated in the second and third European LeukemiaNet (ELN) MDS workshops. In the present report, minimal requirements to analyze dysplasia are refined. The proposed core markers should enable a categorization of FC results in cytopenic patients as ‘normal’, ‘suggestive of’, or ‘diagnostic of’ MDS. An FC report should include a description of validated FC abnormalities such as aberrant marker expression on myeloid progenitors and, furthermore, dysgranulopoiesis and/or dysmonocytopoiesis, if at least two abnormalities are evidenced. The working group is dedicated to initiate further studies to establish robust diagnostic and prognostic FC panels in MDS. An ultimate goal is to refine and improve diagnosis and prognostic scoring systems. Finally, the working group stresses that FC should be part of an integrated diagnosis rather than a separate technique.
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References
Cazzola M, la Porta MG, Travaglino E, Malcovati L . Classification and prognostic evaluation of myelodysplastic syndromes. Semin Oncol 2011; 38: 627–634.
Valent P, Horny HP, Bennett JM, Fonatsch C, Germing U, Greenberg P et al. Definitions and standards in the diagnosis and treatment of the myelodysplastic syndromes: consensus statements and report from a working conference. Leuk Res 2007; 31: 727–736.
Ogata K, Nakamura K, Yokose N, Tamura H, Tachibana M, Taniguchi O et al. Clinical significance of phenotypic features of blasts in patients with myelodysplastic syndrome. Blood 2002; 100: 3887–3896.
Wells DA, Benesch M, Loken MR, Vallejo C, Myerson D, Leisenring WM et al. Myeloid and monocytic dyspoiesis as determined by flow cytometric scoring in myelodysplastic syndrome correlates with the IPSS and with outcome after hematopoietic stem cell transplantation. Blood 2003; 102: 394–403.
van de Loosdrecht AA, Westers TM, Westra AH, Drager AM, van der Velden VHJ, Ossenkoppele GJ . Identification of distinct prognostic subgroups in low- and intermediate-1-risk myelodysplastic syndromes by flow cytometry. Blood 2008; 111: 1067–1077.
van de Loosdrecht AA, Alhan C, Bene MC, Della Porta MG, Drager AM, Feuillard J et al. Standardization of flow cytometry in myelodysplastic syndromes: report from the first European LeukemiaNet working conference on flow cytometry in myelodysplastic syndromes. Haematologica 2009; 94: 1124–1134.
Brunning R, Orazi A, Germing U, LeBeau MM, Porwit A, Baumann I et al. Myelodysplastic syndromes/neoplasms. In: Swerdlow SH et al. (eds). WHO Classification of Tumours and Haematopoietic and Lymphoid Tissues. IARC: Lyon, 2008, pp 88–93.
Arroyo JL, Fernandez ME, Hernandez JM, Orfao A, San Miguel JF, Del Canizo MC . Impact of immunophenotype on prognosis of patients with myelodysplastic syndromes. Its value in patients without karyotypic abnormalities. Hematol J 2004; 5: 227–233.
Malcovati L, Della Porta MG, Lunghi M, Pascutto C, Vanelli L, Travaglino E et al. Flow cytometry evaluation of erythroid and myeloid dysplasia in patients with myelodysplastic syndrome. Leukemia 2005; 19: 776–783.
Scott BL, Wells DA, Loken MR, Myerson D, Leisenring WM, Deeg HJ . Validation of a flow cytometric scoring system as a prognostic indicator for posttransplantation outcome in patients with myelodysplastic syndrome. Blood 2008; 112: 2681–2686.
Valent P, Orazi A, Busche G, Schmitt-Graff A, George TI, Sotlar K et al. Standards and impact of hematopathology in myelodysplastic syndromes (MDS). Oncotarget 2010; 1: 483–496.
Stelzer GT, Shults KE, Loken MR . CD45 gating for routine flow cytometric analysis of human bone marrow specimens. Ann NY Acad Sci 1993; 677: 265–280.
Loken MR, van de Loosdrecht AA, Ogata K, Orfao A, Wells DA . Flow cytometry in myelodysplastic syndromes: report from a working conference. Leuk Res 2008; 32: 5–17.
Bjornsson S, Wahlstrom S, Norstrom E, Bernevi I, O’Neill U, Johansson E et al. Total nucleated cell differential for blood and bone marrow using a single tube in a five-color flow cytometer. Cytometry B Clin Cytom 2008; 74: 91–103.
Allan RW, Nsari-Lari MA, Jordan S . DRAQ5-based, no-lyse, no-wash bone marrow aspirate evaluation by flow cytometry. Am J Clin Pathol 2008; 129: 706–713.
Loken MR, Chu SC, Fritschle W, Kalnoski M, Wells DA . Normalization of bone marrow aspirates for hemodilution in flow cytometric analyses. Cytometry B Clin Cytom 2008; 76B: 27–36.
Brooimans RA, Kraan J, van Putten W, Cornelissen JJ, Lowenberg B, Gratama JW . Flow cytometric differential of leukocyte populations in normal bone marrow: Influence of peripheral blood contamination1. Cytometry B Clin Cytom 2008; 76B: 18–26.
Knipp S, Strupp C, Gattermann N, Hildebrandt B, Schapira M, Giagounidis A et al. Presence of peripheral blasts in refractory anemia and refractory cytopenia with multilineage dysplasia predicts an unfavourable outcome. Leuk Res 2008; 32: 33–37.
Cesana C, Klersy C, Brando B, Nosari A, Scarpati B, Scampini L et al. Prognostic value of circulating CD34+ cells in myelodysplastic syndromes. Leuk Res 2008; 32: 1715–1723.
Alhan C, Westers TM, Ossenkoppele GJ, van de Loosdrecht AA . Do peripheral blasts count in myelodysplastic syndromes? Leuk Res 2009; 33: 209–211.
Braulke F, Schanz J, Jung K, Shirneshan K, Schulte K, Schuetze C et al. FISH analysis of circulating CD34+ cells as a new tool for genetic monitoring in MDS: verification of the method and application to 27 MDS patients. Leuk Res 2010; 34: 1296–1301.
Stetler-Stevenson M, Arthur DC, Jabbour N, Xie XY, Molldrem J, Barrett AJ et al. Diagnostic utility of flow cytometric immunophenotyping in myelodysplastic syndrome. Blood 2001; 98: 979–987.
Benesch M, Deeg HJ, Wells D, Loken M . Flow cytometry for diagnosis and assessment of prognosis in patients with myelodysplastic syndromes. Hematology 2004; 9: 171–177.
Stachurski D, Smith BR, Pozdnyakova O, Andersen M, Xiao Z, Raza A et al. Flow cytometric analysis of myelomonocytic cells by a pattern recognition approach is sensitive and specific in diagnosing myelodysplastic syndrome and related marrow diseases: emphasis on a global evaluation and recognition of diagnostic pitfalls. Leuk Res 2008; 32: 215–224.
Matarraz S, Lopez A, Barrena S, Fernandez C, Jensen E, Flores J et al. The immunophenotype of different immature, myeloid and B-cell lineage-committed CD34+ hematopoietic cells allows discrimination between normal/reactive and myelodysplastic syndrome precursors. Leukemia 2008; 22: 1175–1183.
Wood BL . Myeloid malignancies: myelodysplastic syndromes, myeloproliferative disorders, and acute myeloid leukemia. Clin Lab Med 2007; 27: 551–575, vii.
Font P, Subira D, Mtnez-Chamorro C, Castanon S, Arranz E, Ramiro S et al. Evaluation of CD7 and terminal deoxynucleotidyl transferase (TdT) expression in CD34+ myeloblasts from patients with myelodysplastic syndrome. Leuk Res 2006; 30: 957–963.
Matarraz S, Lopez A, Barrena S, Fernandez C, Jensen E, Flores-Montero J et al. Bone marrow cells from myelodysplastic syndromes show altered immunophenotypic profiles that may contribute to the diagnosis and prognostic stratification of the disease: a pilot study on A series of 56 patients. Cytometry B Clin Cytom 2010; 78: 154–168.
Monreal MB, Pardo ML, Pavlovsky MA, Fernandez I, Corrado CS, Giere I et al. Increased immature hematopoietic progenitor cells CD34+/CD38dim in myelodysplasia. Cytometry B Clin Cytom 2006; 70: 63–70.
Goardon N, Nikolousis E, Sternberg A, Chu WK, Craddock C, Richardson P et al. Reduced CD38 expression on CD34+ cells as a diagnostic test in myelodysplastic syndromes. Haematologica 2009; 94: 1160–1163.
Westers TM, van der Velden VH, Alhan C, Bekkema R, Bijkerk A, Brooimans RA et al. Implementation of flow cytometry in the diagnostic work-up of myelodysplastic syndromes in a multicenter approach: Report from the Dutch Working Party on Flow Cytometry in MDS. Leuk Res 2012; 36: 422–430.
Marsh JC, Ball SE, Cavenagh J, Darbyshire P, Dokal I, Gordon-Smith EC et al. Guidelines for the diagnosis and management of aplastic anaemia. Br J Haematol 2009; 147: 43–70.
Ogata K, Della Porta MG, Malcovati L, Picone C, Yokose N, Matsuda A et al. Diagnostic utility of flow cytometry in low-grade myelodysplastic syndromes: a prospective validation study. Haematologica 2009; 94: 1066–1074.
Vikentiou M, Psarra K, Kapsimali V, Liapis K, Michael M, Tsionos K et al. Distinct neutrophil subpopulations phenotype by flow cytometry in myelodysplastic syndromes. Leuk Lymphoma 2009; 50: 401–409.
Cherian S, Moore J, Bantly A, Vergilio JA, Klein P, Luger S et al. Peripheral blood MDS score: a new flow cytometric tool for the diagnosis of myelodysplastic syndromes. Cytometry B Clin Cytom 2005; 64: 9–17.
Lorand-Metze I, Ribeiro E, Lima CS, Batista LS, Metze K . Detection of hematopoietic maturation abnormalities by flow cytometry in myelodysplastic syndromes and its utility for the differential diagnosis with non-clonal disorders. Leuk Res 2007; 31: 147–155.
Huizinga TW, van der Schoot CE, Jost C, Klaassen R, Kleijer M, von dem Borne AE et al. The PI-linked receptor FcRIII is released on stimulation of neutrophils. Nature 1988; 333: 667–669.
de Haas M, Kleijer M, van Zwieten R, Roos D, von dem Borne AEG . Neutrophil Fc gamma RIIIb deficiency, nature, and clinical consequences: a study of 21 individuals from 14 families. Blood 1995; 86: 2403–2413.
Moulding DA, Hart CA, Edwards SW . Regulation of neutrophil FcgammaRIIIb (CD16) surface expression following delayed apoptosis in response to GM-CSF and sodium butyrate. J Leukoc Biol 1999; 65: 875–882.
Wang L, Wells DA, Deeg HJ, Loken MR . Flow cytometric detection of nonneoplastic antigenic polymorphisms of donor origin after allogeneic marrow transplant: a report of two cases. Am J Clin Pathol 2004; 122: 135–140.
Elghetany MT, Davis BH . Impact of preanalytical variables on granulocytic surface antigen expression: a review. Cytometry B Clin Cytom 2005; 65: 1–5.
Wang H, Chuhjo T, Yasue S, Omine M, Nakao S . Clinical significance of a minor population of paroxysmal nocturnal hemoglobinuria-type cells in bone marrow failure syndrome. Blood 2002; 100: 3897–3902.
Nakao S, Sugimori C, Yamazaki H . Clinical significance of a small population of paroxysmal nocturnal hemoglobinuria-type cells in the management of bone marrow failure. Int J Hematol 2006; 84: 118–122.
Wang SA, Pozdnyakova O, Jorgensen JL, Medeiros LJ, Stachurski D, Anderson M et al. Detection of paroxysmal nocturnal hemoglobinuria clones in patients with myelodysplastic syndromes and related bone marrow diseases, with emphasis on diagnostic pitfalls and caveats. Haematologica 2009; 94: 29–37.
Ando K, Tanaka Y, Hashimoto Y, Ohyashiki JH, Sugimori N, Nakao S et al. PNH-phenotype cells in patients with idiopathic cytopenia of undetermined significance (ICUS) with megakaryocytic hypoplasia and thrombocytopenia. Br J Haematol 2010; 150: 705–707.
Borowitz MJ, Craig FE, Digiuseppe JA, Illingworth AJ, Rosse W, Sutherland DR et al. Guidelines for the diagnosis and monitoring of paroxysmal nocturnal hemoglobinuria and related disorders by flow cytometry. Cytometry B Clin Cytom 2010; 78: 211–230.
Sconocchia G, Fujiwara H, Rezvani K, Keyvanfar K, El OF, Grube M et al. G-CSF-mobilized CD34+ cells cultured in interleukin-2 and stem cell factor generate a phenotypically novel monocyte. J Leukoc Biol 2004; 76: 1214–1219.
Xu Y, McKenna RW, Karandikar NJ, Pildain AJ, Kroft SH . Flow cytometric analysis of monocytes as a tool for distinguishing chronic myelomonocytic leukemia from reactive monocytosis. Am J Clin Pathol 2005; 124: 799–806.
Subira D, Font P, Villalon L, Serrano C, Askari E, Gongora E et al. Immunophenotype in chronic myelomonocytic leukemia: is it closer to myelodysplastic syndromes or to myeloproliferative disorders? Transl Res 2008; 151: 240–245.
Lacronique-Gazaille C, Chaury MP, Le GA, Faucher JL, Bordessoule D, Feuillard J . A simple method for detection of major phenotypic abnormalities in myelodysplastic syndromes: expression of CD56 in CMML. Haematologica 2007; 92: 859–860.
Sternberg A, Killick S, Littlewood T, Hatton C, Peniket A, Seidl T et al. Evidence for reduced B-cell progenitors in early (low-risk) myelodysplastic syndrome. Blood 2005; 106: 2982–2991.
Ogata K, Kishikawa Y, Satoh C, Tamura H, Dan K, Hayashi A . Diagnostic application of flow cytometric characteristics of CD34+ cells in low-grade myelodysplastic syndromes. Blood 2006; 108: 1037–1044.
Maftoun-Banankhah S, Maleki A, Karandikar NJ, Arbini AA, Fuda FS, Wang HY et al. Multiparameter flow cytometric analysis reveals low percentage of bone marrow hematogones in myelodysplastic syndromes. Am J Clin Pathol 2008; 129: 300–308.
Ogata K . Diagnostic flow cytometry for low-grade myelodysplastic syndromes. Hematol Oncol 2008; 26: 193–198.
Loken MR, Shah VO, Dattilio KL, Civin CI . Flow cytometric analysis of human bone marrow: I. Normal erythroid development. Blood 1987; 69: 255–263.
Della Porta MG, Malcovati L, Invernizzi R, Travaglino E, Pascutto C, Maffioli M et al. Flow cytometry evaluation of erythroid dysplasia in patients with myelodysplastic syndrome. Leukemia 2006; 20: 549–555.
Maynadie M, Picard F, Husson B, Chatelain B, Cornet Y, Le RG et al. Immunophenotypic clustering of myelodysplastic syndromes. Blood 2002; 100: 2349–2356.
Pfeilstocker M, Karlic H, Nosslinger T, Sperr W, Stauder R, Krieger O et al. Myelodysplastic syndromes, aging, and age: correlations, common mechanisms, and clinical implications. Leuk Lymphoma 2007; 48: 1900–1909.
Chu SC, Wang TF, Li CC, Kao RH, Li DK, Su YC et al. Flow cytometric scoring system as a diagnostic and prognostic tool in myelodysplastic syndromes. Leuk Res 2011; 35: 868–873.
Clark RE, Smith SA, Jacobs A . Myeloid surface antigen abnormalities in myelodysplasia: relation to prognosis and modification by 13-cis retinoic acid. J Clin Pathol 1987; 40: 652–656.
Horny HP, Sotlar K, Sperr WR, Valent P . Systemic mastocytosis with associated clonal haematological non-mast cell lineage diseases: a histopathological challenge. J Clin Pathol 2004; 57: 604–608.
Kussick SJ, Fromm JR, Rossini A, Li Y, Chang A, Norwood TH et al. Four-color flow cytometry shows strong concordance with bone marrow morphology and cytogenetics in the evaluation for myelodysplasia. Am J Clin Pathol 2005; 124: 170–181.
Satoh C, Dan K, Yamashita T, Jo R, Tamura H, Ogata K . Flow cytometric parameters with little interexaminer variability for diagnosing low-grade myelodysplastic syndromes. Leuk Res 2008; 32: 699–707.
Truong F, Smith BR, Stachurski D, Cerny J, Medeiros LJ, Woda BA et al. The utility of flow cytometric immunophenotyping in cytopenic patients with a non-diagnostic bone marrow: a prospective study. Leuk Res 2009; 33: 1039–1046.
Kern W, Haferlach C, Schnittger S, Haferlach T . Clinical utility of multiparameter flow cytometry in the diagnosis of 1013 patients with suspected myelodysplastic syndrome: correlation to cytomorphology, cytogenetics, and clinical data. Cancer 2010; 116: 4549–4563.
Falco P, Levis A, Stacchini A, Ciriello MM, Geuna M, Notari P et al. Prognostic relevance of cytometric quantitative assessment in patients with myelodysplastic syndromes. Eur J Haematol 2011; 87: 409–418.
Acknowledgements
We express our gratitude to Wolfgang Kern (MLL, Munich, Germany) and Robin Ireland (King's College Hospital, London, UK) for their excellent organization and facilitation of the ELN Working Group meetings in 2009 and 2010, respectively. Furthermore, we acknowledge financial support from the European LeukemiaNet (ELN WP8), the ‘Elimination of Leukaemia Fund’ (UK), Leukaemia and Lymphoma Research (UK), Alexion (UK), Amgen Oncology (UK, Beckman Coulter (Krefeld, Germany), Becton Dickinson (UK), and Celgene (Germany and UK), and Cooperative Research Thematic Network on Cancer (RD06/0020/0035) from the Ministerio de Ciencia e Innovación (Madrid, Spain).
Author Contributions
All persons listed as co-authors contributed to pre-conference and post-conference discussions (June 2009 until November 2010), and actively participated in the Standardization Conferences (Munich, Germany, 30–31 October 2009; London, UK, 5–6 November 2010). All co-authors contributed equally by discussing criteria, standards, algorithms and recommendations at the Working Conference. In addition, all persons listed as co-authors provided essential input by drafting parts of the manuscript and by approving the final version of the document.
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Westers, T., Ireland, R., Kern, W. et al. Standardization of flow cytometry in myelodysplastic syndromes: a report from an international consortium and the European LeukemiaNet Working Group. Leukemia 26, 1730–1741 (2012). https://doi.org/10.1038/leu.2012.30
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DOI: https://doi.org/10.1038/leu.2012.30
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