Nov 7, 2013 - 7Division of Oncology and Children's Research Center, University Children's Hospital, .... common acute lymphoblastic leukemia (cALL) with a.
Cytometry Part B (Clinical Cytometry) 86B:288–291 (2014)
Brief Communication
Flow Diagnostics Essential Code: A Simple and Brief Format for the Summary of Leukemia Phenotyping Ondrej Hrusak,1* Giuseppe Basso,2 Richard Ratei,3 Giuseppe Gaipa,4 Drorit Luria,5 Ester Mejstrıkov a,1 Leonid Karawajew,3 Barbara Buldini,2 Eti Rozenthal,6 Jean Pierre Bourquin,7 Tom as Kalina,1 Mary Sartor,8 and Michael N. Dworzak,9 on behalf of the AIEOP-BFM Flow Network 1
Department of Pediatric Hematology and Oncology, Charles University 2nd Faculty of Medicine and University Hospital Motol, Prague, Czech Republic 2 Dipartimento di Salute della Donna e del Bambino, Padova, Italy 3 Department of Pediatric Oncology/Hematology, Charite– Universit€atsmedizin Berlin, Germany 4 Laboratorio di Terapia Cellulare Stefano Verri, Monza, Italy 5 Schneider Children’s Medical Center, Pediatric Hemato-Oncology, Israel 6 Hematology Institute, Sheba Medical Center, Tel Hashomer, Israel 7 Division of Oncology and Children’s Research Center, University Children’s Hospital, University of Zurich, Zurich, Switzerland 8 Department of Haematology, Westmead Childrens Hospital, Sydney, Australia 9 Department of Pediatrics, St. Anna Children’s Hospital and Children’s Cancer Research Institute, Medical University of Vienna, Vienna, Austria; coordinator of the AIEOP-BFM Flow Network
Background: Flow cytometry is a valuable part in the routine diagnostics of acute leukemia (AL). Although internationally recognized definitions of main AL subsets are available, there is currently no consensus format for the short summary of clinical flow cytometry reports. Since clinical reports are too long for most database purposes, there is a need for a standardized format of their short summaries. Methods: The Associazione Italiana Ematologia Oncologia Pediatrica—Berlin Frankfurt Muenster (AIEOP-BFM) Flow Network that encompasses reference diagnostics laboratories in Australia, Austria, Czechia, Germany, Israel, Italy, and Switzerland have designed a pro-forma for the summary of flow cytometry results in the diagnosis of leukemia. The process involved several meetings and other communications, during which the group established a consensus on the essentials that lead to the diagnostic conclusions in childhood AL. Results: The “Flow Diagnostics Essential (FDE) Code” is a result from an agreement within the AIEOP-BFM Flow Network. In a standardized format, it reports the extent of the infiltration by a malignant clone, followed by description antigen expression as strong, weak or negative, and a diagnostic conclusion. Conclusions: A consensus brief format (the “FDE Code”) has been designed as a brief summary of the diagnostic immunophenotype of childhood AL. It is also applicable for the diagnostic investigation of other malignancies by flow cytometry. The FDE code may be included in the final clinical report and/or C 2013 International Clinical Cytometry Society used in the setting of a multicenter clinical trial database. V Key terms: acute lymphoblastic leukemia; acute myeloid leukemia; cytometry; immunophenotyping; standardization; clinical reporting
*Correspondence to: Ondrej Hrus ak, MD, PhD, CLIP Cytometry Laboratory, Department of Pediatric Hematology and Oncology, V uvalu 84, 15006 Praha 5, Czech Republic. E-mail: Ondrej.Hrusak@lfmotol. cuni.cz
C 2013 International Clinical Cytometry Society V
Received 23 July 2013; Revised 30 October 2013; Accepted 31 October 2013 Published online 7 November 2013 in Wiley Online Library (wileyonlinelibrary.com). DOI: 10.1002/cyto.b.21144
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How to cite this article: Hrusak O, Basso G, Ratei R, Gaipa G, Luria D, Mejstrıkova E, Karawajew L, Buldini B, Rozenthal E, Bourquin JP, Kalina T, Sartor M, and Dworzak MN. Flow Diagnostics Essential Code: A Simple and Brief Format for the Summary of Leukemia Phenotyping. Cytometry Part B 2014; 86B: 288–291.
The growing complexity of cytometric investigation of leukemias partly contrasts with the need of clear and unequivocal reporting of its findings to the clinicians. At the beginning of the analysis, a cytometrist has raw data files in flow cytometry standard format, which contain information on each acquired event, both relevant and irrelevant, both cellular and debris, and cells of both malignant and nonmalignant origin. This is combined with other information that may affect the result of cytometric investigation, such as quality of the specimen, time duration between its collection and the acquisition to the cytometer, or possible technical problems related to the instrument or the laboratory preparation of the specimen (1). During the analysis of the cytometric data on a leukemic specimen, the cytometrist collects principal information on each antigen assessed on the malignant cells: whether it is positive, negative or partially positive, or that its expression cannot be determined conclusively. Additional details can be described on each antigen, such as the intensity of staining (bright, dim, negative), the staining distribution (heterogeneous, homogeneous), quantitative description of the histogram (percentage of cells in positive area or in various intensity regions, positions of a median, mode(s) or a mean), qualitative description of the histogram (unimodal, bimodal, trimodal etc., skewness of the curve, presence or absence of a gap between cell subsets, etc.), description of the relationships among several antigens (coexpression or mutual exclusivity, presence or absence of “empty spaces” etc.) and even more features may be described. Some of the characteristics of positivity may be combined and can lead to the distinction between “strong” and “weak” positivity, which is typically used in the World Health Organization (WHO) classification (2). Many of the details related to the quality and quantity of expression of each antigen may be considered. Nevertheless, the report should be finished with an unequivocal diagnostic conclusion. Unfortunately, routine clinical practice also brings examples of cytometric reports where the diagnostic conclusion is not clearly stated and is replaced by some of the aforementioned parameters. The resulting confusion has been previously criticized (3). The great variability among laboratory report formats has created a need of a condensed summary such as the Flow Diagnostics Essential (FDE) Code which is written in a unique format (examples in Table 1). The FDE code can be used in the diagnosis of leukemia, inter laboratory communication and in clinical trial databases. Laboratories that agree to the principle should use the FDE Code as part of their routine diagnostic reports that
Cytometry Part B: Clinical Cytometry
describe the immunophenotype of atypical (suspected malignant) cells. It starts with letters “FDE” followed by a colon, percentage of atypical cells (if necessary preceded with > or < signs), and a semicolon. It continues with a description of the expression of the investigated antigens followed by a simplified conclusion. Items other than the conclusion are not separated by spaces. The lists of antigens starts with those that are strongly expressed (i.e., antigens with unequivocal positivity on the majority of cells), followed by weakly expressed antigens (i.e., antigens that are positive on a relevant subset of blasts, however, either due to low percentage of cells or due to low intensity are not considered “strong”), and by nonexpressed antigens (considered “negative”). Each list of antigens starts with a designation (STRONG, WEAK, and NEGATIVE) and a colon followed by CD antigens listed in an ascending order, separated by commas, while the characters “CD” are only at the beginning of each list; following the CD antigens are other antigens in alphabetical order. Intracellularly assessed antigens or molecules are prefixed with an “i” and this prefix does not affect the alphanumeric order of the antigen. Antigens that could not be evaluated are typically not mentioned, unless they are required for the diagnosis definition (in such case, their list is labeled as “NOT EVALUATED”). Following the lists of antigens, the diagnostic classification is concluded in the briefest possible way (Table 2). In acute leukemias (AL), the diagnostic conclusions may be abbreviated, for example, as B-I to B-IV or T-I to T-IV (4) or, in acute myeloid leukemia (AML), the differentiation along the granulocytic, monocytic, erythroid or megakaryocytic lineages may be specified (5,6). Importantly, since only two-way classification of positive antigens is allowed in the FDE, all positive antigens that are not considered “WEAK” should be reported as “STRONG” and vice versa. “STRONG” expression thus refers to unequivocal positivity, not necessarily to bright expression. If the percentage of blasts is lower than conventional thresholds accepted for making the diagnosis of AL (usually 20–30% of nucleated cells) but the cytometric assessment unambiguously discerns their pathological immunophenotype, the conclusion should not anticipate a formal diagnosis of AL but state the observation of a pathological (or probably malignant) blast population suggestive of an AL with a low percentage of blasts. If malignant or atypical cells are not detected at all, then this is mentioned together with the level of sensitivity for such conclusion. The conclusion should state the leukemic lineage and subtype as per the differentiation. The definitions of the
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Table 1 Examples of FDE Codes Condition Intermediate T-ALL No blasts detected with a sensitivitya of 5% cALL or preB ALL with aberrant CD33 expression
Mature B cell ALL AML with aberrant CD19 expression
Pathological myeloid cell population, possibly malignant Bilineal leukemia
Biphenotypic AL (cALL with a high myeloid EGIL score)
Example of FDE Code FDE:41%;STRONG:CD1a,2,i3,4,7,8,45,iTdT,NEGATIVE: CD3,10,13,14,15,19,22,33,34,i79a,iMPO,TCRab,TCRgd; T-ALL T-III. FDE:
