CD200 has an important role in the differential diagnosis of mature B-cell neoplasms by multiparameter flow cytometry

2013 ◽  
pp. n/a-n/a ◽  
Author(s):  
Alex F. Sandes ◽  
Maria de Lourdes Chauffaille ◽  
Cláudia Regina M.C. Oliveira ◽  
Yumi Maekawa ◽  
Nair Tamashiro ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Differential Diagnosis ◽  
B Cell ◽  
Multiparameter Flow Cytometry

Multiparameter Flow Cytometry (MFC) for Identification of the Waldenström's Clone in IgM MGUS and Waldenstrom's Macroglobulinemia (WM): New Criteria for Differential Diagnosis and Risk Stratification

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 936-936
Author(s):  
Bruno Paiva ◽  
Maria-Carmen Montes ◽  
Ramón García-Sanz ◽  
Jennifer Alonso ◽  
Natalia de las Heras ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Differential Diagnosis ◽  
B Cells ◽  
B Cell ◽  
Light Chain ◽  
Conflicts Of Interest ◽  
Phenotypic Characterization ◽  
International Prognostic Scoring System ◽  
Multiparameter Flow Cytometry ◽  
Risk Of Progression

Abstract Abstract 936 Demonstration of bone marrow (BM) infiltration by lymphoplasmacytic lymphoma is essential to the diagnosis of WM, and a trephine biopsy is considered mandatory for this assessment. Multiparameter flow cytometry (MFC) has demonstrated its clinical relevance in MGUS and myeloma; however, immunophenotypic studies on IgM monoclonal gammopathies are scanty, and focus only in patients with WM. Herein, MFC immunophenotyping was performed on BM samples from 244 patients, including 67 IgM MGUS, 77 smoldering, and 100 symptomatic WM newly diagnosed patients according to the Second International Workshop. A four color panel that systematically allowed the identification of B cells and plasma cells (PC), and their phenotypic characterization for a total of 24 antigens was used. We first analyzed the percentage of B cells and PC in BM and the percentage of light chain restricted cells in both compartments. Our results show a progressive increment of B cells from IgM MGUS to smoldering and symptomatic WM (medians of 2%, 9% and 12%; P<.001), as well of light chain restricted B cells (75%, 96% and 99%; P<.001). In contrast, no differences were found for the percentage of PC (median of 0.3%), but light chain restricted PC progressively increased from IgM MGUS to smoldering and symptomatic WM (70%, 85% and 97%; P<.001). Accordingly, only 1% of IgM MGUS patients showed >10% B cells, in contrast to 34% and 55% of smoldering and symptomatic WM (P<.001). Likewise, only 1% of IgM MGUS patients showed 100% light chain restricted B cells, in contrast to 19% and 40% of smoldering and symptomatic WM (P<.001); similar results being also found using a cutoff of 100% light chain restricted PC. Subsequently, we explored whether the percentages of BM and light chain restricted B cells and PC could predict time to progression (TTP) from smoldering into symptomatic WM, as well as overall survival (OS) in symptomatic WM. In smoldering WM, B cells (>10% vs ≤10%: median TTP of 47m vs 145m; P=.016) and light chain restricted B cells (100% vs <100%: 26m vs 145m; P<.001) but not PC, predicted risk of progression. On the multivariate analysis that included serum M-spike (±3g/dL), BM infiltration (±50% lymphoplasmacytic cells), BM B-cells and light chain restricted B cells (by MFC), only the later retained independent prognostic value (HR: 19.8, P=.001). Upon analyzing factors influencing survival in symptomatic WM patients, cases with >10% B cells showed a trend for inferior OS (P=.080), and significant differences emerged when comparing patients with 100% vs <100% light chain restricted B cells (median OS 44m vs 78m; P=.001). The later marker was independent (HR: 2.6; P=.004) of the International Prognostic Scoring System (HR: 2.2; P=.006). Focusing on the antigenic profiles of B cells and PC, we noted that within the B-cell compartment there was a progressive increment of CD22dim (69%, 92% and 88%; P<.001), CD25+ (61%, 88% and 90%; P<.001) and sIgM+ (88%, 95% and 97%; P=.002) B cells from IgM MGUS to smoldering and symptomatic WM. This underlies that the accumulating light chain restricted clonal B cells show a characteristic Waldenstrom's phenotype (CD22dim/CD25+/IgM+). Of note, a bimodal (from - to +) expression for the B cell memory marker CD27 was found in >50% of WM patients, which raises the possibility that the WM clone may arise, at least in some cases, before antigenic stimulation; subsequent maturation of the clone into PC would explain the typical presence of somatic hypermutations. On the other hand, B-cells from IgM MGUS and WM patients were negative in ≥90% of cases for CD5, CD10, CD11c and CD103, which can be useful to differentiate between WM and other B-NHL. Finally, the antigenic profile of PC in IgM MGUS and WM was similar to that of normal PC, and different from myeloma PC by consistently showing a CD27+ and CD56- phenotype, in addition to sIgM+ expression in ≥87% of all cases. Similarly to B-cells, we also noted that within the PC compartment there was a progressive increment of CD19+, CD45+ and sIgM+ CD20+ PC from IgM MGUS to smoldering and symptomatic WM. This underlies that this transition is asssociated with an accumulation of light chain restricted clonal PC displaying an immature/plasmablastic phenotype. In summary, our results highlight the potential value of MFC immunophenotyping for the characterization of the Waldenström's clone, as well as for the differential diagnosis, risk of progression and survival in WM. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Expert-independent classification of mature B-cell neoplasms using standardized flow cytometry: a multicentric study

2021 ◽  
Author(s):  
Sebastian Böttcher ◽  
Robby Engelmann ◽  
Georgiana Grigore ◽  
Paula Carolina Fernandez ◽  
Joana Caetano ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Differential Diagnosis ◽  
Follicular Lymphoma ◽  
B Cell ◽  
Diagnostic Algorithm ◽  
Lymphocytic Leukemia ◽  
B Cell Lymphomas ◽  
Multicentric Study ◽  
Predictive Values ◽  
Large B Cell

Reproducible expert-independent flow-cytometric criteria for the differential diagnoses between mature B-cell neoplasms are lacking. We developed an algorithm-driven classification for these lymphomas by flow cytometry and compared it to the WHO gold standard diagnosis. Overall, 662 samples from 662 patients representing nine disease categories were analyzed at 9 laboratories using the previously published EuroFlow 5-tube-8-color B-cell chronic lymphoproliferative disease antibody panel. Expression levels of all 26 markers from the panel were plotted by B-cell entity to construct a univariate, fully standardized diagnostic reference library. For multivariate data analysis we subsequently utilized Canonical Correlation Analysis of 176 training cases to project the multi-dimensional space of all 26 immunophenotypic parameters into 36 two-dimensional plots for each possible pair-wise differential diagnosis. Diagnostic boundaries were fitted according to the distribution of the immunophenotypes of a given differential diagnosis. A diagnostic algorithm based on these projections was developed and subsequently validated using 486 independent cases. Negative predictive values exceeding 92.1% were observed for all disease categories except for follicular lymphoma. Particularly high positive predictive values were returned in chronic lymphocytic leukemia (99.1%), hairy cell leukemia (97.2%), follicular lymphoma (97.2%) and mantle cell lymphoma (95.4%). Burkitt and CD10+ diffuse large B-cell lymphomas were difficult to distinguish by the algorithm. A similar ambiguity was observed between marginal zone, lymphoplasmacytic, and CD10- diffuse large B-cell lymphomas. The specificity of the approach exceeded 98% for all entities. The univariate immunophenotypic library and the multivariate expert-independent diagnostic algorithm might contribute to increased reproducibility of future diagnostics in mature B-cell neoplasms.

Detection of t(14;18)(q32;q21) in B-Cell Chronic Lymphocytic Leukemia

2005 ◽  
Vol 129 (3) ◽  
pp. 410-411
Author(s):  
Wolfgang Kern ◽  
Torsten Haferlach ◽  
Susanne Schnittger ◽  
Claudia Schoch
Keyword(s):  
Flow Cytometry ◽  
In Situ Hybridization ◽  
Chronic Lymphocytic Leukemia ◽  
Follicular Lymphoma ◽  
B Cell ◽  
Lymphocytic Leukemia ◽  
Multiparameter Flow Cytometry ◽  
Lymphoma Therapy ◽  
Cell Chronic Lymphocytic Leukemia

Abstract Cytomorphologic testing and multiparameter flow cytometry are the mainstays in diagnosing B-cell chronic lymphocytic leukemia, whereas fluorescence in situ hybridization that targets the translocation t(14;18)(q32;q21) often is used to identify follicular lymphoma. Therapy is highly diverse between both diseases. We describe a case with cytomorphologically and immunologically proven B-cell chronic lymphocytic leukemia in which t(14;18)(q32;q21) was found.

scholarly journals Hematologist‐Level Classification of Mature B‐Cell Neoplasm Using Deep Learning on Multiparameter Flow Cytometry Data

2020 ◽  
Vol 97 (10) ◽  
pp. 1073-1080
Author(s):  
Max Zhao ◽  
Nanditha Mallesh ◽  
Alexander Höllein ◽  
Richard Schabath ◽  
Claudia Haferlach ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Deep Learning ◽  
B Cell ◽  
Multiparameter Flow Cytometry ◽  
Flow Cytometry Data ◽  
Cell Neoplasm

Utility of Multiparameter Flow Cytometry Immunophenotypic Studies In Patients with Systemic Light Chain (AL) Amyloidosis

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 4051-4051
Author(s):  
Bruno Paiva ◽  
María-Belén Vidriales ◽  
Jose J. Perez ◽  
Maria-Consuelo López-Berges ◽  
Ramón García-Sanz ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Flow Cytometry ◽  
Differential Diagnosis ◽  
Plasma Cell ◽  
Light Chain ◽  
Cell Cycle Analysis ◽  
Disease Assessment ◽  
Multiparameter Flow Cytometry ◽  
Al Amyloidosis

Abstract Abstract 4051 Multiparameter flow cytometry (MFC) immunophenotyping has shown to be of value for differential diagnosis and minimal residual disease assessment in multiple myeloma. However, the clinical value of MFC immunophenotyping in other plasma cell disorders (PCD) remains largely unexplored. Systemic light chain (AL) amyloidosis is a rare PCD characterized by the accumulation of monoclonal light chain fragments leading to end-organ damage and short survival. Bone marrow (BM) plasma cell (PC) infiltration in AL is usually low and thus the identification of clonal PC can be often difficult by immunohistochemistry and/or immunofluorescence. In the present study we focused on 34 BM samples sent to our institution with a suspected diagnosis of AL. MFC immunophenotypic studies were performed using the following 4-color combinations of MoAbs (FITC/PE/PerCP-Cy5.5/APC): CD38/CD56/CD19/CD45 (n=34); in addition cy-Kappa/cy-Lambda/CD19/CD38 staining was add to confirm the clonal or polyclonal nature of BMPC in equivocal cases. Ploidy and cell cycle analysis were additionally performed in a subset of cases (n=12/34). From the total 34 cases included in the present study, 28 had a confirmed diagnosis of AL. The remaining 6 cases were finally diagnosed with localized - amyloidoma - (n=2) and familial (n=1) forms of amyloidosis, multiple myeloma-associated amyloid (n=2) and congestive pericarditis (n=1). Interestingly, the presence of clonal PC was detected by MFC in 27 of the 28 (96%) patients with AL; in turn, clonal PC were undetectable in the BM of all cases with localized and familial forms of amyloidosis. The median overall level of PC (M-PC plus N-PC) seen in MFC immunophenotypic analyses of BM samples of the 28 patients with AL was 1.9% (range: 0.1% - 15%), with a significant positive correlation between PC enumerated by MFC and conventional morphology (r=0.5; p=.01). Within the BMPC compartment, the median proportion of clonal PC was of 94% (mean 81% ± 29%); in 6 cases all BMPC were clonal while in the remaining 22 patients residual normal PC persisted (median of normal PC/BMPC 13% ± 31%). The most common aberrant phenotypes were down-regulation of CD19 (92%) and CD45 (83%), followed by overexpression of CD56 (56%) and infra-expression of CD38 (42%). Aneuploidy was only found in 18% of cases, all of them hyperdiploid. Cell cycle analysis showed a median % of S-phase and G2-Mitosis PC of 0.7% and 3.5%, respectively. Concerning patients' outcome, cases with undetectable normal PC (6/28, 21%) had a significantly decreased overall survival (OS) compared to patients with persistent BM normal PC at diagnosis (22/28, 79%) with 3-year OS rates of 0% vs. 59%, respectively (p=.001). In summary, these preliminary data suggests that MFC immunophenotyping investigations may be clinically relevant in patients with suspected amyloidosis for i) differential diagnosis between AL and other forms of amyloidosis and, ii) prognostication of patients with AL according to the presence or absence of baseline persistent normal PC. Disclosures: No relevant conflicts of interest to declare.

Multicentric Analyses of the CD148, CD180, and CD200 Combination for the Diagnosis of Mature B-Cell Neoplasm Using Flow Cytometry

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2662-2662 ◽  
Author(s):  
Laurent Miguet ◽  
Luc Fornecker ◽  
Marie Wyrwas ◽  
Sarah Cianferani ◽  
Raoul Herbrecht ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Differential Diagnosis ◽  
B Cell ◽  
Expression Profile ◽  
Standard Error ◽  
Conflicts Of Interest ◽  
Lymphocytic Leukemia ◽  
Strong Expression ◽  
Group A ◽  
Weak Expression

Abstract Introduction Diagnosis of mature B-cells proliferations, especially those involving the spleen, do not always falls into any of the WHO types of B-cell neoplasms using standart diagnosis tools. This situation in notably encountered in the case of the differential diagnosis of marginal zone lymphoma (MZL), atypical chronic lymphocytic leukemia (aCLL), mantle cell lymphoma (MCL), and lymphoplasmacytic lymphoma (LPL), mostly due to the lack of immunological positive markers. In order to find new markers to discriminate between these different malignancies, we have previously developed a proteomic strategy based on the analyses of plasma membrane microparticles and proposed two new specific markers: CD148 and CD1801,2 for MCL and MZL respectively. The simultaneous use of these two markers, together with the CD200 that is positive in most cases of CLL and negative in MCL could be of great interest to better assess the differential diagnosis. Methods Flow cytometry analyses have been realized in Nancy and Strasbourg hospitals by combining these three markers: CD148 (Clone 143-41 FITC); CD180 (Clone G28.8 PE) and CD200 (Clone OX104 APC). Expression profile of these proteins was established on a well characterized set of patients (N=287): CLL with a Matutes score > 3 (N=81); MCL harboring t(11;14) translocation or CCND1 overexpression (N=44); LPL (N=58) classified following cytological morphology, IgM peak and the positivity of CD38 and/or Myd88 mutation, MZL (N=84), displaying a CD5- CD23- immunophenotype associated to a splenomegaly and 20 controls. For each group the mean of fluorescence intensity and Standard Error have been determined. Results MCL exhibited a strong expression of CD148 combined with a weak expression of CD180 and CD200. A weak expression of CD148 and CD180 coupled to a strong expression of CD200 was typical of the CLL group and a weak expression of CD148 and CD200 coupled to a strong expression of CD180 was observed in the MZL group. A moderate expression of these three markers was observed in the LPL group. A threshold corresponding to MFI +/- 4 standard error was then calculated for each group, and patients were categorized following the expression profile of these 3 markers (see figures). In this cohort, the above described profiles correctly identified MCL cases with a specificity of 92% and a sensitivity of 64%, aCLL cases with a specificity of 100% and a sensitivity of 47%, LPL cases with a specificity of 90% and a sensitivity of 54% and MZL cases with a specificity of 99% and a sensitivity of 60%. Conclusion These results strongly suggest that the incorporation of these three markers CD148 CD180 and CD200 in addition of the routinely used flow cytometry panel can be helpful in a number of cases for which the diagnosis is difficult. References: 1) Miguet et al leukemia 2013 2) Miguet et al journal of proteome research 2009 Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

Multiparameter flow cytometry in the differential diagnosis of aberrant T-cell clones of unclear significance

2014 ◽  
Vol 56 (3) ◽  
pp. 639-644 ◽  
Author(s):  
Anna Flammiger ◽  
Ulrike Bacher ◽  
Maximilian Christopeit ◽  
Christiane Horn ◽  
Elke Rühlmann ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Differential Diagnosis ◽  
T Cell ◽  
Multiparameter Flow Cytometry ◽  
T Cell Clones ◽  
Cell Clones

Flow Cytometric Analysis of Acute Leukemias

2003 ◽  
Vol 127 (1) ◽  
pp. 42-48 ◽  
Author(s):  
Zahid Kaleem ◽  
Eric Crawford ◽  
M. Hanif Pathan ◽  
Leah Jasper ◽  
Michael A. Covinsky ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Acute Leukemia ◽  
B Cell ◽  
Flow Cytometric Analysis ◽  
Multiparameter Flow Cytometry ◽  
Correct Identification ◽  
Acute Leukemias ◽  
Cytochemical Analysis ◽  
Flow Cytometric ◽  
Hla Dr

Abstract Context.—Acute leukemia displays characteristic patterns of surface antigen expression (CD antigens), which facilitate their identification and proper classification and hence play an important role in instituting proper treatment plans. In addition to enzyme cytochemical analysis, multiparameter flow cytometric analysis has become commonplace in most laboratories for that purpose. The essential role and caveats of flow cytometry in that regard, however, have received little scrutiny. Objective.—To evaluate the expression of commonly used immunomarkers and patterns in various acute leukemias to help define the best use and role of multiparameter flow cytometry in the diagnosis and proper classification of acute leukemias. Design.—We have retrospectively analyzed the immunophenotypic data from 508 de novo adult and pediatric acute leukemia patients, as studied using multiparameter flow cytometry in addition to routine morphologic and enzyme cytochemical analysis. Cytogenetic and/or molecular data were correlated in all 41 cases of acute promyelocytic leukemia (APL) and in 203 other cases of acute leukemia where those data were available. We have also determined the positive and negative predictive values of a combined CD34 and HLA-DR expression pattern for the differentiation of APL from other myeloid leukemias. Results.—In acute myeloid leukemia (AML) other than APL, expression of CD34 was seen in 62% and expression of HLA-DR in 86% of all cases. Twenty-six (10%) of 259 cases of non-APL AML were negative for both CD34 and HLA-DR as opposed to 33 (80%) of 41 cases of APL. None of the cases of APL were positive for both CD34 and HLA-DR in contrast to 149 (58%) of 259 cases of non-APL AML. Fifty-three cases were found to be examples of minimally differentiated AML (AML M0) based on the lack of expression of cytoplasmic CD3 and cytoplasmic CD79a and expression of one or more myelomonocytic-associated antigens and/or myeloperoxidase. Expression of CD20 was seen in 40 (24%) of 168 cases of precursor B-cell acute lymphoblastic leukemia (pB-ALL) and 52 (29%) lacked CD34 expression. Five of 180 cases of pB-ALL and 2 cases of precursor T-cell ALL (pT-ALL) were negative for terminal deoxynucleotidyl transferase (TdT). Aside from cytoplasmic CD3, CD5 and CD7 were the most sensitive antigens present in all 21 cases of pT-ALL. CD33 was more sensitive but less specific than CD13 for myeloid lineage. Conclusion.—Aside from identification of blasts, flow cytometry was found to be especially useful in the correct identification of AML M0, differentiation of APL from AML M1/M2, and correct identification of TdT-negative ALL and unusual variants, such as transitional B-cell ALL and undifferentiated and biphenotypic acute leukemias.

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