Research

16 pages, 1795 KiB

Open AccessArticle

CD20 Expression as a Possible Novel Prognostic Marker in CLL: Application of EuroFlow Standardization Technique and Normalization Procedures in Flow Cytometric Expression Analysis

by Anke Schilhabel, Peter Jonas Walter, Paula Cramer, Julia von Tresckow, Saskia Kohlscheen, Monika Szczepanowski, Anna Laqua, Kirsten Fischer, Barbara Eichhorst, Sebastian Böttcher, Christof Schneider, Eugen Tausch, Monika Brüggemann, Michael Kneba, Michael Hallek and Matthias Ritgen

Cancers 2022, 14(19), 4917; https://doi.org/10.3390/cancers14194917 - 07 Oct 2022

Cited by 3 | Viewed by 1674

Abstract

Background: CD20 expression is a controversial issue regarding response prediction to anti-CD20 therapy in chronic lymphocytic leukemia (CLL). Methods: Median fluorescence intensities (MFIs) of standard fluorescence beads from the daily calibration of flow cytometers according to EuroFlow protocols were used to establish a [...] Read more.

Background: CD20 expression is a controversial issue regarding response prediction to anti-CD20 therapy in chronic lymphocytic leukemia (CLL). Methods: Median fluorescence intensities (MFIs) of standard fluorescence beads from the daily calibration of flow cytometers according to EuroFlow protocols were used to establish a normalization approach to study CD20 expression on CLL cells. CD20 MFI was retrospectively assessed prior to and during treatment from flow cytometric measurements of peripheral blood in patients with different depths of molecular response in the four phase-II CLL2-BXX trials (BIG; BAG; BIO; BCG; N = 194) administering either Obinutuzumab or Ofatumumab in combination with targeted agents. Results: No significant difference was observed between the normalized and measured MFIs of CD19 and CD20 on CLL cells. During treatment, CD20 expression levels on CLL cells did not significantly differ between the four investigated different treatment schemes, but a strong molecular response to Ofatumumab seemed to correlate with higher CD20 expression prior to therapy. Conclusions: Standardized staining and instrument monitoring enable a robust assessment of longitudinal biological variations of marker expression based on MFI values. Obinutuzumab showed a higher proportion of patients with a strong MRD response independent from initial CD20 expression, whereas high pre-therapeutic CD20 expression levels seem to correlate with a profound response to Ofatumumab. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

18 pages, 17795 KiB

Open AccessArticle

Bone Marrow Stromal Cell Regeneration Profile in Treated B-Cell Precursor Acute Lymphoblastic Leukemia Patients: Association with MRD Status and Patient Outcome

by Elen Oliveira, Elaine S. Costa, Juana Ciudad, Giuseppe Gaipa, Łukasz Sedek, Susana Barrena, Tomasz Szczepanski, Chiara Buracchi, Daniela Silvestri, Patrícia F. R. Siqueira, Fabiana V. Mello, Rafael C. Torres, Leonardo M. R. Oliveira, Isabelle V. C. Fay-Neves, Edwin Sonneveld, Vincent H. J. van der Velden, Esther Mejstrikova, Josep-Maria Ribera, Valentino Conter, Martin Schrappe, Jacques J. M. van Dongen, Marcelo G. P. Land and Alberto Orfao Show full author list Hide full author list

Cancers 2022, 14(13), 3088; https://doi.org/10.3390/cancers14133088 - 23 Jun 2022

Cited by 4 | Viewed by 2061

Abstract

For the last two decades, measurable residual disease (MRD) has become one of the most powerful independent prognostic factors in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, the effect of therapy on the bone marrow (BM) microenvironment and its potential relationship with the [...] Read more.

For the last two decades, measurable residual disease (MRD) has become one of the most powerful independent prognostic factors in B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, the effect of therapy on the bone marrow (BM) microenvironment and its potential relationship with the MRD status and disease free survival (DFS) still remain to be investigated. Here we analyzed the distribution of mesenchymal stem cells (MSC) and endothelial cells (EC) in the BM of treated BCP-ALL patients, and its relationship with the BM MRD status and patient outcome. For this purpose, the BM MRD status and EC/MSC regeneration profile were analyzed by multiparameter flow cytometry (MFC) in 16 control BM (10 children; 6 adults) and 1204 BM samples from 347 children and 100 adult BCP-ALL patients studied at diagnosis (129 children; 100 adults) and follow-up (824 childhood samples; 151 adult samples). Patients were grouped into a discovery cohort (116 pediatric BCP-ALL patients; 338 samples) and two validation cohorts (74 pediatric BCP-ALL, 211 samples; and 74 adult BCP-ALL patients; 134 samples). Stromal cells (i.e., EC and MSC) were detected at relatively low frequencies in all control BM (16/16; 100%) and in most BCP-ALL follow-up samples (874/975; 90%), while they were undetected in BCP-ALL BM at diagnosis. In control BM samples, the overall percentage of EC plus MSC was higher in children than adults (p = 0.011), but with a similar EC/MSC ratio in both groups. According to the MRD status similar frequencies of both types of BM stromal cells were detected in BCP-ALL BM studied at different time points during the follow-up. Univariate analysis (including all relevant prognostic factors together with the percentage of stromal cells) performed in the discovery cohort was used to select covariates for a multivariate Cox regression model for predicting patient DFS. Of note, an increased percentage of EC (>32%) within the BCP-ALL BM stromal cell compartment at day +78 of therapy emerged as an independent unfavorable prognostic factor for DFS in childhood BCP-ALL in the discovery cohort—hazard ratio (95% confidence interval) of 2.50 (1–9.66); p = 0.05—together with the BM MRD status (p = 0.031). Further investigation of the predictive value of the combination of these two variables (%EC within stromal cells and MRD status at day +78) allowed classification of BCP-ALL into three risk groups with median DFS of: 3.9, 3.1 and 1.1 years, respectively (p = 0.001). These results were confirmed in two validation cohorts of childhood BCP-ALL (n = 74) (p = 0.001) and adult BCP-ALL (n = 40) (p = 0.004) treated at different centers. In summary, our findings suggest that an imbalanced EC/MSC ratio in BM at day +78 of therapy is associated with a shorter DFS of BCP-ALL patients, independently of their MRD status. Further prospective studies are needed to better understand the pathogenic mechanisms involved. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

15 pages, 2663 KiB

Open AccessArticle

Quality Assessment of a Large Multi-Center Flow Cytometric Dataset of Acute Myeloid Leukemia Patients—A EuroFlow Study

by Anne E. Bras, Sergio Matarraz, Stefan Nierkens, Paula Fernández, Jan Philippé, Carmen-Mariana Aanei, Fabiana Vieira de Mello, Leire Burgos, Alita J. van der Sluijs-Gelling, Georgiana Emilia Grigore, Jacques J. M. van Dongen, Alberto Orfao, Vincent H. J. van der Velden and on behalf of the EuroFlow Consortium

Cancers 2022, 14(8), 2011; https://doi.org/10.3390/cancers14082011 - 15 Apr 2022

Cited by 3 | Viewed by 1960

Abstract

Flowcytometric analysis allows for detailed identification and characterization of large numbers of cells in blood, bone marrow, and other body fluids and tissue samples and therefore contributes to the diagnostics of hematological malignancies. Novel data analysis tools allow for multidimensional analysis and comparison [...] Read more.

Flowcytometric analysis allows for detailed identification and characterization of large numbers of cells in blood, bone marrow, and other body fluids and tissue samples and therefore contributes to the diagnostics of hematological malignancies. Novel data analysis tools allow for multidimensional analysis and comparison of patient samples with reference databases of normal, reactive, and/or leukemia/lymphoma patient samples. Building such reference databases requires strict quality assessment (QA) procedures. Here, we compiled a dataset and developed a QA methodology of the EuroFlow Acute Myeloid Leukemia (AML) database, based on the eight-color EuroFlow AML panel consisting of six different antibody combinations, including four backbone markers. In total, 1142 AML cases and 42 normal bone marrow samples were included in this analysis. QA was performed on 803 AML cases using multidimensional analysis of backbone markers, as well as tube-specific markers, and data were compared using classical analysis employing median and peak expression values. Validation of the QA procedure was performed by re-analysis of >300 cases and by running an independent cohort of 339 AML cases. Initial evaluation of the final cohort confirmed specific immunophenotypic patterns in AML subgroups; the dataset therefore can reliably be used for more detailed exploration of the immunophenotypic variability of AML. Our data show the potential pitfalls and provide possible solutions for constructing large flowcytometric databases. In addition, the provided approach may facilitate the building of other databases and thereby support the development of novel tools for (semi)automated QA and subsequent data analysis. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

16 pages, 2022 KiB

Open AccessArticle

Immunophenotypic Analysis of Acute Megakaryoblastic Leukemia: A EuroFlow Study

by Nienke Brouwer, Sergio Matarraz, Stefan Nierkens, Mattias Hofmans, Michaela Nováková, Elaine Sobral da Costa, Paula Fernandez, Anne E. Bras, Fabiana Vieira de Mello, Ester Mejstrikova, Jan Philippé, Georgiana Emilia Grigore, Carlos E. Pedreira, Jacques J. M. van Dongen, Alberto Orfao, Vincent H. J. van der Velden and on behalf of the EuroFlow Consortium

Cancers 2022, 14(6), 1583; https://doi.org/10.3390/cancers14061583 - 21 Mar 2022

Cited by 11 | Viewed by 3321

Abstract

Acute megakaryoblastic leukemia (AMKL) is a rare and heterogeneous subtype of acute myeloid leukemia (AML). We evaluated the immunophenotypic profile of 72 AMKL and 114 non-AMKL AML patients using the EuroFlow AML panel. Univariate and multivariate/multidimensional analyses were performed to identify most relevant [...] Read more.

Acute megakaryoblastic leukemia (AMKL) is a rare and heterogeneous subtype of acute myeloid leukemia (AML). We evaluated the immunophenotypic profile of 72 AMKL and 114 non-AMKL AML patients using the EuroFlow AML panel. Univariate and multivariate/multidimensional analyses were performed to identify most relevant markers contributing to the diagnosis of AMKL. AMKL patients were subdivided into transient abnormal myelopoiesis (TAM), myeloid leukemia associated with Down syndrome (ML-DS), AML—not otherwise specified with megakaryocytic differentiation (NOS-AMKL), and AMKL—other patients (AML patients with other WHO classification but with flowcytometric features of megakaryocytic differentiation). Flowcytometric analysis showed good discrimination between AMKL and non-AMKL patients based on differential expression of, in particular, CD42a.CD61, CD41, CD42b, HLADR, CD15 and CD13. Combining CD42a.CD61 (positive) and CD13 (negative) resulted in a sensitivity of 71% and a specificity of 99%. Within AMKL patients, TAM and ML-DS patients showed higher frequencies of immature CD34+/CD117+ leukemic cells as compared to NOS-AMKL and AMKL-Other patients. In addition, ML-DS patients showed a significantly higher expression of CD33, CD11b, CD38 and CD7 as compared to the other three subgroups, allowing for good distinction of these patients. Overall, our data show that the EuroFlow AML panel allows for straightforward diagnosis of AMKL and that ML-DS is associated with a unique immunophenotypic profile. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

18 pages, 3075 KiB

Open AccessArticle

Impact of Pre-Analytical and Analytical Variables Associated with Sample Preparation on Flow Cytometric Stainings Obtained with EuroFlow Panels

by Łukasz Sędek, Juan Flores-Montero, Alita van der Sluijs, Jan Kulis, Jeroen te Marvelde, Jan Philippé, Sebastian Böttcher, Marieke Bitter, Joana Caetano, Vincent H. J. van der Velden, Edwin Sonneveld, Chiara Buracchi, Ana Helena Santos, Margarida Lima, Tomasz Szczepański, Jacques J. M. van Dongen and Alberto Orfao

Cancers 2022, 14(3), 473; https://doi.org/10.3390/cancers14030473 - 18 Jan 2022

Cited by 3 | Viewed by 2581

Abstract

Objective interpretation of FC results may still be hampered by limited technical standardization. The EuroFlow consortium conducted a series of experiments to determine the impact of different variables on the relative distribution and the median fluorescence intensity (MFI) of markers stained on different [...] Read more.

Objective interpretation of FC results may still be hampered by limited technical standardization. The EuroFlow consortium conducted a series of experiments to determine the impact of different variables on the relative distribution and the median fluorescence intensity (MFI) of markers stained on different cell populations, from both healthy donors and patients’ samples with distinct hematological malignancies. The use of different anticoagulants; the time interval between sample collection, preparation, and acquisition; pH of washing buffers; and the use of cell surface membrane-only (SM) vs. cell surface plus intracytoplasmic (SM+CY) staining protocols, were evaluated. Our results showed that only monocytes were represented at higher percentages in EDTA- vs. heparin-anticoagulated samples. Application of SM or SM+CY protocols resulted in slight differences in the percentage of neutrophils and debris determined only with particular antibody combinations. In turn, storage of samples for 24 h at RT was associated with greater percentage of debris and cell doublets when the plasma cell disorder panel was used. Furthermore, 24 h storage of stained cells at RT was selectively detrimental for MFI levels of CD19 and CD45 on mature B- and T-cells (but not on leukemic blasts, clonal B- and plasma cells, neutrophils, and NK cells). The obtained results showed that the variables evaluated might need to be tailored for sample and cell type(s) as well as to the specific markers compared; however, defining of well-balanced boundaries for storage time, staining-to-acquisition delay, and pH of washing buffer would be a valid recommendation for most applications and circumstances described herein. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

10 pages, 3893 KiB

Open AccessCommunication

High-Sensitive TRBC1-Based Flow Cytometric Assessment of T-Cell Clonality in Tαβ-Large Granular Lymphocytic Leukemia

by Noemí Muñoz-García, F. Javier Morán-Plata, Neus Villamor, Margarida Lima, Susana Barrena, Sheila Mateos, Carolina Caldas, Jacques J. M. van Dongen, Alberto Orfao and Julia Almeida

Cancers 2022, 14(2), 408; https://doi.org/10.3390/cancers14020408 - 14 Jan 2022

Cited by 10 | Viewed by 2354

Abstract

Flow cytometric (FCM) analysis of the constant region 1 of the T-cell receptor β chain (TRBC1) expression for assessing Tαβ-cell clonality has been recently validated. However, its utility for the diagnosis of clonality of T-large granular lymphocytic leukemia (T-LGLL) needs to be confirmed, [...] Read more.

Flow cytometric (FCM) analysis of the constant region 1 of the T-cell receptor β chain (TRBC1) expression for assessing Tαβ-cell clonality has been recently validated. However, its utility for the diagnosis of clonality of T-large granular lymphocytic leukemia (T-LGLL) needs to be confirmed, since more mature Tαβ cells (i.e., T-LGL normal-counterpart) show broader TRBC1⁺/TRBC1⁻ ratios vs. total Tαβ cells. We compared the distribution and absolute counts of TRBC1⁺ and TRBC1⁻ Tαβ-LGL in blood containing polyclonal (n = 25) vs. clonal (n = 29) LGL. Overall, polyclonal TRBC1⁺ or TRBC1⁻ Tαβ-LGL ranged between 0.36 and 571 cells/μL (3.2–91% TRBC1⁺ cells), whereas the clonal LGL cases showed between 51 and 11,678 cells/μL (<0.9% or >96% TRBC1⁺ cells). Among the distinct TCRVβ families, the CD28⁻ effector-memory and terminal-effector polyclonal Tαβ cells ranged between 0 and 25 TRBC1⁺ or TRBC1⁻ cells/μL and between 0 and 100% TRBC1⁺ cells, while clonal LGL ranged between 32 and 5515 TRBC1⁺ or TRBC1⁻ cells/μL, representing <1.6% or >98% TRBC1⁺ cells. Our data support the utility of the TRBC1-FCM assay for detecting T-cell clonality in expansions of Tαβ-LGL suspected of T-LGLL based on altered percentages of TRBC1⁺ Tαβ cells. However, in the absence of lymphocytosis or in the case of TαβCD4-LGL expansion, the detection of increased absolute cell counts by the TRBC1-FCM assay for more accurately defined subpopulations of Tαβ-LGL-expressing individual TCRVβ families, allows the detection of T-cell clonality, even in the absence of phenotypic aberrations. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

27 pages, 3434 KiB

Open AccessArticle

Flow Cytometry Immunophenotyping for Diagnostic Orientation and Classification of Pediatric Cancer Based on the EuroFlow Solid Tumor Orientation Tube (STOT)

by Cristiane de Sá Ferreira-Facio, Vitor Botafogo, Patrícia Mello Ferrão, Maria Clara Canellas, Cristiane B. Milito, Sérgio Romano, Daiana V. Lopes, Lisandra C. Teixeira, Elen Oliveira, Enrico Bruno-Riscarolli, Fabiana V. Mello, Patrícia F. R. Siqueira, Patrícia Moura, Francisco Nicanor Macedo, Danielle N. Forny, Luíza Simião, Ana Luíza Pureza, Marcelo Gerardin Poirot Land, Carlos Eduardo Pedreira, Jacques J. M. van Dongen, Alberto Orfao and Elaine Sobral da Costa Show full author list Hide full author list

Cancers 2021, 13(19), 4945; https://doi.org/10.3390/cancers13194945 - 30 Sep 2021

Cited by 5 | Viewed by 6257

Abstract

Early diagnosis of pediatric cancer is key for adequate patient management and improved outcome. Although multiparameter flow cytometry (MFC) has proven of great utility in the diagnosis and classification of hematologic malignancies, its application to non-hematopoietic pediatric tumors remains limited. Here we designed [...] Read more.

Early diagnosis of pediatric cancer is key for adequate patient management and improved outcome. Although multiparameter flow cytometry (MFC) has proven of great utility in the diagnosis and classification of hematologic malignancies, its application to non-hematopoietic pediatric tumors remains limited. Here we designed and prospectively validated a new single eight-color antibody combination—solid tumor orientation tube, STOT—for diagnostic screening of pediatric cancer by MFC. A total of 476 samples (139 tumor mass, 138 bone marrow, 86 lymph node, 58 peripheral blood, and 55 other body fluid samples) from 296 patients with diagnostic suspicion of pediatric cancer were analyzed by MFC vs. conventional diagnostic procedures. STOT was designed after several design–test–evaluate–redesign cycles based on a large panel of monoclonal antibody combinations tested on 301 samples. In its final version, STOT consists of a single 8-color/12-marker antibody combination (CD99-CD8/_numyogenin/CD4-EpCAM/CD56/GD2/_smCD3-CD19/_cyCD3-CD271/CD45). Prospective validation of STOT in 149 samples showed concordant results with the patient WHO/ICCC-3 diagnosis in 138/149 cases (92.6%). These included: 63/63 (100%) reactive/disease-free samples, 43/44 (98%) malignant and 4/4 (100%) benign non-hematopoietic tumors together with 28/38 (74%) leukemia/lymphoma cases; the only exception was Hodgkin lymphoma that required additional markers to be stained. In addition, STOT allowed accurate discrimination among the four most common subtypes of malignant CD45⁻ CD56⁺⁺ non-hematopoietic solid tumors: 13/13 (GD2⁺⁺ _numyogenin⁻ CD271^−/+ _nuMyoD1⁻ CD99⁻ EpCAM⁻) neuroblastoma samples, 5/5 (GD2⁻ _numyogenin⁺⁺ CD271⁺⁺ _nuMyoD1⁺⁺ CD99^−/+ EpCAM⁻) rhabdomyosarcomas, 2/2 (GD2^−/+ _numyogenin⁻ CD271⁺ _nuMyoD1⁻ CD99⁺ EpCAM⁻) Ewing sarcoma family of tumors, and 7/7 (GD2⁻ _numyogenin⁻ CD271⁺ _nuMyoD1⁻ CD99⁻ EpCAM⁺) Wilms tumors. In summary, here we designed and validated a new standardized antibody combination and MFC assay for diagnostic screening of pediatric solid tumors that might contribute to fast and accurate diagnostic orientation and classification of pediatric cancer in routine clinical practice. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

19 pages, 1571 KiB

Open AccessArticle

Anti-TRBC1 Antibody-Based Flow Cytometric Detection of T-Cell Clonality: Standardization of Sample Preparation and Diagnostic Implementation

by Noemí Muñoz-García, Margarida Lima, Neus Villamor, F. Javier Morán-Plata, Susana Barrena, Sheila Mateos, Carolina Caldas, Ana Balanzategui, Miguel Alcoceba, Alejandro Domínguez, Fabio Gómez, Anton W. Langerak, Jacques J. M. van Dongen, Alberto Orfao and Julia Almeida

Cancers 2021, 13(17), 4379; https://doi.org/10.3390/cancers13174379 - 30 Aug 2021

Cited by 16 | Viewed by 4088

Abstract

A single antibody (anti-TRBC1; JOVI-1 antibody clone) against one of the two mutually exclusive T-cell receptor β-chain constant domains was identified as a potentially useful flow-cytometry (FCM) marker to assess Tαβ-cell clonality. We optimized the TRBC1-FCM approach for detecting clonal Tαβ-cells and validated [...] Read more.

A single antibody (anti-TRBC1; JOVI-1 antibody clone) against one of the two mutually exclusive T-cell receptor β-chain constant domains was identified as a potentially useful flow-cytometry (FCM) marker to assess Tαβ-cell clonality. We optimized the TRBC1-FCM approach for detecting clonal Tαβ-cells and validated the method in 211 normal, reactive and pathological samples. TRBC1 labeling significantly improved in the presence of CD3. Purified TRBC1⁺ and TRBC1⁻ monoclonal and polyclonal Tαβ-cells rearranged TRBJ1 in 44/47 (94%) and TRBJ1+TRBJ2 in 48 of 48 (100%) populations, respectively, which confirmed the high specificity of this assay. Additionally, TRBC1⁺/TRBC1⁻ ratios within different Tαβ-cell subsets are provided as reference for polyclonal cells, among which a bimodal pattern of TRBC1-expression profile was found for all TCRVβ families, whereas highly-variable TRBC1⁺/TRBC1⁻ ratios were observed in more mature vs. naïve Tαβ-cell subsets (vs. total T-cells). In 112/117 (96%) samples containing clonal Tαβ-cells in which the approach was validated, monotypic expression of TRBC1 was confirmed. Dilutional experiments showed a level of detection for detecting clonal Tαβ-cells of ≤10⁻⁴ in seven out of eight pathological samples. These results support implementation of the optimized TRBC1-FCM approach as a fast, specific and accurate method for assessing T-cell clonality in diagnostic-FCM panels, and for minimal (residual) disease detection in mature Tαβ⁺ leukemia/lymphoma patients. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

18 pages, 2353 KiB

Open AccessArticle

B-Cell Regeneration Profile and Minimal Residual Disease Status in Bone Marrow of Treated Multiple Myeloma Patients

by Robéria Mendonça de Pontes, Juan Flores-Montero, Luzalba Sanoja-Flores, Noemi Puig, Roberto J. Pessoa de Magalhães, Alba Corral-Mateos, Anna Beatriz Salgado, Omar García-Sánchez, José Pérez-Morán, Maria-Victoria Mateos, Leire Burgos, Bruno Paiva, Jeroen te Marvelde, Vincent H. J. van der Velden, Carlos Aguilar, Abelardo Bárez, Aranzazú García-Mateo, Jorge Labrador, Pilar Leoz, Carmen Aguilera-Sanz, Brian Durie, Jacques J. M. van Dongen, Angelo Maiolino, Elaine Sobral da Costa and Alberto Orfao Show full author list Hide full author list

Cancers 2021, 13(7), 1704; https://doi.org/10.3390/cancers13071704 - 03 Apr 2021

Cited by 6 | Viewed by 2519

Abstract

B-cell regeneration during therapy has been considered as a strong prognostic factor in multiple myeloma (MM). However, the effects of therapy and hemodilution in bone marrow (BM) B-cell recovery have not been systematically evaluated during follow-up. MM (n = 177) and adult (≥50y) [...] Read more.

B-cell regeneration during therapy has been considered as a strong prognostic factor in multiple myeloma (MM). However, the effects of therapy and hemodilution in bone marrow (BM) B-cell recovery have not been systematically evaluated during follow-up. MM (n = 177) and adult (≥50y) healthy donor (HD; n = 14) BM samples were studied by next-generation flow (NGF) to simultaneously assess measurable residual disease (MRD) and residual normal B-cell populations. BM hemodilution was detected in 41 out of 177 (23%) patient samples, leading to lower total B-cell, B-cell precursor (BCP) and normal plasma cell (nPC) counts. Among MM BM, decreased percentages (vs. HD) of BCP, transitional/naïve B-cell (TBC/NBC) and nPC populations were observed at diagnosis. BM BCP increased after induction therapy, whereas TBC/NBC counts remained abnormally low. At day+100 postautologous stem cell transplantation, a greater increase in BCP with recovered TBC/NBC cell numbers but persistently low memory B-cell and nPC counts were found. At the end of therapy, complete response (CR) BM samples showed higher CD19⁻ nPC counts vs. non-CR specimens. MRD positivity was associated with higher BCP and nPC percentages. Hemodilution showed a negative impact on BM B-cell distribution. Different BM B-cell regeneration profiles are present in MM at diagnosis and after therapy with no significant association with patient outcome. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Figure 1

17 pages, 4841 KiB

Open AccessArticle

Monocyte Subsets and Serum Inflammatory and Bone-Associated Markers in Monoclonal Gammopathy of Undetermined Significance and Multiple Myeloma

by Daniela Damasceno, Julia Almeida, Cristina Teodosio, Luzalba Sanoja-Flores, Andrea Mayado, Alba Pérez-Pons, Noemi Puig, Paula Arana, Bruno Paiva, Fernando Solano, Alfonso Romero, Sergio Matarraz, Wouter B. L. van den Bossche, Juan Flores-Montero, Brian Durie, Jacques J. M. van Dongen and Alberto Orfao

Cancers 2021, 13(6), 1454; https://doi.org/10.3390/cancers13061454 - 22 Mar 2021

Cited by 10 | Viewed by 3299

Abstract

Background. Monocyte/macrophages have been shown to be altered in monoclonal gammopathy of undetermined significance (MGUS), smoldering (SMM) and active multiple myeloma (MM), with an impact on the disruption of the homeostasis of the normal bone marrow (BM) microenvironment. Methods: We investigated the distribution [...] Read more.

Background. Monocyte/macrophages have been shown to be altered in monoclonal gammopathy of undetermined significance (MGUS), smoldering (SMM) and active multiple myeloma (MM), with an impact on the disruption of the homeostasis of the normal bone marrow (BM) microenvironment. Methods: We investigated the distribution of different subsets of monocytes (Mo) in blood and BM of newly-diagnosed untreated MGUS (n = 23), SMM (n = 14) and MM (n = 99) patients vs. healthy donors (HD; n = 107), in parallel to a large panel of cytokines and bone-associated serum biomarkers. Results: Our results showed normal production of monocyte precursors and classical Mo (cMo) in MGUS, while decreased in SMM and MM (p ≤ 0.02), in association with lower blood counts of recently-produced CD62L⁺ cMo in SMM (p = 0.004) and of all subsets of (CD62L⁺, CD62L⁻ and FcεRI⁺) cMo in MM (p ≤ 0.02). In contrast, intermediate and end-stage non-classical Mo were increased in BM of MGUS (p ≤ 0.03), SMM (p ≤ 0.03) and MM (p ≤ 0.002), while normal (MGUS and SMM) or decreased (MM; p = 0.01) in blood. In parallel, increased serum levels of interleukin (IL)1β were observed in MGUS (p = 0.007) and SMM (p = 0.01), higher concentrations of serum IL8 were found in SMM (p = 0.01) and MM (p = 0.002), and higher serum IL6 (p = 0.002), RANKL (p = 0.01) and bone alkaline phosphatase (BALP) levels (p = 0.01) with decreased counts of FcεRI⁺ cMo, were restricted to MM presenting with osteolytic lesions. This translated into three distinct immune/bone profiles: (1) normal (typical of HD and most MGUS cases); (2) senescent-like (increased IL1β and/or IL8, found in a minority of MGUS, most SMM and few MM cases with no bone lesions); and (3) pro-inflammatory-high serum IL6, RANKL and BALP with significantly (p = 0.01) decreased blood counts of immunomodulatory FcεRI⁺ cMo-, typical of MM presenting with bone lesions. Conclusions: These results provide new insight into the pathogenesis of plasma cell neoplasms and the potential role of FcεRI⁺ cMo in normal bone homeostasis. Full article

(This article belongs to the Special Issue Leukemia and Lymphoma Immunophenotyping)

► Show Figures

Graphical abstract

Journal Menu

Journal Browser

Leukemia and Lymphoma Immunophenotyping

Share This Special Issue

Special Issue Editors

Special Issue Information

Keywords

Published Papers (10 papers)

Research

Further Information

Guidelines

MDPI Initiatives

Follow MDPI