New Immunophenotypic Profile of Waldenstrom’s Macroglobulinemia: Interest of CD80 and CD86 Staining

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5277-5277
Author(s):  
Andrea Toma ◽  
Magali Le Garff-Tavernier ◽  
Martine Brissard ◽  
Patrick Bonnemye ◽  
Lucille Musset ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
Cell Population ◽  
Light Chain ◽  
Waldenström’S Macroglobulinemia ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia ◽  
Immunophenotypic Analysis

Abstract The immunophenotypic characterization is an essential tool in the diagnosis of hematological malignancies but the immunophenotypic features in Waldenstrom’s macroglobulinemia (WM) remain not clearly defined. We studied 96 cases of WM diagnosed by monoclonal IgM in the serum and morphological lymphoplasmacytic bone marrow infiltration, and we compared results to 33 cases of other chronic B-cell lymphoproliferative disorders (LPD), including marginal zone (MZL)(n=23), mantle cell (MCL)(n=8) and follicular (FL)(n=2) lymphomas. Patients with a Matutes score >3 (chronic lymphocytic leukemia) and with pathognomonic immunophenotype (hairy cell leukemia) were excluded. Immunophenotypic analysis was performed by flow cytometry using six-colour staining (FACS Canto II, Becton Dickinson). In WM and LPD groups, a monoclonal B-cell population was identified in blood (31 and 28 patients, respectively), blood and bone marrow (28 and 4 patients) or bone marrow samples (23 and 1 patients). Overall, 61% of WM patients showed a monoclonal B-cell population in blood. Neoplastic cells of WM and LPD patients with blood and/or bone marrow involvement expressed a monoclonal immunoglobulin light chain kappa (in 70% and 73% of cases respectively) or lambda (30% and 27%). The intensity of expression of the light chain was heterogeneous in both groups (high, normal or low expression in 43%, 27% or 30% of WM, and in 52%, 33% or 15% of LPD, respectively). All pan-B antigens (CD20, CD19, CD79b) were positive for at least 97% of patients. Results obtained with other antigens in WM compared to LPD were: CD10 = 10% vs 7% of patients, CD23 = 33% vs 56%, CD5 = 14% vs 26%, FMC7 = 76% vs 89%, CD38 = 56% vs 41%, CD25 = 86% vs 84%, CD43 = 12% vs 16%, and CD11c = 10% vs 36%. The intensity of expression of these antigens was heterogeneous in both groups. Among the antigens only tested in the WM group, CD1c and CD27 were positive for 70% of patients, IgM and IgD for 95% of patients, and CD103 as well as CD117 were negative in all cases. No difference was found between blood and bone marrow for all previous antigens. Plasma cells (CD38/CD138 positive cells) were found at low levels (less than 2.5% of B-cells) for 46% of WM in blood and/or bone marrow samples. Among the 10 WM patients tested for ZAP-70 expression, 9 were negative and 1 showed a low intensity expression. These results confirm that the immunophenotypic analysis usually performed with standard antigens does not allow defining a typical profile of WM. In order to tentatively identify the WM among the B-cell malignancies, we studied the expression of molecules known to be involved in B-cell development or in costimulatory pathways of antigenic activation, namely CD69, CD83, CD80 and CD86. We first analyzed blood samples of 24 WM patients showing a peripheral monoclonal B-cell population. CD80 was positive (> 20% of B-cells) in all cases and CD83, CD69 and CD86 were always negative. Among these WM patients, 13 were also studied for the bone marrow phenotype. No difference was found between blood and bone marrow phenotype in 11/13 WM cases. We then studied 11 LPD with blood tumoral involvement (MZL(n=7), MCL(n=2) and FL(n=2)). In these LPD, CD69 and CD83 were always negative and, in most cases (9/11 patients), CD80 and CD86 were also negative. Interestingly, CD80 was found positive in 2 patients with MZL, but the CD80 positivity was always associated to the CD86 positivity. Altogether, these data suggest that the inclusion of CD80 and CD86 in the panel of cytometric analysis allow to discriminate WM from other B-LPD with peripheral blood involvement.

Immunophenotypic Analysis of Waldenstrom’s Macroglobulinemia: A Study of 63 Cases.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4399-4399
Author(s):  
Andrea Toma ◽  
Magali Le Garff-Tavernier ◽  
Martine Brissard ◽  
Patrick Bonnemye ◽  
Lucile Musset ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
Light Chain ◽  
Plasma Cells ◽  
Lymphoproliferative Disorders ◽  
Waldenström’S Macroglobulinemia ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia ◽  
Immunophenotypic Analysis

Abstract The immunophenotypic characterization of Waldenstrom’s macroglobulinemia (WM) is still unclear, despite being an essential tool in the diagnosis of hematological malignancies. We retrospectively reviewed the immunophenotypic profile of 63 cases of WM showing monoclonal IgM in the serum and morphological lymphoplasmacytic bone marrow infiltration, and of 26 cases of other chronic B-cell lymphoproliferative disorders having monoclonal IgM in the serum (LPD-M), including marginal zone (n=16), mantle cell (n=8) and follicular (n=2) lymphomas. The median age at diagnosis was 64[46–92] years. Immunophenotypic analysis was performed by flow cytometry between January 1998 and July 2007, using four or six monoclonal antibody combinations. In WM and LPD-M groups, all patients showed a monoclonal tumoral B-cell population, detected and studied in blood (21 and 23 patients, respectively), blood and bone marrow (19 and 2 patients, respectively) or bone marrow samples (23 and 1 patient, respectively). Patients with a Matutes score > 3 were excluded. Neoplastic cells, in all cases, expressed a monoclonal immunoglobulin light chain (kappa for 70% WM and 73% LPD-M, lambda for 30% WM and 27% LPD-M). The intensity of expression of the monoclonal light chain was particularly heterogeneous in both groups: high, normal or low expression in 43%, 27% or 30% of WM, and in 52%, 33% or 15% of LPD-M, respectively. Pan-B antigens (CD20, CD19, CD79b) were positive for at least 97% of patients. The results obtained with other antigens in WM compared to LPD-M were: CD10 = 10 versus 7%, CD23 = 33 versus 56%, CD5 = 14 versus 26%, FMC7 = 76 versus 89%, CD38 = 56 versus 41%, CD25 = 86 versus 84%, CD43 = 12 versus 16%, and CD11c = 10 versus 36%. The intensity of expression of these antigens was heterogeneous in both groups. Among the antigens only tested in the WM group, CD1c and CD27 were positive for 70% of patients, IgM and IgD for 95% of patients, and CD103 as well as CD117 were negative in all cases. Considering all (blood and/or bone marrow) WM samples, plasma cells (CD38/CD138 positive cells) were found at low levels (less than 2.5% of B-cells) for 46% of WM. Comparing blood versus bone marrow WM samples, no differences were found for all previous antigens except the higher frequency of low counts of plasma cells in bone marrow (observed in 71% of WM) versus blood samples (28% of WM). Among the 10 WM patients tested for ZAP-70 expression, nine were negative and one showed a low intensity expression. In conclusion, our results show that the immunophenotypic analysis usually performed with standard antigens in WM overlaps with other B-cell lymphoproliferative disorders. Studies involving the expression of new antigens and/or other biological approaches are required to identify the WM among the B-cell malignancies and are ongoing in our group.

Immunophenotypic Analysis Using CD80 and CD86 Staining Improves the Diagnosis of Waldenstrom's Macroglobulinemia by Evidencing a CD80+/CD86- Profile.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4389-4389
Author(s):  
Andrea Toma ◽  
Stephanie Poulain ◽  
Aurore Grelier ◽  
Magali Le Garff-Tavernier ◽  
Frederic Davi ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Population ◽  
Peripheral Blood ◽  
Waldenström’S Macroglobulinemia ◽  
The Mean ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia ◽  
Immunophenotypic Analysis

Abstract Abstract 4389 The immunophenotypic characterization is an essential tool in the diagnosis of hematological malignancies but the immunophenotypic profile of Waldenstrom's macroglobulinemia (WM) B cells remains not clearly defined. In order to improve the diagnosis of WM by an immunophenotypic peripheral blood test, we studied within a multicentric protocol the expression of costimulatory molecules CD80 (B7.1) and CD86 (B7.2) on blood B-cells of WM patients diagnosed by monoclonal IgM in the serum and morphological lymphoplasmacytic bone marrow (BM) infiltration. Results were compared to those obtained in healthy controls and other chronic B-cell lymphoproliferative disorders (LPD). Immunophenotypic analysis was performed by flow cytometry using six-colour staining. The CD80 and CD86 staining was concomitant with usual analysis of standard antigens (kappa/lambda immunoglobulin light chains, CD20, CD19, CD79b, CD5, CD10, CD23, FMC7, CD38, CD25, CD43, CD11c, CD1c, CD27, IgM, IgD). We first analyzed 65 cases of WM and compared results to 13 healthy control subjects. Among the WM patients, 45 (69%) showed a monotypic kappa or lambda peripheral blood B-cell population with a Matutes score '3. In these patients, CD80 was always positive (>21% of B-cells) and CD86 negative (<18%). The mean responses obtained in WM compared to controls were: CD80 = 44.2±19.5% versus 13±4.6% (p<0.00001), CD86 = 5.4±5.3% versus 4.5±2.1% (p=0.18). Among the 20 patients without monotypic kappa or lambda B-cell population in peripheral blood, 13 showed a CD80+/CD86- profile (CD80 = 33±7.4%, CD86 = 7.6±4.4%). Six of them were studied by molecular aproach (PCR) showing the presence of a clonal B-cell population in all cases. Overall, 90% of WM patients showed a CD80+/CD86- profile in blood. Seven patients without peripheral monotypic kappa or lambda B-cell population evidenced a CD80-/CD86- profile; among them, one was tested by PCR and showed the lack of a clonal B-cell population in blood. We then studied other chronic B-cell LPD with peripheral blood neoplastic involvement by a monotypic kappa/lambda population, including 15 marginal zone lymphoma (MZL), 4 non-Hodgkin lymphomas (NHL) and 28 chronic lymphocytic leukemia (CLL). The mean expression of CD80 was 37±35.6%, 2.5±1.7%, 12.4±20.6%, and of CD86 30.5±38%, 19±27.5%, 15.4±19.3%, respectively. The expression of these antigens in MZL was heterogeneous, but a WM-type CD80+/CD86- profile was never observed in peripheral blood B cells. No NHL and 4 CLL (14%) exhibited a CD80+/CD86- profile. In conclusion, while immunophenotypic analysis usually performed with standard antigens does not allow to define a typical profile of WM, additional CD80 and CD86 staining in blood samples usefully improves the diagnosis of WM in the context of chronic B-cell LPD with peripheral blood involvement having a Matutes score '3. Furthermore, even in the absence of blood involvement detected by classical imunophenotypical aproach, the CD80 and CD86 staining may prove the presence of a peripheral blood monoclonal population, discriminating WM from other B-LPD with plasmatic IgM and lymphoplasmacytic BM infiltration, and diagnosing a WM in 90% of cases. Disclosures: No relevant conflicts of interest to declare.

B-Lymphocyte Stimulator (BLyS) Is Highly Expressed in Waldenstrom’s Macroglobulinemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2291-2291
Author(s):  
Stephen M. Ansell ◽  
Deanna M. Grote ◽  
Steven C. Ziesmer ◽  
Thomas E. Witzig ◽  
Robert A. Kyle ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cells ◽  
B Cell ◽  
B Lymphocyte ◽  
Waldenström’S Macroglobulinemia ◽  
B Lymphocyte Stimulator ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia ◽  
Normal Controls

Abstract Waldenstrom’s macroglobulinemia is a serious and frequently fatal illness, however many of the mechanisms leading to this disease are not yet known. It is clear, however, that there is dysregulation of the balance between cell proliferation and programmed cell death. BLyS (B-lymphocyte stimulator) is a newly identified TNF family member expressed by monocytes, macrophages, and dendritic cells. BLyS has been shown to be critical for maintenance of normal B cell development and homeostasis, and has been found to stimulate lymphocyte growth. BLyS is overexpressed in a variety of B-cell malignancies and has been shown to inhibit apoptosis in malignant B-cells. Studies of the effects of BLyS on B cell physiology have shown that it also regulates immunoglobulin secretion. To determine the relevance of the BLyS receptor-ligand system in Waldenstrom’s macroglobulinemia, we examined malignant B cells from 5 patients with Waldenstrom’s macroglobulinemia for their ability to bind soluble BLyS and for the expression of the known BLyS receptors, TACI, BAFF-R, or BCMA. The malignant B cells were found to bind BLyS and express BAFF-R and TACI. BCMA expression was undetectable. We then determined the expression of BLyS in bone marrow specimens from 5 patients with Waldenstrom’s macroglobulinemia by immunohistochemistry and compared it to the expression in 5 normal bone marrow specimens. The lymphoplasmacytic cell infiltrate in the bone marrow of patients with Waldenstrom’s macroglobulinemia showed significantly increased BLyS expression. We further determined the serum BLyS levels by ELISA in stored serum specimens from patients with Waldenstrom’s macroglobulinemia (n=20), and compared them to serum BLyS levels in other patients with lymphoplasmacytic lymphoma without elevated immunoglobulin levels (n=10) and to serum levels in normal controls (n=50). Serum BLyS levels in Waldenstrom’s patients (mean: 49.6ng/ml) as well as those in patients with lymphoplasmacytic lymphoma (mean; 46.7ng/ml) were significantly higher than normal controls (mean 12.6ng/ml). In conclusion, we have demonstrated that malignant B cells from patients with Waldenstrom’s macroglobulinemia express the receptors for BLyS and can bind soluble BLyS. Furthermore, we have found that serum BLyS levels are significantly elevated in patients with Waldenstrom’s macroglobulinemia when compared to controls. Strategies to inhibit BLyS may potentially have significant therapeutic efficacy in Waldenstrom’s macroglobulinemia.

Role of B-Lymphocyte Stimulator (BLyS) in Waldenstrom’s Macroglobulinemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 601-601
Author(s):  
Sherine F. Elsawa ◽  
Anne J. Novak ◽  
Deanna M. Grote ◽  
Steven C. Zeismer ◽  
Thomas E. Witzig ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
B Lymphocyte ◽  
Waldenström’S Macroglobulinemia ◽  
Immunoglobulin Production ◽  
B Lymphocyte Stimulator ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia

Abstract Waldenstrom’s macroglobulinemia (WM) is a serious and frequently fatal disorder characterized by the production of a monoclonal IgM protein, a lymphoplasmacytic infiltrate in the bone marrow, and associated symptoms including anemia, lymphadenopathy and hyperviscosity. Many of the mechanisms leading to this disease are not yet known. It is clear, however, that there is dysregulation of the balance between cell proliferation and programmed cell death. BLyS (B-lymphocyte stimulator) is a TNF family member expressed by monocytes, neutrophils, macrophages, and dendritic cells. BLyS has been shown to be critical for maintenance of normal B cell development and homeostasis, and has been found to stimulate lymphocyte growth. BLyS is overexpressed in a variety of B-cell malignancies and has been shown to inhibit apoptosis in malignant B-cells. Studies of the effects of BLyS on B cell physiology have shown that it also regulates immunoglobulin secretion. In previous work, we have shown that malignant B cells from patients with WM are able to bind soluble BLyS and variably express the BLyS receptors, BAFF-R, TACI and BCMA. We also found expression of BLyS in bone marrow specimens by immunohistochemistry and elevated serum BLyS levels in patients with WM. The goal of this study was to determine the functional role of BLyS-receptor ligand system in Waldenstrom’s macroglobulinemia and its relevance to the increased immunoglobulin production seen in this disease. Using cells from WM patients, we first examined the ability of BLyS to increase the secretion of IgM by malignant B cells. BLyS, alone or in combination with cytokines that induce plasmacytic differentiation and immunoglobulin production (IL-2, IL-6, IL-10 and IL-12), was found to increase IgM secretion by malignant B cells. Mean baseline IgM levels significantly increased in cells treated with BLyS (p=0.03), cytokines (p=0.0002) and a combination of BLyS and cytokines (p<0.0001). We then determined the effect of BLyS on the survival of malignant B cells using Annexin-V/PI staining. Compared to cells cultured in media alone, BLyS was found to increase viability of malignant B cells from WM patients. Cell viability was normalized relative to the media-alone control and the median relative viability increased significantly compared to controls (median increase 41.2%; range 8 – 46%). Next, we examined the ability of BLyS to modulate cell proliferation using thymidine incorporation. Using WM patient samples, BLyS was found to significantly enhance the proliferation of malignant B cells (p=0.0002). Furthermore, addition of anti-Ig antibody further enhanced the ability of BLyS to promote the proliferation of malignant B cells (p<0.0001). In summary, we have demonstrated that BLyS enhances IgM secretion by malignant B cells from patients with Waldenstrom’s macroglobulinemia. We have also demonstrated the ability of BLyS to enhance the survival and proliferation of malignant B cells. Strategies to inhibit BLyS may potentially have therapeutic efficacy in Waldenstrom’s macroglobulinemia.

Molecular Characterization of Clonotypic VDJ Sequences in Waldenstrom’s Macroglobulinemia Suggests Independent Transformation Events in the Development of Biclonal B Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4624-4624
Author(s):  
Jitra Kriangkum ◽  
Brian J. Taylor ◽  
Steven P. Treon ◽  
Michael J. Mant ◽  
Tony Reiman ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Waldenström’S Macroglobulinemia ◽  
Fragment Analysis ◽  
Genealogical Tree ◽  
Cell Clones ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
B Lineage ◽  
Waldenström's Macroglobulinemia

Abstract Malignant B lineage cells in Waldenstrom’s macroglobulinemia (WM) express a unique clonotypic VDJ associated with IgM. Studies of WM patients revealed a frequent incidence of biclonal B cells (16%) as defined by the presence of two distinct clonotypic VDJ sequences. This is the first report to estimate the frequency of biclonality in WM. Four WM cases are reported: two with distinct IgM clones (WM1-19 and WM1-09), one with distinct IgM/IgA clones (WM1-18), and one with related but diversified IgM/IgG clones (WM10). In 2 cases (WM1-19 and WM10), the two clonotypic signatures were found to be abundant in bone marrow (BM) but less frequent in blood, reminiscent of monoclonal WM cases. In WM1-19, single cell analysis showed that only one partner VDJ was expressed per cell, excluding the possibility of aberrant biallelic rearrangements. The distribution ratio between the two tumor clones was 2:1, suggestive of their mutual role in the clinical manifestation. In the other 2 cases (WM1-09 and WM1-18), partner clones were shown by CDR3 fragment analysis to be anatomically distinct, with one BM clonotypic signature and one in blood. In these 2 patients, the BM clone was hypermutated (6.2% and 3.8%) while the blood clone was germline or minimally mutated (0% and 1.0%). Partner clones lacked intraclonal diversity and Ig class switching, characteristic of malignant WM clones, suggesting relatively frequent transformation events throughout B lineage differentiation. The biological events leading to the appearance of clones in two different anatomic sites and the clinical implications remain to be understood. CDR3 fragment analysis in the longitudinal studies of WM1-09 and WM1-18 suggested that the repertoire of blood B cells may recover some level of diversity after successive cycles of treatment (WM1-18), while the persistence of the monoclonal peak (WM1-09) likely reflects a tumor that does not respond to treatment. In WM10, biclonal IgM/IgG B cell clones shared common VDJ gene rearrangement. The IgG clone displayed a higher mutation rate than did the IgM clone (9.7% vs. 6.1%). Increased somatic hypermutation in IgG consists mostly of replacement mutations that are clustered within the CDR regions, strongly supporting a contention that the IgG clone has undergone affinity maturation. There are 11 point mutations in the IgM that are not present in the IgG, suggesting that the two clones are distinctly different and belong to different branches of the genealogical tree. These distinct mutational signatures suggest that biclonal IgM/IgG clones are unlikely derived from clonal evolution. Overall, our results suggest that for the four biclonal WM evaluated here, the partner B cell clones appear to have undergone separate transformation events in the development of biclonality. The extent to which each partner clone contributes to disease progression and death, whether separately or in synergy, is as yet unknown.

IL-21 in the Bone Marrow Microenvironment Contributes to IgM Secretion and Proliferation of Malignant Cells in Waldenstrom's Macroglobulinemia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 770-770
Author(s):  
Lucy S. Hodge ◽  
Steve Ziesmer ◽  
Thomas E. Witzig ◽  
Anne J. Novak ◽  
Stephen M. Ansell
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
Cell Line ◽  
Cellular Proliferation ◽  
Bone Marrow Microenvironment ◽  
Waldenström’S Macroglobulinemia ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia ◽  
Dependent Mechanism

Abstract Abstract 770 Background: Waldenstrom's macroglobulinemia (WM) is a B-cell lymphoma characterized by high serum monoclonal IgM and infiltration of lymphoplasmacytic cells into the bone marrow. As with many hematologic malignancies, cytokines within the tumor microenvironment play an important role in supporting the growth and survival of malignant WM cells. IL-21 is a pleiotropic cytokine involved in the differentiation of B cells into plasma cells. During malignancy, IL-21 has demonstrated diverse effects promoting the growth of myeloma and Hodgkin lymphoma cells while inducing apoptosis in chronic lymphocytic leukemia. However, the biologic significance of IL-21 has not been examined in WM. Our objective here was to assess the expression of IL-21 and its receptor in WM cells and to examine whether IL-21 contributes to the biology of WM. Results: When compared to normal bone marrows, immunohistochemistry revealed significant IL-21 staining in the bone marrow of patients with WM (n=5). To determine whether WM cells are susceptible to IL-21 in the microenvironment, expression of the IL-21 receptor (IL-21R) was assessed via PCR in CD19+CD138+ cells isolated by positive selection from patients with WM (n=8) and in the newly characterized WM cell line, MWCL-1. Nearly all (7/8) CD19+CD138+ WM cells expressed IL-21R, as did MWCL-1 cells. Using flow cytometry we detected expression of IL-21R protein on the surface of WM cells as well. The contribution of microenvironmental IL-21 to the biology of WM tumors was then examined. In MWCL-1 cells, IL-21 (100 ng/mL) increased proliferation by 37% (p=0.005) over untreated controls as determined by thymidine incorporation at 72 hr, and in primary WM cells, proliferation increased by nearly 50% (p=0.003). Interestingly, the immortalized B cell line, IM-9, responded to IL-21 with a significant decrease in proliferation, consistent with previous data indicating differential effects of IL-21 depending on the pathological status of the B-cell in question. IL-21 also significantly induced (p<0.0005) IgM secretion in WM as measured by ELISA (MWCL-1 5,956 +/− 393 ng/mL vs. 10,013 +/− 730 ng/mL; CD19+138+ WM 504 +/− 33 ng/mL vs. 811 +/− 32.5 ng/mL). Annexin/PI staining was used to assess viability, but no apoptotic effects were associated with IL-21 in WM. To better understand the mechanisms through which IL-21 increases cellular proliferation and IgM secretion in WM, we characterized STAT activation in response to this cytokine. In MWCL-1 cells, IL-21 significantly increased the phosphorylation of both STAT1 and STAT3, and to a lesser extent, STAT5. Treatment with a STAT3 inhibitor completely abolished the effects of IL-21 on cellular proliferation and IgM secretion suggesting IL-21 mediates its biologic activity through a STAT3-dependent mechanism. The expression of transcription factors involved in B-cell differentiation was also measured in MWCL-1 cells treated with IL-21. Both BLIMP-1 and Bcl-6 levels significantly increased upon addition of IL-21, whereas PAX5 was significantly decreased. IL-21 had no effect on the expression of XBP-1, which is involved in regulating Ig secretion, suggesting that the increase in IgM secretion in MWCL-1 cells may occur secondary to the increase in proliferation, as opposed to an actual increase in the production of IgM. Lastly, IL-21 significantly enhanced IL-10 secretion from MWCL-1 cells (669 +/− 152 pg/mL vs. 1,948 +/− 279 pg/mL, p=0.0002). While the interplay between IL-10 and IL-21 in WM remains to be examined, IL-10 is known to be involved in normal B-cell development and may have synergistic effects with IL-21 in malignant WM cells. Overall our data indicate that IL-21 in the bone marrow microenvironment significantly affects the biology of WM tumor cells through a STAT3-dependent mechanism. Disclosures: No relevant conflicts of interest to declare.

Trisomy 4, a New Chromosomal Abnormality in Waldenstrom’s Macroglobulinemia.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2841-2841
Author(s):  
C. Terre ◽  
F. N. Guyen-Khac ◽  
C. Barin ◽  
M.J. Moziconacci ◽  
V. Eclache ◽  
...  
Keyword(s):  
Bone Marrow ◽  
B Cell ◽  
Chromosomal Abnormality ◽  
Partial Trisomy ◽  
Waldenström’S Macroglobulinemia ◽  
Genetic Event ◽  
The Mean ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
Waldenström's Macroglobulinemia

Abstract Background: Waldenstrom’s Macroglobulinemia(WM) is a rare B-cell disorder characterized by the accumulation of monoclonal lymphoplasmocytic cells and a serum monoclonal IgM protein. The genetic basis of this disorder is poorly understood. Molecular cytogenetic abnormalities differ from those reported for multiple myeloma, lymphomas, and B-cell chronic lymphocytic leukemia. One recurrent abnormality, deletion of the long arm of chromosome 6, was reported with a high prevalence (23–50%), but seems associated with advanced disease and clonal evolutions. Aims : As data on molecular changes in WM are limited, we used interphase fluorescence in situ hydidization (IP-FISH) to evaluate the prevalence of trisomy 4 in WM patients. Material and methods: 39 patients with a diagnosis of WM , according to the Workshop classification, were included: 28 men and 11 women. Mean age at diagnosis was 66 years (48–84 years). Mean percentage of bone marrow lymphoplasmocytic cells was 47% (13–97%). Conventional cytogenetic was performed after 72 or 96 hours culture with TPA, at diagnosis for 23 patients and 3.8 years median (6 months–9 years) after for 13. FISH study was carried on cytogenetic cryopreserved pellets using a CEP-4 probe (Qbiogene, Illkirsch, France) Results: Karyotypes were available for 37cases (2 failures): 24 were normal, 3 had a trisomy 4, (2 as the sole abnormality), 2 a partial loss of 17p (one deletion and one additional material), 2 a loss of sexual chromosome, 1 a t(11;18), 1 a partial duplication of 17q, 1 a partial monosomy 5q with insertion of unknown material in 4q, 1 a complex karyotype with a partial trisomy 4 and 2 with abnormal independent clones. IP-FISH: The 3 complete trisomies 4, already seen by conventional cytogenetic, were confirmed and 4 others were detected. The mean percentage of cells with trisomy 4 was 31% (11–52%) (cut-off 2%, median+4SD) Discussion: We found that 7 of 39 patients (18%) with WM harboured a complete trisomy 4 with IP-FISH, contrary to only three (8%) with conventional cytogenetic. This study, performed in interphase, is independent of the proliferation rate. Therefore the prevalence is probably underestimated since IP-FISH was realized in unselected cells: in 11 patients the percentage of clonal involvement of the bone marrow was under 30% and it was unknown in 6 patients. The mean percentage of bone marrow lymphoplasmocytic cells was higher for patients with trisomy 4 (63%) than in others (43%). The trisomy 4 was the only abnormality in 2 of the 3 karyotypes where it was identified and in one sporadic case reported previously. Thus it could be a specific primary genetic event in the WM. Moreover, the detection of one partial trisomy 4 define an interesting region with potential oncogene. cKIT, located at 4q12, was a good candidat gene and molecular studies are in process. Conclusion: We identified a new recurrent chromosomal abnormality in WM, the trisomy 4, with a prevalence of at least 18%. The exact prevalence of this abnormality should be determined by IP-FISH study in CD19+ and CD138+ selected cells. This abnormality could be primary and could lead to elusive the molecular pathogenesis of WM.

Identification of two DNA methylation subtypes of Waldenström's macroglobulinemia with plasma and memory B cell features

Blood ◽  
2020 ◽  
Author(s):  
Damien Roos-Weil ◽  
Brian Giacopelli ◽  
Marine Armand ◽  
Véronique Della Valle ◽  
Hussein Ghamlouch ◽  
...  
Keyword(s):  
Dna Methylation ◽  
B Cell ◽  
Plasma Cells ◽  
Surface Expression ◽  
B Cell Differentiation ◽  
Waldenström’S Macroglobulinemia ◽  
Waldenstrom's Macroglobulinemia ◽  
Waldenstrom’S Macroglobulinemia ◽  
B Cell Subsets ◽  
Waldenström's Macroglobulinemia

Epigenetic changes during B cell differentiation generates distinct DNA methylation signatures specific for B cell subsets, including memory B cells (MBCs) and plasma cells (PCs). Waldenström's macroglobulinemia (WM) is a complex B cell malignancy uniquely comprised of a mixture of lymphocytic and plasmacytic phenotypes. Here we integrated genome-wide DNA methylation, transcriptome, mutation and other phenotypic features of tumor cells from 35 MYD88-mutated WM patients in relation to normal plasma and B cell subsets. We discovered that WM patients naturally segregate into two groups according to DNA methylation patterns, related to normal MBC and PC profiles, and reminiscent of other memory and plasma cell-derived malignancies. Concurrent analysis of DNA methylation changes in normal and WM development were used to capture tumor-specific events, highlighting a selective reprogramming of enhancer regions in MBC-like WM and repressed and heterochromatic regions in PC-like WM. MBC-like WM hypomethylation was enriched in motifs belonging to PU.1, TCF3 and OCT2 transcription factors and involved elevated MYD88/TLR pathway activity. PC-like WM displayed marked global hypomethylation and selective overexpression of histone genes. Finally, WM subtypes exhibited differential genetic, phenotypic and clinical features. MBC-like WM harbored significantly more clonal CXCR4 mutations (P=0.015), deletion 13q (P=0.006), splenomegaly (P=0.02) and thrombocytopenia (P=0.004), while PC-like WM harbored more deletion 6q (P=0.012), gain 6p (P=0.033), had increased frequencies of IGHV3 genes (P=0.002), CD38 surface expression (P=4.1e-5), and plasmacytic differentiation features (P=0.008). Together our findings illustrate a novel approach to subclassify WM patients using patterns of DNA methylation and reveal divergent molecular signatures among WM patients.

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