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Immunoglobulin M Concentration in Waldenström Macroglobulinemia: Correlation With Bone Marrow B Cells and Plasma Cells
IWWM 2012 Proceedings
Immunoglobulin M Concentration in Waldenström Macroglobulinemia: Correlation With Bone Marrow B Cells and Plasma Cells Ruth M. de Tute, Andy C. Rawstron, Roger G. Owen Clinical Lymphoma, Myeloma & Leukemia, Vol. 13, No. 2, 211-3 © 2013 Elsevier Inc. All rights reserved. Keywords: Immunoglobulin M, Plasma cell, Waldenström macroglobulinemia
Abstract
Patients and Methods
Serum immunoglobulin (Ig) M levels vary considerably among patients with Waldenström macroglobulinemia, and previous studies have failed to demonstrate a correlation with overall bone marrow disease burden. In this study, bone marrow B cells and plasma cells were enumerated by flow cytometry and correlated with serum IgM concentrations. Monotypic B cells comprised a median of 6% of bone marrow leukocytes but did not correlate with IgM levels (r ⫽ 0.071, P ⫽ .5). Plasma cells, although typically present in lower numbers (median, 0.52%) did show a correlation with IgM (r ⫽ 0.452, P ⫽ .01). IgM levels in Waldenström macroglobulinemia, at least in part, correlate with the degree of plasma cell differentiation seen within the tumor.
Eighty-seven patients (49 men, 38 women) fulfilling consensus panel and World Health Organization diagnostic criteria1,2 for WM were included in this analysis. The median age was 75 years (range, 37-91 years) and the median IgM (M component concentration determined by densitometry) was 11 g/L (range, 1-51 g/L). All the patients were assessed at initial presentation and were untreated. Multiparameter flow cytometry was performed in a single supraregional hematopathology laboratory (HMDS Laboratory, Leeds, UK), and BM samples were received from 25 individual clinical hematology centers. IgM concentrations were determined in local laboratories. Leukocytes were prepared from fresh BM aspirate samples by incubation in a 10-fold excess of ammonium chloride (8.6 g/L in distilled water; Vickers Laboratories, Pudsey, UK) for 5 minutes at 37°C. Cells were then washed twice in buffer (FACSFlow, which contained 0.3% bovine serum albumin; Sigma-Aldrich, Dorset, UK). Leukocytes (approximately 1 ⫻ 106) were stained with 30 L of a preprepared cocktail of antibodies. For plasma cell enumeration, this comprised the following: CD27 fluorescein isothiocyanate (FITC), CD56 phycoerythrin (PE), CD19 peridin chlorophyll protein-cyanine 5.5 (PerCP-Cy5.5), CD38 phycoerythrincyanine (PE-Cy7), CD45 allophycocyanin-cyanine (APC-H7) (all from Becton Dickinson, Oxford, UK), and CD138 allophycocyanin (APC) (Miltenyi Biotec, Bisley, UK). The panel used for B-cell enumeration comprised Lambda FITC, Kappa PE, CD19 PerCP-Cy5.5, CD5 PE-Cy7, CD20 APC, and CD45 APC-H7 (all from Becton Dickinson). After incubation in the dark at 4°C for 20 minutes, the cells were washed twice in buffer and resuspended in FACSFlow. Samples were acquired on a FACSCanto II analyzer by using FACSDiva software (BD Biosciences, Oxford, UK), and a minimum of 100,000 events were acquired for each screening tube. The Pearson correlation coefficient was used to investigate the relationship between total BM B-cell and plasma cell numbers (expressed as a percentage of BM leukocytes) and serum IgM concentration.
Introduction Waldenström macroglobulinemia (WM) is a B-cell lymphoproliferative disorder characterized by immunoglobulin (Ig) M monoclonal gammopathy and bone marrow (BM) infiltration by lymphoplasmacytic lymphoma.1,2 It is well recognized that there is considerable clinical heterogeneity, and, in particular, IgM concentrations can vary widely from patient to patient. Previous studies failed to demonstrate a correlation between IgM concentration and overall BM disease burden.3,4 From a pathologic perspective, it is known that lymphoplasmacytic lymphoma shows a spectrum of morphologies from mature lymphocytes to plasma cells and that there can be considerable interpatient variability in this cellular content.1-3,5,6 In this study, we chose to explore the relationship between BM cellular heterogeneity and IgM concentration by accurately enumerating both B-cell and plasma cell components with multiparameter flow cytometry and correlating this with serum IgM concentrations.
HMDS Laboratory, Leeds Teaching Hospitals NHS Trust, Leeds, UK Address for correspondence: Roger G. Owen, MD, HMDS Laboratory, Level 3, Bexley Wing’s University Hospital, Beckett Street, Leeds LS9 7TF, UK Fax: ⫹44 113 206 7883; e-mail contact: [email protected].
2152-2650/$ - see frontmatter © 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.clml.2013.02.018
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IgM Concentration and BM Plasma Cells in WM Figure 1 Correlation of Bone Marrow B Cells and Plasma Cells With Serum Immunoglobulin (Ig) M Concentration in Waldenström Macroglobulinemia. (A) B-cell Numbers Do Not Appear to Correlate With IgM (r ⴝ 0.071, P ⴝ .5); (B) Whereas Some Correlation Is Noted With Bone Marrow Plasma Cell Numbers (r ⴝ 0.452, P ⴝ .01)
A
80 70
B-cell percentage
60 50 40 30 20 10 0
B
0
10
20
30 IgM level
40
50
60
0
10
20
30 IgM level
40
50
60
9 8
Plasma cell percentage
7 6 5 4 3 2 1 0
Results and Discussion WM is a heterogeneous disorder in which there is significant variation in serum IgM levels and BM cellular content. Some previous studies failed to demonstrate a correlation between serum IgM levels and overall BM cellular burden.3,4 Similarly, prognostic factor analyses have failed to demonstrate a reproducible effect of IgM concentration on overall outcome except in the context of very high levels when additional factors, eg, cardiovascular may contribute to poor overall survival.7,8 Given this cellular heterogeneity, we chose to evaluate the relationship between both BM B cells and plasma cells, and serum IgM on the assumption that the plasma cells would be responsible for the majority of IgM secretion. Monotypic B cells were demonstrable in all patients and comprised a median of 6% of BM leukocytes (range, 0.19%-67%) but did not appear to correlate with serum IgM concentration (r ⫽ 0.071, P ⫽ .5) (Figure 1A). Plasma cells were demonstrable in all cases but were typically present at lower
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levels, comprising a median 0.52% of BM leukocytes (range, 0.05%8.4%). In contrast to B cells, plasma cell numbers did show some correlation with IgM concentration (r ⫽ 0.452, P ⫽ .01) (Figure 1B). These data would support the hypothesis that IgM levels in WM are, at least in part, related to the degree of plasma cell differentiation within the tumor population. This is further supported by 2 recent publications. Pasricha et al9 estimated BM B-cell and plasma-cell infiltration by using CD20 and CD138 immunostaining on trephine biopsy sections, and found that IgM levels correlated with the plasma cell burden but not with the B-cell burden. Similarly, Kyrtsonis et al10 found that the overall level of CD138⫹ expression, and, by implication, the proportion of plasma cells as assessed by immunohistochemistry also correlated with IgM levels. It is interesting to that note that plasma cells are found in relatively small numbers in WM but still appear to correlate with IgM levels. In
Ruth M. de Tute et al the current study, we demonstrated a median of 0.52% plasma cells, which is almost identical to that reported by Morice et al6 (median, 0.7%) who used similar flow cytometric methodology. These levels are consistently lower than that reported for CD138 staining of trephine biopsy sections (median, 5%-10%),6,9 and there are likely to be a number of methodologic factors to account for this apparent discrepancy. It is well established that flow cytometry consistently underestimates the overall plasma cell burden in myeloma, principally as a consequence of sample dilution, but, despite this, it appears to be a more relevant prognostic factor than standard morphologic assessment.11,12 Plasma cell distribution may also be a factor in WM because clonal plasma cells have been reported distinct from areas of lymphoid infiltration by some investigators.6 There now are some emerging data regarding the clinical significance of the heterogeneity of cellular content in WM. It is clear that monotypic plasma cells, typically CD20⫺ and CD138⫹ are demonstrable in virtually all patients with WM, but they usually are present in lower numbers than monotypic B cells.5,6,9 Plasma cells, however, may be the dominant cell type in some patients, and, in this setting, the rare entity of IgM myeloma becomes part of the differential diagnosis.6,9 The latter, however, is readily distinguished from WM on the basis of an aberrant plasma cell phenotype (plasma cells are typically CD19– in IgM myeloma but CD19⫹ in WM) and the presence of IGH translocations and t(11;14) in particular.6,13 Recent studies have also demonstrated the significance of the CD138⫹ plasma cell component in the context of response assessment because it appears that some therapies, particularly monoclonal antibody and purine analogue– based therapies appear to selectively deplete the B-cell component but spare the plasma cell component, which can result in delayed IgM responses.14-16 Furthermore, it also has been noted that the cellular composition seen in some patients can vary over time because plasma cell–rich lesions that mimic plasmacytoma have been described at progression with some patients.17,18 Therapeutic strategies in WM have traditionally targeted the monotypic B-cell component of the disease, but it is clear that antigenic targets for monoclonal antibodies, such as CD20 and CD52, are expressed at significantly lower levels in plasma cells.14 It may be prudent to consider plasma cell– directed therapies in certain clinical scenarios, such as the hyperviscosity syndrome in which rapid disease control is required, or IgM-related syndromes and amyloidosis in which the clinical sequelae are a consequence of the M protein rather than tumor bulk per se. In this context, it is clear that bortezomibbased therapies can result in rapid IgM responses, whereas plasma cell–specific monoclonal antibodies, eg, elotuzumab, may also have a role in these clinical situations.19,20
Conclusion We would conclude that IgM levels in WM appear to correlate more with the degree of plasma cell differentiation than the overall
BM disease burden. Appreciation of the plasma cell component in WM is becoming increasingly relevant in the routine clinical management of patients and may indeed be a legitimate therapeutic target in certain clinical scenarios.
Disclosure The authors have stated that they have no conflicts of interest.
References 1. Owen RG, Treon SP, Al-Katib A, et al. Clinicopathological definition of Waldenstrom’s macroglobulinemia: consensus panel recommendations from the Second International Workshop on Waldenstrom’s Macroglobulinemia. Semin Oncol 2003; 30:110-5. 2. Swerdlow SH, Campo E, Harris NL, et al. World Health Organization classification of tumours of haematopoietic and lymphoid tissues. Lyon, France: IARC; 2008. 3. Bartl R, Frisch B, Mahl G, et al. Bone marrow histology in Waldenstrom’s macroglobulinaemia. Clinical relevance of subtype recognition. Scand J Haematol 1983; 31:359-75. 4. Treon SP. How I treat Waldenström macroglobulinemia. Blood 2009; 114:2375-85. 5. Remstein ED, Hanson CA, Kyle RA, et al. Despite apparent morphologic and immunophenotypic heterogeneity, Waldenstrom’s macroglobulinemia is consistently composed of cells along a morphologic continuum of small lymphocytes, plasmacytoid lymphocytes and plasma cells. Semin Oncol 2003; 30:182-6. 6. Morice WG, Chen D, Kurtin PJ, et al. Novel immunophenotypic features of marrow lymphoplasmacytic lymphoma and correlation with Waldenstrom’s macroglobulinemia. Mod Pathol 2009; 22:807-16. 7. Morel P, Duhamel A, Gobbi P, et al. International prognostic scoring system for Waldenström macroglobulinemia. Blood 2009; 113:4163-70. 8. Morel P, Merlini G. Risk stratification in Waldenström macroglobulinemia. Expert Rev Hematol 2012; 5:187-99. 9. Pasricha SR, Juneja SK, Westerman DA, et al. Bone-marrow plasma cell burden correlates with IgM paraprotein concentration in Waldenström macroglobulinemia. J Clin Pathol 2011; 64:520-3. 10. Kyrtsonis MC, Levidou G, Korkolopoulou P, et al. CD138 expression helps distinguish Waldenstrom’s macroglobulinemia (WM) from splenic marginal zone lymphoma. Clin Lymphoma Myeloma Leuk 2011; 11:99-102. 11. Rawstron AC, Orfao A, Beksac M, et al. Report of the European myeloma network on multi-parametric flow cytometry in multiple myeloma and related disorders. Haematologica 2008; 93:431-8. 12. Paiva B, Vidriales MB, Pérez JJ, et al. Multi-parameter flow cytometry quantification of bone marrow plasma cells at diagnosis provides more prognostic information than morphological assessment in myeloma patients. Haematologica 2009; 94: 1599-602. 13. Feyler S, O’Connor SJ, Rawstron AC, et al. IgM myeloma: a rare entity characterized by a CD20-CD56-CD117- immuno-phenotype and the t(11;14). Br J Haematol 2008; 140:547-51. 14. Owen RG, Hillmen P, Rawstron AC. CD52 expression in Waldenstrom’s macroglobulinemia: implications for alemtuzumab therapy and response assessment. Clin Lymphoma 2005; 5:278-81. 15. Varghese AM, Rawstron AC, Ashcroft AJ, et al. Assessment of bone marrow response in Waldenstrom’s macroglobulinemia. Clin Lymphoma Myeloma 2009; 9:53-5. 16. Barakat FH, Medeiros LJ, Wei EX, et al. Residual monotypic plasma cells in patients with Waldenström macroglobulinemia after therapy. Am J Clin Pathol 2011; 135:365-73. 17. Bashir Q, Lee CK, Stuart RW, et al. Phenotypic evolution of Waldenstrom’s macroglobulinemia to extramedullary plasmacytoma. J Clin Oncol 2008; 26:2408-10. 18. Owen RG, Bynoe AG, Varghese A, et al. Heterogeneity of histological transformation events in Waldenstrom’s macroglobulinemia (WM) and related disorders. Clin Lymphoma Myeloma Leuk 2011; 11:176-9. 19. Treon SP, Ioakimidis L, Soumerai JD, et al. Primary therapy of Waldenström macroglobulinemia with bortezomib, dexamethasone, and rituximab: WMCTG clinical trial 05-180. J Clin Oncol 2009; 27:3830-5. 20. Hsi ED, Steinle R, Balasa B, et al. CS1, a potential new therapeutic antibody target for the treatment of multiple myeloma. Clin Cancer Res 2008; 14:2775-84.
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Immunoglobulin M Concentration in Waldenström Macroglobulinemia: Correlation With Bone Marrow B Cells and Plasma Cells Ruth M. de Tute, Andy C. Rawstron, Roger G. Owen Clinical Lymphoma, Myeloma & Leukemia, Vol. 13, No. 2, 211-3 © 2013 Elsevier Inc. All rights reserved. Keywords: Immunoglobulin M, Plasma cell, Waldenström macroglobulinemia
Abstract
Patients and Methods
Serum immunoglobulin (Ig) M levels vary considerably among patients with Waldenström macroglobulinemia, and previous studies have failed to demonstrate a correlation with overall bone marrow disease burden. In this study, bone marrow B cells and plasma cells were enumerated by flow cytometry and correlated with serum IgM concentrations. Monotypic B cells comprised a median of 6% of bone marrow leukocytes but did not correlate with IgM levels (r ⫽ 0.071, P ⫽ .5). Plasma cells, although typically present in lower numbers (median, 0.52%) did show a correlation with IgM (r ⫽ 0.452, P ⫽ .01). IgM levels in Waldenström macroglobulinemia, at least in part, correlate with the degree of plasma cell differentiation seen within the tumor.
Eighty-seven patients (49 men, 38 women) fulfilling consensus panel and World Health Organization diagnostic criteria1,2 for WM were included in this analysis. The median age was 75 years (range, 37-91 years) and the median IgM (M component concentration determined by densitometry) was 11 g/L (range, 1-51 g/L). All the patients were assessed at initial presentation and were untreated. Multiparameter flow cytometry was performed in a single supraregional hematopathology laboratory (HMDS Laboratory, Leeds, UK), and BM samples were received from 25 individual clinical hematology centers. IgM concentrations were determined in local laboratories. Leukocytes were prepared from fresh BM aspirate samples by incubation in a 10-fold excess of ammonium chloride (8.6 g/L in distilled water; Vickers Laboratories, Pudsey, UK) for 5 minutes at 37°C. Cells were then washed twice in buffer (FACSFlow, which contained 0.3% bovine serum albumin; Sigma-Aldrich, Dorset, UK). Leukocytes (approximately 1 ⫻ 106) were stained with 30 L of a preprepared cocktail of antibodies. For plasma cell enumeration, this comprised the following: CD27 fluorescein isothiocyanate (FITC), CD56 phycoerythrin (PE), CD19 peridin chlorophyll protein-cyanine 5.5 (PerCP-Cy5.5), CD38 phycoerythrincyanine (PE-Cy7), CD45 allophycocyanin-cyanine (APC-H7) (all from Becton Dickinson, Oxford, UK), and CD138 allophycocyanin (APC) (Miltenyi Biotec, Bisley, UK). The panel used for B-cell enumeration comprised Lambda FITC, Kappa PE, CD19 PerCP-Cy5.5, CD5 PE-Cy7, CD20 APC, and CD45 APC-H7 (all from Becton Dickinson). After incubation in the dark at 4°C for 20 minutes, the cells were washed twice in buffer and resuspended in FACSFlow. Samples were acquired on a FACSCanto II analyzer by using FACSDiva software (BD Biosciences, Oxford, UK), and a minimum of 100,000 events were acquired for each screening tube. The Pearson correlation coefficient was used to investigate the relationship between total BM B-cell and plasma cell numbers (expressed as a percentage of BM leukocytes) and serum IgM concentration.
Introduction Waldenström macroglobulinemia (WM) is a B-cell lymphoproliferative disorder characterized by immunoglobulin (Ig) M monoclonal gammopathy and bone marrow (BM) infiltration by lymphoplasmacytic lymphoma.1,2 It is well recognized that there is considerable clinical heterogeneity, and, in particular, IgM concentrations can vary widely from patient to patient. Previous studies failed to demonstrate a correlation between IgM concentration and overall BM disease burden.3,4 From a pathologic perspective, it is known that lymphoplasmacytic lymphoma shows a spectrum of morphologies from mature lymphocytes to plasma cells and that there can be considerable interpatient variability in this cellular content.1-3,5,6 In this study, we chose to explore the relationship between BM cellular heterogeneity and IgM concentration by accurately enumerating both B-cell and plasma cell components with multiparameter flow cytometry and correlating this with serum IgM concentrations.
HMDS Laboratory, Leeds Teaching Hospitals NHS Trust, Leeds, UK Address for correspondence: Roger G. Owen, MD, HMDS Laboratory, Level 3, Bexley Wing’s University Hospital, Beckett Street, Leeds LS9 7TF, UK Fax: ⫹44 113 206 7883; e-mail contact: [email protected].
2152-2650/$ - see frontmatter © 2013 Elsevier Inc. All rights reserved. http://dx.doi.org/10.1016/j.clml.2013.02.018
Clinical Lymphoma, Myeloma & Leukemia April 2013
211
IgM Concentration and BM Plasma Cells in WM Figure 1 Correlation of Bone Marrow B Cells and Plasma Cells With Serum Immunoglobulin (Ig) M Concentration in Waldenström Macroglobulinemia. (A) B-cell Numbers Do Not Appear to Correlate With IgM (r ⴝ 0.071, P ⴝ .5); (B) Whereas Some Correlation Is Noted With Bone Marrow Plasma Cell Numbers (r ⴝ 0.452, P ⴝ .01)
A
80 70
B-cell percentage
60 50 40 30 20 10 0
B
0
10
20
30 IgM level
40
50
60
0
10
20
30 IgM level
40
50
60
9 8
Plasma cell percentage
7 6 5 4 3 2 1 0
Results and Discussion WM is a heterogeneous disorder in which there is significant variation in serum IgM levels and BM cellular content. Some previous studies failed to demonstrate a correlation between serum IgM levels and overall BM cellular burden.3,4 Similarly, prognostic factor analyses have failed to demonstrate a reproducible effect of IgM concentration on overall outcome except in the context of very high levels when additional factors, eg, cardiovascular may contribute to poor overall survival.7,8 Given this cellular heterogeneity, we chose to evaluate the relationship between both BM B cells and plasma cells, and serum IgM on the assumption that the plasma cells would be responsible for the majority of IgM secretion. Monotypic B cells were demonstrable in all patients and comprised a median of 6% of BM leukocytes (range, 0.19%-67%) but did not appear to correlate with serum IgM concentration (r ⫽ 0.071, P ⫽ .5) (Figure 1A). Plasma cells were demonstrable in all cases but were typically present at lower
212
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levels, comprising a median 0.52% of BM leukocytes (range, 0.05%8.4%). In contrast to B cells, plasma cell numbers did show some correlation with IgM concentration (r ⫽ 0.452, P ⫽ .01) (Figure 1B). These data would support the hypothesis that IgM levels in WM are, at least in part, related to the degree of plasma cell differentiation within the tumor population. This is further supported by 2 recent publications. Pasricha et al9 estimated BM B-cell and plasma-cell infiltration by using CD20 and CD138 immunostaining on trephine biopsy sections, and found that IgM levels correlated with the plasma cell burden but not with the B-cell burden. Similarly, Kyrtsonis et al10 found that the overall level of CD138⫹ expression, and, by implication, the proportion of plasma cells as assessed by immunohistochemistry also correlated with IgM levels. It is interesting to that note that plasma cells are found in relatively small numbers in WM but still appear to correlate with IgM levels. In
Ruth M. de Tute et al the current study, we demonstrated a median of 0.52% plasma cells, which is almost identical to that reported by Morice et al6 (median, 0.7%) who used similar flow cytometric methodology. These levels are consistently lower than that reported for CD138 staining of trephine biopsy sections (median, 5%-10%),6,9 and there are likely to be a number of methodologic factors to account for this apparent discrepancy. It is well established that flow cytometry consistently underestimates the overall plasma cell burden in myeloma, principally as a consequence of sample dilution, but, despite this, it appears to be a more relevant prognostic factor than standard morphologic assessment.11,12 Plasma cell distribution may also be a factor in WM because clonal plasma cells have been reported distinct from areas of lymphoid infiltration by some investigators.6 There now are some emerging data regarding the clinical significance of the heterogeneity of cellular content in WM. It is clear that monotypic plasma cells, typically CD20⫺ and CD138⫹ are demonstrable in virtually all patients with WM, but they usually are present in lower numbers than monotypic B cells.5,6,9 Plasma cells, however, may be the dominant cell type in some patients, and, in this setting, the rare entity of IgM myeloma becomes part of the differential diagnosis.6,9 The latter, however, is readily distinguished from WM on the basis of an aberrant plasma cell phenotype (plasma cells are typically CD19– in IgM myeloma but CD19⫹ in WM) and the presence of IGH translocations and t(11;14) in particular.6,13 Recent studies have also demonstrated the significance of the CD138⫹ plasma cell component in the context of response assessment because it appears that some therapies, particularly monoclonal antibody and purine analogue– based therapies appear to selectively deplete the B-cell component but spare the plasma cell component, which can result in delayed IgM responses.14-16 Furthermore, it also has been noted that the cellular composition seen in some patients can vary over time because plasma cell–rich lesions that mimic plasmacytoma have been described at progression with some patients.17,18 Therapeutic strategies in WM have traditionally targeted the monotypic B-cell component of the disease, but it is clear that antigenic targets for monoclonal antibodies, such as CD20 and CD52, are expressed at significantly lower levels in plasma cells.14 It may be prudent to consider plasma cell– directed therapies in certain clinical scenarios, such as the hyperviscosity syndrome in which rapid disease control is required, or IgM-related syndromes and amyloidosis in which the clinical sequelae are a consequence of the M protein rather than tumor bulk per se. In this context, it is clear that bortezomibbased therapies can result in rapid IgM responses, whereas plasma cell–specific monoclonal antibodies, eg, elotuzumab, may also have a role in these clinical situations.19,20
Conclusion We would conclude that IgM levels in WM appear to correlate more with the degree of plasma cell differentiation than the overall
BM disease burden. Appreciation of the plasma cell component in WM is becoming increasingly relevant in the routine clinical management of patients and may indeed be a legitimate therapeutic target in certain clinical scenarios.
Disclosure The authors have stated that they have no conflicts of interest.
References 1. Owen RG, Treon SP, Al-Katib A, et al. Clinicopathological definition of Waldenstrom’s macroglobulinemia: consensus panel recommendations from the Second International Workshop on Waldenstrom’s Macroglobulinemia. Semin Oncol 2003; 30:110-5. 2. Swerdlow SH, Campo E, Harris NL, et al. World Health Organization classification of tumours of haematopoietic and lymphoid tissues. Lyon, France: IARC; 2008. 3. Bartl R, Frisch B, Mahl G, et al. Bone marrow histology in Waldenstrom’s macroglobulinaemia. Clinical relevance of subtype recognition. Scand J Haematol 1983; 31:359-75. 4. Treon SP. How I treat Waldenström macroglobulinemia. Blood 2009; 114:2375-85. 5. Remstein ED, Hanson CA, Kyle RA, et al. Despite apparent morphologic and immunophenotypic heterogeneity, Waldenstrom’s macroglobulinemia is consistently composed of cells along a morphologic continuum of small lymphocytes, plasmacytoid lymphocytes and plasma cells. Semin Oncol 2003; 30:182-6. 6. Morice WG, Chen D, Kurtin PJ, et al. Novel immunophenotypic features of marrow lymphoplasmacytic lymphoma and correlation with Waldenstrom’s macroglobulinemia. Mod Pathol 2009; 22:807-16. 7. Morel P, Duhamel A, Gobbi P, et al. International prognostic scoring system for Waldenström macroglobulinemia. Blood 2009; 113:4163-70. 8. Morel P, Merlini G. Risk stratification in Waldenström macroglobulinemia. Expert Rev Hematol 2012; 5:187-99. 9. Pasricha SR, Juneja SK, Westerman DA, et al. Bone-marrow plasma cell burden correlates with IgM paraprotein concentration in Waldenström macroglobulinemia. J Clin Pathol 2011; 64:520-3. 10. Kyrtsonis MC, Levidou G, Korkolopoulou P, et al. CD138 expression helps distinguish Waldenstrom’s macroglobulinemia (WM) from splenic marginal zone lymphoma. Clin Lymphoma Myeloma Leuk 2011; 11:99-102. 11. Rawstron AC, Orfao A, Beksac M, et al. Report of the European myeloma network on multi-parametric flow cytometry in multiple myeloma and related disorders. Haematologica 2008; 93:431-8. 12. Paiva B, Vidriales MB, Pérez JJ, et al. Multi-parameter flow cytometry quantification of bone marrow plasma cells at diagnosis provides more prognostic information than morphological assessment in myeloma patients. Haematologica 2009; 94: 1599-602. 13. Feyler S, O’Connor SJ, Rawstron AC, et al. IgM myeloma: a rare entity characterized by a CD20-CD56-CD117- immuno-phenotype and the t(11;14). Br J Haematol 2008; 140:547-51. 14. Owen RG, Hillmen P, Rawstron AC. CD52 expression in Waldenstrom’s macroglobulinemia: implications for alemtuzumab therapy and response assessment. Clin Lymphoma 2005; 5:278-81. 15. Varghese AM, Rawstron AC, Ashcroft AJ, et al. Assessment of bone marrow response in Waldenstrom’s macroglobulinemia. Clin Lymphoma Myeloma 2009; 9:53-5. 16. Barakat FH, Medeiros LJ, Wei EX, et al. Residual monotypic plasma cells in patients with Waldenström macroglobulinemia after therapy. Am J Clin Pathol 2011; 135:365-73. 17. Bashir Q, Lee CK, Stuart RW, et al. Phenotypic evolution of Waldenstrom’s macroglobulinemia to extramedullary plasmacytoma. J Clin Oncol 2008; 26:2408-10. 18. Owen RG, Bynoe AG, Varghese A, et al. Heterogeneity of histological transformation events in Waldenstrom’s macroglobulinemia (WM) and related disorders. Clin Lymphoma Myeloma Leuk 2011; 11:176-9. 19. Treon SP, Ioakimidis L, Soumerai JD, et al. Primary therapy of Waldenström macroglobulinemia with bortezomib, dexamethasone, and rituximab: WMCTG clinical trial 05-180. J Clin Oncol 2009; 27:3830-5. 20. Hsi ED, Steinle R, Balasa B, et al. CS1, a potential new therapeutic antibody target for the treatment of multiple myeloma. Clin Cancer Res 2008; 14:2775-84.
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