Defining Lymphoplasmacytic Lymphoma: Does MYD88L265P Define a Pathologically Distinct Entity Among Patients With an IgM Paraprotein and Bone Marrow–Based Low-Grade B-Cell Lymphomas With Plasmacytic Differentiation?

Defining Lymphoplasmacytic Lymphoma: Does MYD88L265P Define a Pathologically Distinct Entity... Abstract Objectives Lymphoplasmacytic lymphoma (LPL) remains a poorly defined entity, even with the discovery of MYD88L265P mutations and association with Waldenström macroglobulinemia (WM). Among bone marrow (BM)–based, low-grade B-cell lymphoma with plasmacytic differentiation (LGBLPD) and immunoglobulin M (IgM) paraproteins, we sought to determine whether MYD88L265P defines a distinct entity and can help refine diagnostic criteria for LPL. Methods BMs diagnosed with LGBLPD or LPL and serum IgM paraprotein were studied (2007-2013). Clinicopathologic features were reviewed and specimens were tested for MYD88L265P. Results In total, 138 (87%) of 159 cases had MYD88L265P, and 158 of 159 were clinically considered WM. MYD88L265P cases had higher disease burden than MYD88WT. Features associated with MYD88L265P include increased mast cells and lymphocyte (not plasma cell)–predominant infiltrate. Hemosiderin, Dutcher bodies, and paratrabecular growth were not associated with MYD88L265P. Conclusions Our data support a clinicopathologic approach to LPL diagnosis and recognition that it may manifest with varying morphologies, phenotypes, and molecular features. Lymphoplasmacytic lymphoma, IgM paraprotein, Waldenström macroglobulinemia, MYD88, Bone marrow Lymphoplasmacytic lymphoma (LPL) is a low-grade B-cell lymphoma (LGBL) exhibiting a spectrum of B-cell differentiation, including small lymphocytes, plasmacytoid lymphocytes, and plasma cells, usually involving bone marrow (BM) and sometimes lymph nodes and spleen, which does not fulfill criteria for any other small B-cell lymphoid neoplasms.1 It typically has nonspecific morphologic features in BM, overlapping with other LGBLs, and can be either plasma cell predominant or lymphocyte predominant. Most patients are clinically diagnosed with Waldenström macroglobulinemia (WM), which is defined as LPL with 10% or more BM involvement and an immunoglobulin M (IgM) monoclonal gammopathy of any concentration. WM can be clinically asymptomatic (smoldering) or symptomatic.2 Common symptoms include constitutional symptoms, hyperviscosity, vision changes, headaches, cytopenias, peripheral neuropathy, cryoglobulinemia, autoimmune phenomena, coagulopathy, hepatosplenomegaly, skin rash (Schnitzler syndrome), and occasionally amyloid light chain (AL) amyloidosis.3,4 In a small fraction of LPLs, the malignant cells synthesize non-IgM immunoglobulin, such as immunoglobulin G or immunoglobulin A,5 or light chains only, or they may be nonsecretory. Distinguishing LPL from other LGBLs, especially marginal zone lymphoma (MZL), in the BM remains a challenge due to overlapping pathologic features. MZL is characterized by small, CD5-negative/CD10-negative neoplastic B lymphocytes in lymph nodes or spleen and may show distinctive marginal zone differentiation with monocytoid cytology. However, these morphologic features are rarely identifiable in the BM. In addition, plasmacytic differentiation, autoimmune manifestations, and monoclonal gammopathy are frequently seen in both LPLs and MZLs.6 Bassarova et al7 showed that LPL might be reliably distinguished from MZL in the BM by assessing a combination of pathologic features, including paratrabecular infiltration, the presence of lymphoplasmacytoid cells and cells with Dutcher bodies, and an increased number of mast cells. However, since none of these features are entirely specific, LBGLs with plasmacytic differentiation (LGBLPD) in the BM are often unable to be definitively classified on morphology and phenotype alone. Although MZL is more often nodal or tissue based and LPL is typically marrow based, splenic marginal zone lymphoma shows some degree of BM involvement in most cases, and conversely, LPL can involve extramedullary sites. Thus, clinical correlation is essential to ensure the pathologist understands the distribution of a given patient’s disease. These cases present a challenge to hematologists who are then faced with deciding a course of therapy based on a nonspecific diagnosis. Next-generation sequencing studies have identified multiple recurring somatic mutations in WM, including MYD88L265P (up to 97%), CXCR4 (30%-40%), ARIDA (17%), and CD79B (8%-15%).8-14MYD88L265P, the first of these to be recognized, plays an important role in the Toll-like receptor signaling pathway and activates nuclear factor–κB, which promotes growth and survival of downstream cells. The MYD88L265P mutation was first identified using whole-genome sequencing and subsequently confirmed to be highly sensitive for the diagnosis of WM.8,10-14 However, MYD88L265P mutations have been observed, although with lower frequency, in other LGBLs as well, including MZL (6%-10%) and chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) (3% to 8%).10-12,14 A second gene, CXCR4, also shows frequent mutations in LPL/WM and, although present at a lower frequency, may be an important therapeutic target in WM.15 As a result of these studies, testing for MYD88L265P mutations has become an integral part of the routine pathologic evaluation of LGBLPD in the BM. However, questions of what exactly defines the pathologic entity of LPL are still faced by the practicing hematopathologist frequently. Although many studies have investigated the incidence of MYD88L265P mutations in well-defined cases of WM/LPL,8-14 and others have compared distinct cases of MZL with well-defined cases of WM/LPL,8,10-14 this study has a different aim. In evaluating a large cohort of LGBLPD in the BM, without a definitive extramedullary diagnosis, we sought to determine whether MYD88L265P mutations define a distinct clinicopathologic entity and can help refine our diagnostic criteria for LPL. As a secondary aim, by beginning with a pathology-derived cohort, we sought to establish whether patients diagnosed with LPL differed clinically or pathologically from those diagnosed with LGBLPD. Materials and Methods Case Selection The study was approved by our institutional review board. The laboratory information system at the Mayo Clinic in Rochester, Minnesota, was queried. A retrospective review of consecutive BM cases from 2007 to 2013 was performed to identify cases diagnosed with either LPL (World Health Organization [WHO]) or LGBLPD with 10% or more BM involvement, serum IgM paraprotein of any size, and frozen cells available for MYD88 testing. Of the 164 cases meeting these criteria, two were excluded for equivocal MYD88 results, two were excluded for a concurrent diagnosis of CLL/SLL, and one was excluded for an extramedullary diagnosis of MZL, leaving 159 cases in the final cohort. Because MYD88L265P was not tested clinically at our institution until 2014, no patients had MYD88L265P testing done during the time of their initial diagnostic evaluation, and thus their initially reported diagnosis was rendered without consideration of MYD88 status. Clinicopathologic Review Wright-Giemsa–stained BM aspirate and H&E core biopsy slides were re-reviewed, when available, without knowledge of MYD88L265P mutation status. Data of other ancillary studies, including flow cytometry and/or cytogenetics, were collected from the electronic medical record when available. Pathologic features, including BM cellularity, percentage of lymphoma involvement, distribution pattern, reactive mast cells, Dutcher bodies, and hemosiderin, were assessed by a hematopathologist (R.L.K.). Cases were assessed by morphologic evaluation for whether the infiltrate was predominantly lymphocytes, predominantly plasma cells, or an approximate equal mixture of both. Pathologic features were assessed on the initial diagnostic BM when available or the first BM performed at our institution when the initial diagnostic marrow was not available. If available, flow cytometric immunophenotyping reports and/or histograms were retrospectively reviewed and the immunophenotype of the clonal B-cell and plasma cell populations was recorded. Flow cytometric immunophenotyping, when performed as part of the diagnostic evaluation, was done according to standard, previously published methods.16-18 Two hematologists (P.K. and J.A.) reviewed the clinical records to record if patients demonstrated a clinical course and features in keeping with a diagnosis of WM, either smoldering or symptomatic. Clinical features associated with symptomatic WM, including vision changes, headache, bleeding, and peripheral neuropathy, were abstracted from the electronic medical record. Diagnosis rendered at time of biopsy (LGBLPD vs LPL) was also recorded. LGBLPD is a less specific diagnosis used in our practice when the pathologist feels there is insufficient evidence to definitively subclassify the infiltrate using a more specific WHO diagnosis. We hypothesized initially, based on anecdotal experience, that these two diagnoses are used somewhat interchangeably in our practice, depending on the pathologist’s comfort level with rendering a definite WHO-based diagnosis. MYD88L265P Mutational Analysis MYD88 mutation status was assessed by the amplification-refractory mutation system (ARMS), a variant of allele-specific polymerase chain reaction (PCR). DNA was extracted using the Qiagen DNeasy kit (Qiagen, Valencia, CA) from archived unsorted BM aspirate sample pellets fixed in methanol–acetic acid. A single-tube multiplex ARMS was performed using primers situated in exon 5 of MYD88 (NM_002468.4), including one primer specifically targeting the c.794T > C; Leu265Pro (L256P) alteration. Reaction products were analyzed using capillary electrophoresis (QIAxcel; Qiagen). MYD88 wild-type control amplification yields a PCR product of 141 base pairs (bp), and if present, an additional specific 72-bp product denotes the L265P mutation. This assay has an analytical sensitivity of approximately 1% mutation detection in a wild-type background. Statistics Statistical analysis was performed using the SAS biostatistical software JMP Pro 10.0.0 (SAS Institute, Cary, NC). Categorical variables were compared using a two-tailed Fisher exact test for 2 × 2 contingency tables or χ2 analysis for larger tables. Continuous variables were compared using the Student t test or Kruskal-Wallis tests. The associations between MYD88 status and the histopathologic features were also analyzed to adjust for potential confounding factors, including age, sex, treatment history, and percentage of BM involvement, using multivariate logistic regression. A P value less than .05 was considered statistically significant. Results Of 159 patients, 138 (87%) had the MYD88L265P mutation. Age and sex were similar between the MYD88L265P and MYD88WT groups Table 1. Upon clinical chart review by our hematologists, virtually all of the patients in this series (158/159, 99%) had clinical features in keeping with a diagnosis of WM and had been managed as such. Among the patients with WM, 138 (87%) of 158 had MYD88L265P. The presence of WM-associated clinical manifestations (vision changes, headache, bleeding, peripheral neuropathy, lymphadenopathy, and organomegaly) and laboratory values did not correlate with MYD88 status. The one non-WM case had cold agglutinin disease with an IgM paraprotein and was MYD88WT. Cases diagnosed with LPL had a slightly lower median hemoglobin (10.2 g/dL vs 11.0 g/dL, P = .04) and slightly higher serum viscosity (2.2 cp vs 1.5 cp, P = .003) than those diagnosed with LGBLPD but otherwise showed no statistically significant difference in the clinical features assessed. In total, 150 patients had treatment data available for review, and 71 (47%) of 150 patients had received some form of therapy prior to being evaluated at our institution. Table 1 Clinical Characteristics, Categorized by MYD88 Mutation Status Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  BM, bone marrow; LPL, lymphoplasmacytic lymphoma. aOthers diagnosed with low-grade B-cell lymphoma with plasmacytic differentiation. View Large Table 1 Clinical Characteristics, Categorized by MYD88 Mutation Status Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  BM, bone marrow; LPL, lymphoplasmacytic lymphoma. aOthers diagnosed with low-grade B-cell lymphoma with plasmacytic differentiation. View Large LPL was originally diagnosed in 52% of MYD88L265P cases compared with 33% of MYD88WT cases, while the remainder were generically classified as LGBLPD (P = .10; Table 1). One case diagnosed with LPL also had concurrent myelodysplastic syndrome. Twenty-two (17 MYD88L265P and five MYD88WT) patients also had biopsy-proven extramedullary tissue or body fluid involvement by their lymphoma, including two patients with transformation to diffuse large B-cell lymphoma. The remaining 20 were diagnosed with either LPL or LGBLPD, in keeping with the BM diagnosis. As described by the WHO, LPL in the BM is composed of small B lymphocytes admixed with a variable amount of plasma cells and plasmacytoid lymphocytes. A representative example of one of the cases in our series is shown in Image 1. Among cases with core biopsy slides available for review, 62 (42%) of 146 had Dutcher bodies and 55 (38%) of 146 had prominent, chunky hemosiderin deposition within macrophages in close association with the lymphoma Image 2A and Image 2B. Lymphomatous infiltrates demonstrated a purely interstitial pattern in 50 (34%) of 146, a nodular/aggregate pattern in 46 (32%) of 146, or both in 50 (34%) of 146. In total, 62 (42%) of 146 cases had paratrabecular aggregates Image 2C. Of cases with aspirate smears available for review, we assessed whether the mast cells were easily identifiable and present in greater than three consecutive high-power fields (×600). By these criteria, 93 (73%) of 127 had increased numbers of cytologically normal mast cells Image 2D. The cases were evaluated for whether the infiltrate was predominantly lymphocytic (112/148, 76%), predominantly mature plasma cells (15/148, 10%), or an approximately equal mixture of both (21/148, 14%) based on the aspirate smear (preferred, when available) or core biopsy specimen. Image 1 View largeDownload slide Bone marrow (BM) features of a representative lymphoplasmacytic lymphoma case from this series. BM biopsy specimen shows extensive involvement (A; H&E, x40) by a relatively monomorphic infiltrate of small lymphocytes and plasma cells (B; H&E, x400). It demonstrates a spectrum of small B lymphocytes, plasmacytoid lymphocytes, and plasma cells (C; Wright-Giemsa, x600). Image 1 View largeDownload slide Bone marrow (BM) features of a representative lymphoplasmacytic lymphoma case from this series. BM biopsy specimen shows extensive involvement (A; H&E, x40) by a relatively monomorphic infiltrate of small lymphocytes and plasma cells (B; H&E, x400). It demonstrates a spectrum of small B lymphocytes, plasmacytoid lymphocytes, and plasma cells (C; Wright-Giemsa, x600). Image 2 View largeDownload slide Commonly described features of lymphoplasmacytic lymphoma in bone marrow include Dutcher bodies (A; H&E, x600), hemosiderin deposition (B; H&E, x600), paratrabecular aggregates (C; H&E, x100), and reactive mast cells (D; Wright-Giemsa, x600). Image 2 View largeDownload slide Commonly described features of lymphoplasmacytic lymphoma in bone marrow include Dutcher bodies (A; H&E, x600), hemosiderin deposition (B; H&E, x600), paratrabecular aggregates (C; H&E, x100), and reactive mast cells (D; Wright-Giemsa, x600). The pathologic features of all cases with slides available for review, segregated by MYD88 status, are described in Table 2. Percentage of BM involvement by lymphoma was higher in MYD88L265P cases than MYD88WT cases (50% vs 20%, P = .0007 after adjustment for age, sex, and prior treatment). Mean (SD) percentage of BM involvement for all cases was 46% (26%; range, 10%-100%). By univariate analysis, histologic features associated with MYD88 status include increased mast cells in the aspirate (78% in MYD88L265P vs 44% in MYD88WT) and lymphocyte (not plasma cell)–predominant infiltrate (78% vs 58%), which remained significant after adjustment for age, sex, prior treatment, and percent BM involvement. Paratrabecular involvement showed a trend toward association with MYD88 status (adjusted P = .07). Presence of Dutcher bodies and prominent hemosiderin were not associated with MYD88 status (adjusted P = .56 and 0.32, respectively). Table 2 Histopathologic Features and Phenotyping by Flow Cytometry, Categorized by MYD88 Mutation Statusa Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  BM, bone marrow. aValues are presented as number (%) unless otherwise indicated. bAdjusted for age, sex, prior treatment (yes/no), and % BM infiltration, unless specified. cAdjusted for age, sex, and prior treatment (yes/no). View Large Table 2 Histopathologic Features and Phenotyping by Flow Cytometry, Categorized by MYD88 Mutation Statusa Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  BM, bone marrow. aValues are presented as number (%) unless otherwise indicated. bAdjusted for age, sex, prior treatment (yes/no), and % BM infiltration, unless specified. cAdjusted for age, sex, and prior treatment (yes/no). View Large Flow cytometric analysis of plasma cells was done in 131 cases and of B cells in 135 cases. Of these, both panels were done in a total of 112 cases. Among the cases with B-cell flow cytometry, a clonal B-cell population was detected in 131 (97%). Among cases with plasma cell flow cytometry, a clonal plasma cell population was identified in 113 (86%). Among cases with both panels performed, clonal B cells and plasma cells were detected in 94 (84%). Of cases with clonal B cells detected by flow cytometry, CD5 was expressed in 20 (18%) of 113 cases and CD10 in three (3%) of 109. Plasma cells in most cases retained expression of CD19 (86/94, 91%) and/or CD45 (89/96, 93%). Of these markers, only CD45 expression by plasma cells was associated with MYD88L265P (adjusted P < .01) (Table 2). Eighty-eight patients had karyotyping and 50 had some type of fluorescent in situ hybridization (FISH) performed on the BM aspirate. Abnormal karyotype (except for isolated –Y) was seen in six (7%) of 88 cases, and an abnormal FISH result was seen in eight (16%) of 50 cases. Results were not specific for a given diagnosis in any case (Supplemental Table 1; all supplemental materials can be found at American Journal of Clinical Pathology online). However, several of the abnormalities seen have been specifically described in LPL, including trisomy 4, 6q deletion, 11q deletion, and gains of chromosome 3.19,20 One case had a t(11;14) identified by karyotype, although breakpoints were not compatible with CCND1-IGH. Other cases had abnormalities of chromosome 12, which have been described in a variety of lymphoid malignancies, albeit not commonly in LPL.21 None of the karyotypic abnormalities were felt to be diagnostic of an underlying myeloid neoplasm or to argue strongly against a diagnosis of LPL. Discussion The diagnosis of LPL is often challenging to pathologists due to morphologic and immunophenotypic overlap with MZL, as well as other LGBLs with plasmacytic differentiation and sometimes multiple myeloma (MM). The discovery of a highly recurrent point mutation, MYD88L265P, in WM has offered an appealing tool for pathologists and clinicians to aid in the differential diagnosis.8-14 However, although MYD88L265P is present in 70% to 100% of patients with WM in various series, at least 10% of MZLs may harbor the mutation, as well,10,14 thereby limiting its utility as a disease-defining mutation. While several studies have evaluated the prevalence of MYD88 mutations in pathologically and clinically well-defined cases of LPL/WM as well as MZL,8,10-14 our study aimed to evaluate the utility of this mutation in identifying a pathologically and clinically distinct group of patients when faced with a LGBLPD in the BM and an IgM paraprotein. In doing so, we sought to determine whether we could better define MYD88L265P-mutated LPL as a distinct pathologic entity. Secondarily, we sought to understand whether cases nonspecifically diagnosed in our practice as LGBLPD represented true LPL or a heterogeneous group. The prevalence of MYD88L265P (87%) in our cohort is similar to that observed in other published cohorts of LPL/WM, supporting our conclusion that most of these cases represent true LPL/WM.13,14,22 Virtually all of the patients studied, with the exception of one patient with IgM cold agglutinin disease (MYD88WT), were, on retrospective review, found to have been considered and treated as having WM clinically. Only 18.9% of these patients had any classic WM symptoms of hyperviscosity, vision changes, headache, bleeding, or neuropathy. Patients diagnosed with LPL (rather than LGBLPD) had a slightly lower median hemoglobin and slightly higher serum viscosity, both of which may be attributable to the slightly higher disease burden among those diagnosed with LPL. Overall, these findings suggest that, despite morphologic, phenotypic, and genetic heterogeneity, these cases seem to all represent a spectrum of true LPL with 158 presenting clinically as WM and one as cold agglutinin disease. In accordance with some prior studies, patients with MYD88L265P had higher disease burden than those with MYD88WT in our cohort.23,24 However, despite initial reports demonstrating adverse prognosis among MYD88WT patients,24 a recent publication from our group, including some overlapping patients with our study as well as others, demonstrated no difference in overall survival between MYD88WT and MYD88L265P patients with WM.25 The current WHO classification acknowledges a large amount of morphologic heterogeneity within the entity of LPL, which was primarily derived from descriptions of this disease process prior to the discovery of the MYD88 mutation.1 Other reports support the assertion that LPL is heterogeneous but indicate that LPL in the BM is identifiable by the presence of paratrabecular aggregates, presence of lymphoplasmacytoid cells, Dutcher bodies, and increased numbers of mast cells.7,26,27 A recent study found these features significantly more often in LPL than in MZL, suggesting their utility in distinguishing the two entities.7 Importantly, this study was done in a cohort of clinically well-defined LPL cases, most of which had MYD88L265P. This raises the question of whether some of the classic morphologic features of LPL are specific to those cases that harbor the mutation, thus supporting MYD88L265P LPL as a unique and identifiable pathologic entity. Our study, however, demonstrates that although some features, including mast cells (78% vs 44%, P = .007), are statistically associated with MYD88L265P, these are present frequently in MYD88WT cases as well. Features such as Dutcher bodies and hemosiderin deposition showed no correlation with MYD88 status in our cohort (Table 2). In addition, there were no significant differences in B-cell immunophenotype between the MYD88WT and MYD88L265P cohorts. Plasma cells in MYD88L265P cases more often showed at least partial CD45 expression compared with MYD88WT. Not surprisingly, most of our cases showed lymphocyte-predominant infiltrates, but a significant minority showed either plasma cell predominance (10%) or approximately equal numbers of plasma cells and lymphocytes (14%), again supporting the morphologic heterogeneity in this disease. In our study, a plasma cell–predominant infiltrate is significantly associated with MYD88WT. Importantly, these seven cases (MYD88WT/plasma cell predominant) did not have lytic bone lesions or other clinical features of MM, showed associated B-cell populations by immunohistochemistry or flow cytometry, and were negative for cyclin D1, making IgM MM unlikely. Although karyotype and FISH are not routinely performed at our institution in the workup of LGBCL in the BM, a subset of our cases did have data available. These results support other studies showing a preponderance of nonspecific abnormalities. Although clearly a useful addition to the routine workup of BM-based LGBLPD, our data suggest that MYD88L265P alone, even within a relatively narrow spectrum of patients with IgM paraproteins and BM-based disease, does not define a single uniform pathologic entity. As previously demonstrated in work by our group, the WHO entity of LPL is not limited to patients with IgM paraproteins and thus is not always synonymous with WM, at least by currently accepted definitions.1,5 The data here further assert that LPL is not limited to patients with MYD88 mutations or those with “typical” pathologic features such as paratrabecular infiltrates and reactive mast cells. There is no doubt that the discovery of MYD88L265P mutations in LPL has helped to clarify both diagnostic and therapeutic issues within the realm of this complex disease. However, as with any new diagnostic test, it has also raised many questions regarding whether histopathologic, molecular, or clinical features should take precedence in disease classification. As our data have shown, even when faced with difficult-to-classify, BM-based lymphoplasmacytic infiltrates, MYD88L265P does not define a distinct phenotypic or morphologic entity. Overall, our findings suggest that despite pathologic heterogeneity, these cases seem to all represent a spectrum of true LPL/WM. What our data most support is a clinicopathologic approach to the diagnosis of LPL and recognition that this intriguing disease may manifest with varying morphologies, phenotypes, and molecular features. From the pathologists’ perspective, our data support a more inclusive use of the WHO-defined diagnosis of LPL. Finally, while MYD88L265P itself may not define LPL as an entity, it clearly represents an exciting biomarker for therapeutic decisions and predicting response to certain therapies. References 1. Swerdlow SH, Campo E, Harris NL, et al.   WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues . Rev. 4th ed. Lyon, France: IARC; 2017. 2. Kyle RA, Benson JT, Larson DR, et al.   Progression in smoldering Waldenstrom macroglobulinemia: long-term results. Blood . 2012; 119: 4462- 4466. Google Scholar CrossRef Search ADS PubMed  3. Dispenzieri A, Buadi F, Kumar SK, et al.   Treatment of immunoglobulin light chain amyloidosis: Mayo stratification of myeloma and risk-adapted therapy (mSMART) consensus statement. Mayo Clin Proc . 2015; 90: 1054- 1081. Google Scholar CrossRef Search ADS PubMed  4. Kapoor P, Ansell SM, Fonseca R, et al.   Diagnosis and management of Waldenström macroglobulinemia: Mayo stratification of macroglobulinemia and risk-adapted therapy (mSMART) guidelines 2016. JAMA Oncol . 2017;3:1257-1265. 5. King RL, Gonsalves WI, Ansell SM, et al.   Lymphoplasmacytic lymphoma with a non-IgM paraprotein shows clinical and pathologic heterogeneity and may harbor MYD88 L265p mutations. Am J Clin Pathol . 2016; 145: 843- 851. Google Scholar CrossRef Search ADS PubMed  6. Berger F, Traverse-Glehen A, Felman P, et al.   Clinicopathologic features of Waldenstrom’s macroglobulinemia and marginal zone lymphoma: are they distinct or the same entity? Clin Lymphoma . 2005; 5: 220- 224. Google Scholar CrossRef Search ADS PubMed  7. Bassarova A, Trøen G, Spetalen S, et al.   Lymphoplasmacytic lymphoma and marginal zone lymphoma in the bone marrow: paratrabecular involvement as an important distinguishing feature. Am J Clin Pathol . 2015; 143: 797- 806. Google Scholar CrossRef Search ADS PubMed  8. Ansell SM, Hodge LS, Secreto FJ, et al.   Activation of TAK1 by MYD88 L265p drives malignant B-cell growth in non-Hodgkin lymphoma. Blood Cancer J . 2014; 4: e183. Google Scholar CrossRef Search ADS PubMed  9. Hunter ZR, Yang G, Xu L, et al.   Genomics, signaling, and treatment of Waldenström macroglobulinemia. J Clin Oncol . 2017; 35: 999- 1001. Google Scholar CrossRef Search ADS   10. Jiménez C, Sebastián E, Chillón MC, et al.   MYD88 L265p is a marker highly characteristic of, but not restricted to, Waldenström’s macroglobulinemia. Leukemia . 2013; 27: 1722- 1728. Google Scholar CrossRef Search ADS PubMed  11. Poulain S, Roumier C, Decambron A, et al.   MYD88 L265p mutation in Waldenstrom macroglobulinemia. Blood . 2013; 121: 4504- 4511. Google Scholar CrossRef Search ADS PubMed  12. Treon SP, Xu L, Yang G, et al.   MYD88 L265p somatic mutation in Waldenström’s macroglobulinemia. N Engl J Med . 2012; 367: 826- 833. Google Scholar CrossRef Search ADS PubMed  13. Varettoni M, Arcaini L, Zibellini S, et al.   Prevalence and clinical significance of the MYD88 (L265p) somatic mutation in Waldenstrom’s macroglobulinemia and related lymphoid neoplasms. Blood . 2013; 121: 2522- 2528. Google Scholar CrossRef Search ADS PubMed  14. Xu L, Hunter ZR, Yang G, et al.   MYD88 L265p in Waldenström macroglobulinemia, immunoglobulin M monoclonal gammopathy, and other B-cell lymphoproliferative disorders using conventional and quantitative allele-specific polymerase chain reaction. Blood . 2013; 121: 2051- 2058. Google Scholar CrossRef Search ADS PubMed  15. Hunter ZR, Xu L, Yang G, et al.   The genomic landscape of Waldenstrom macroglobulinemia is characterized by highly recurring MYD88 and WHIM-like CXCR4 mutations, and small somatic deletions associated with B-cell lymphomagenesis. Blood . 2014; 123: 1637- 1646. Google Scholar CrossRef Search ADS PubMed  16. Morice WG, Chen D, Kurtin PJ, Hanson CA, McPhail ED. Novel immunophenotypic features of marrow lymphoplasmacytic lymphoma and correlation with Waldenström’s macroglobulinemia. Mod Pathol . 2009; 22: 807-8 16. Google Scholar CrossRef Search ADS PubMed  17. Morice WG, Hanson CA, Kumar S, et al.   Novel multi-parameter flow cytometry sensitively detects phenotypically distinct plasma cell subsets in plasma cell proliferative disorders. Leukemia . 2007; 21: 2043- 2046. Google Scholar CrossRef Search ADS PubMed  18. Rosado FG, Morice WG, He R, Howard MT, Timm M, McPhail ED. Immunophenotypic features by multiparameter flow cytometry can help distinguish low grade B-cell lymphomas with plasmacytic differentiation from plasma cell proliferative disorders with an unrelated clonal B-cell process. Br J Haematol . 2015; 169: 368-3 76. Google Scholar CrossRef Search ADS PubMed  19. Nguyen-Khac F, Lambert J, Chapiro E, et al.  ; Groupe Français d’Etude de la Leucémie Lymphoïde Chronique et Maladie de Waldenström (GFCLL/MW); Groupe Ouest-Est d’étude des Leucémie Aiguës et Autres Maladies du Sang (GOELAMS); Groupe d’Etude des Lymphomes de l’Adulte (GELA). Chromosomal aberrations and their prognostic value in a series of 174 untreated patients with Waldenström’s macroglobulinemia. Haematologica . 2013; 98: 649- 654. Google Scholar CrossRef Search ADS PubMed  20. Braggio E, Keats JJ, Leleu X, et al.   High-resolution genomic analysis in Waldenström’s macroglobulinemia identifies disease-specific and common abnormalities with marginal zone lymphomas. Clin Lymphoma Myeloma . 2009; 9: 39- 42. Google Scholar CrossRef Search ADS PubMed  21. Hoeve MA, Gisbertz IA, Schouten HC, et al.   Gastric low-grade MALT lymphoma, high-grade MALT lymphoma and diffuse large B cell lymphoma show different frequencies of trisomy. Leukemia . 1999; 13: 799- 807. Google Scholar CrossRef Search ADS PubMed  22. Gachard N, Parrens M, Soubeyran I, et al.   IGHV gene features and MYD88 L265p mutation separate the three marginal zone lymphoma entities and Waldenström macroglobulinemia/lymphoplasmacytic lymphomas. Leukemia . 2013; 27: 183- 189. Google Scholar CrossRef Search ADS PubMed  23. Patkar N, Subramanian PG, Deshpande P, et al.   MYD88 mutant lymphoplasmacytic lymphoma/Waldenström macroglobulinemia has distinct clinical and pathological features as compared to its mutation negative counterpart. Leuk Lymphoma . 2015; 56: 420- 425. Google Scholar CrossRef Search ADS PubMed  24. Treon SP, Cao Y, Xu L, et al.   Somatic mutations in MYD88 and CXCR4 are determinants of clinical presentation and overall survival in Waldenstrom macroglobulinemia. Blood . 2014; 123: 2791- 2796. Google Scholar CrossRef Search ADS PubMed  25. Abeykoon JP, Paludo J, King RL, et al.   MYD88 mutation status does not impact overall survival in Waldenström macroglobulinemia. Am J Hematol . 2018; 93: 187- 194. Google Scholar CrossRef Search ADS PubMed  26. 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- 115. Google Scholar CrossRef Search ADS PubMed  27. Tournilhac O, Santos DD, Xu L, et al.   Mast cells in Waldenstrom’s macroglobulinemia support lymphoplasmacytic cell growth through CD154/CD40 signaling. Ann Oncol . 2006; 17: 1275- 1282. Google Scholar CrossRef Search ADS PubMed  © American Society for Clinical Pathology, 2018. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com This article is published and distributed under the terms of the Oxford University Press, Standard Journals Publication Model (https://academic.oup.com/journals/pages/about_us/legal/notices) http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png American Journal of Clinical Pathology Oxford University Press

Defining Lymphoplasmacytic Lymphoma: Does MYD88L265P Define a Pathologically Distinct Entity Among Patients With an IgM Paraprotein and Bone Marrow–Based Low-Grade B-Cell Lymphomas With Plasmacytic Differentiation?

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Abstract

Abstract Objectives Lymphoplasmacytic lymphoma (LPL) remains a poorly defined entity, even with the discovery of MYD88L265P mutations and association with Waldenström macroglobulinemia (WM). Among bone marrow (BM)–based, low-grade B-cell lymphoma with plasmacytic differentiation (LGBLPD) and immunoglobulin M (IgM) paraproteins, we sought to determine whether MYD88L265P defines a distinct entity and can help refine diagnostic criteria for LPL. Methods BMs diagnosed with LGBLPD or LPL and serum IgM paraprotein were studied (2007-2013). Clinicopathologic features were reviewed and specimens were tested for MYD88L265P. Results In total, 138 (87%) of 159 cases had MYD88L265P, and 158 of 159 were clinically considered WM. MYD88L265P cases had higher disease burden than MYD88WT. Features associated with MYD88L265P include increased mast cells and lymphocyte (not plasma cell)–predominant infiltrate. Hemosiderin, Dutcher bodies, and paratrabecular growth were not associated with MYD88L265P. Conclusions Our data support a clinicopathologic approach to LPL diagnosis and recognition that it may manifest with varying morphologies, phenotypes, and molecular features. Lymphoplasmacytic lymphoma, IgM paraprotein, Waldenström macroglobulinemia, MYD88, Bone marrow Lymphoplasmacytic lymphoma (LPL) is a low-grade B-cell lymphoma (LGBL) exhibiting a spectrum of B-cell differentiation, including small lymphocytes, plasmacytoid lymphocytes, and plasma cells, usually involving bone marrow (BM) and sometimes lymph nodes and spleen, which does not fulfill criteria for any other small B-cell lymphoid neoplasms.1 It typically has nonspecific morphologic features in BM, overlapping with other LGBLs, and can be either plasma cell predominant or lymphocyte predominant. Most patients are clinically diagnosed with Waldenström macroglobulinemia (WM), which is defined as LPL with 10% or more BM involvement and an immunoglobulin M (IgM) monoclonal gammopathy of any concentration. WM can be clinically asymptomatic (smoldering) or symptomatic.2 Common symptoms include constitutional symptoms, hyperviscosity, vision changes, headaches, cytopenias, peripheral neuropathy, cryoglobulinemia, autoimmune phenomena, coagulopathy, hepatosplenomegaly, skin rash (Schnitzler syndrome), and occasionally amyloid light chain (AL) amyloidosis.3,4 In a small fraction of LPLs, the malignant cells synthesize non-IgM immunoglobulin, such as immunoglobulin G or immunoglobulin A,5 or light chains only, or they may be nonsecretory. Distinguishing LPL from other LGBLs, especially marginal zone lymphoma (MZL), in the BM remains a challenge due to overlapping pathologic features. MZL is characterized by small, CD5-negative/CD10-negative neoplastic B lymphocytes in lymph nodes or spleen and may show distinctive marginal zone differentiation with monocytoid cytology. However, these morphologic features are rarely identifiable in the BM. In addition, plasmacytic differentiation, autoimmune manifestations, and monoclonal gammopathy are frequently seen in both LPLs and MZLs.6 Bassarova et al7 showed that LPL might be reliably distinguished from MZL in the BM by assessing a combination of pathologic features, including paratrabecular infiltration, the presence of lymphoplasmacytoid cells and cells with Dutcher bodies, and an increased number of mast cells. However, since none of these features are entirely specific, LBGLs with plasmacytic differentiation (LGBLPD) in the BM are often unable to be definitively classified on morphology and phenotype alone. Although MZL is more often nodal or tissue based and LPL is typically marrow based, splenic marginal zone lymphoma shows some degree of BM involvement in most cases, and conversely, LPL can involve extramedullary sites. Thus, clinical correlation is essential to ensure the pathologist understands the distribution of a given patient’s disease. These cases present a challenge to hematologists who are then faced with deciding a course of therapy based on a nonspecific diagnosis. Next-generation sequencing studies have identified multiple recurring somatic mutations in WM, including MYD88L265P (up to 97%), CXCR4 (30%-40%), ARIDA (17%), and CD79B (8%-15%).8-14MYD88L265P, the first of these to be recognized, plays an important role in the Toll-like receptor signaling pathway and activates nuclear factor–κB, which promotes growth and survival of downstream cells. The MYD88L265P mutation was first identified using whole-genome sequencing and subsequently confirmed to be highly sensitive for the diagnosis of WM.8,10-14 However, MYD88L265P mutations have been observed, although with lower frequency, in other LGBLs as well, including MZL (6%-10%) and chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) (3% to 8%).10-12,14 A second gene, CXCR4, also shows frequent mutations in LPL/WM and, although present at a lower frequency, may be an important therapeutic target in WM.15 As a result of these studies, testing for MYD88L265P mutations has become an integral part of the routine pathologic evaluation of LGBLPD in the BM. However, questions of what exactly defines the pathologic entity of LPL are still faced by the practicing hematopathologist frequently. Although many studies have investigated the incidence of MYD88L265P mutations in well-defined cases of WM/LPL,8-14 and others have compared distinct cases of MZL with well-defined cases of WM/LPL,8,10-14 this study has a different aim. In evaluating a large cohort of LGBLPD in the BM, without a definitive extramedullary diagnosis, we sought to determine whether MYD88L265P mutations define a distinct clinicopathologic entity and can help refine our diagnostic criteria for LPL. As a secondary aim, by beginning with a pathology-derived cohort, we sought to establish whether patients diagnosed with LPL differed clinically or pathologically from those diagnosed with LGBLPD. Materials and Methods Case Selection The study was approved by our institutional review board. The laboratory information system at the Mayo Clinic in Rochester, Minnesota, was queried. A retrospective review of consecutive BM cases from 2007 to 2013 was performed to identify cases diagnosed with either LPL (World Health Organization [WHO]) or LGBLPD with 10% or more BM involvement, serum IgM paraprotein of any size, and frozen cells available for MYD88 testing. Of the 164 cases meeting these criteria, two were excluded for equivocal MYD88 results, two were excluded for a concurrent diagnosis of CLL/SLL, and one was excluded for an extramedullary diagnosis of MZL, leaving 159 cases in the final cohort. Because MYD88L265P was not tested clinically at our institution until 2014, no patients had MYD88L265P testing done during the time of their initial diagnostic evaluation, and thus their initially reported diagnosis was rendered without consideration of MYD88 status. Clinicopathologic Review Wright-Giemsa–stained BM aspirate and H&E core biopsy slides were re-reviewed, when available, without knowledge of MYD88L265P mutation status. Data of other ancillary studies, including flow cytometry and/or cytogenetics, were collected from the electronic medical record when available. Pathologic features, including BM cellularity, percentage of lymphoma involvement, distribution pattern, reactive mast cells, Dutcher bodies, and hemosiderin, were assessed by a hematopathologist (R.L.K.). Cases were assessed by morphologic evaluation for whether the infiltrate was predominantly lymphocytes, predominantly plasma cells, or an approximate equal mixture of both. Pathologic features were assessed on the initial diagnostic BM when available or the first BM performed at our institution when the initial diagnostic marrow was not available. If available, flow cytometric immunophenotyping reports and/or histograms were retrospectively reviewed and the immunophenotype of the clonal B-cell and plasma cell populations was recorded. Flow cytometric immunophenotyping, when performed as part of the diagnostic evaluation, was done according to standard, previously published methods.16-18 Two hematologists (P.K. and J.A.) reviewed the clinical records to record if patients demonstrated a clinical course and features in keeping with a diagnosis of WM, either smoldering or symptomatic. Clinical features associated with symptomatic WM, including vision changes, headache, bleeding, and peripheral neuropathy, were abstracted from the electronic medical record. Diagnosis rendered at time of biopsy (LGBLPD vs LPL) was also recorded. LGBLPD is a less specific diagnosis used in our practice when the pathologist feels there is insufficient evidence to definitively subclassify the infiltrate using a more specific WHO diagnosis. We hypothesized initially, based on anecdotal experience, that these two diagnoses are used somewhat interchangeably in our practice, depending on the pathologist’s comfort level with rendering a definite WHO-based diagnosis. MYD88L265P Mutational Analysis MYD88 mutation status was assessed by the amplification-refractory mutation system (ARMS), a variant of allele-specific polymerase chain reaction (PCR). DNA was extracted using the Qiagen DNeasy kit (Qiagen, Valencia, CA) from archived unsorted BM aspirate sample pellets fixed in methanol–acetic acid. A single-tube multiplex ARMS was performed using primers situated in exon 5 of MYD88 (NM_002468.4), including one primer specifically targeting the c.794T > C; Leu265Pro (L256P) alteration. Reaction products were analyzed using capillary electrophoresis (QIAxcel; Qiagen). MYD88 wild-type control amplification yields a PCR product of 141 base pairs (bp), and if present, an additional specific 72-bp product denotes the L265P mutation. This assay has an analytical sensitivity of approximately 1% mutation detection in a wild-type background. Statistics Statistical analysis was performed using the SAS biostatistical software JMP Pro 10.0.0 (SAS Institute, Cary, NC). Categorical variables were compared using a two-tailed Fisher exact test for 2 × 2 contingency tables or χ2 analysis for larger tables. Continuous variables were compared using the Student t test or Kruskal-Wallis tests. The associations between MYD88 status and the histopathologic features were also analyzed to adjust for potential confounding factors, including age, sex, treatment history, and percentage of BM involvement, using multivariate logistic regression. A P value less than .05 was considered statistically significant. Results Of 159 patients, 138 (87%) had the MYD88L265P mutation. Age and sex were similar between the MYD88L265P and MYD88WT groups Table 1. Upon clinical chart review by our hematologists, virtually all of the patients in this series (158/159, 99%) had clinical features in keeping with a diagnosis of WM and had been managed as such. Among the patients with WM, 138 (87%) of 158 had MYD88L265P. The presence of WM-associated clinical manifestations (vision changes, headache, bleeding, peripheral neuropathy, lymphadenopathy, and organomegaly) and laboratory values did not correlate with MYD88 status. The one non-WM case had cold agglutinin disease with an IgM paraprotein and was MYD88WT. Cases diagnosed with LPL had a slightly lower median hemoglobin (10.2 g/dL vs 11.0 g/dL, P = .04) and slightly higher serum viscosity (2.2 cp vs 1.5 cp, P = .003) than those diagnosed with LGBLPD but otherwise showed no statistically significant difference in the clinical features assessed. In total, 150 patients had treatment data available for review, and 71 (47%) of 150 patients had received some form of therapy prior to being evaluated at our institution. Table 1 Clinical Characteristics, Categorized by MYD88 Mutation Status Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  BM, bone marrow; LPL, lymphoplasmacytic lymphoma. aOthers diagnosed with low-grade B-cell lymphoma with plasmacytic differentiation. View Large Table 1 Clinical Characteristics, Categorized by MYD88 Mutation Status Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  Category  MYD88L265P  MYD88WT  P Value  Basic characteristics   Age, mean ± SD, y  67 ± 11  69 ± 12  .44   Female sex, No. (%)  37/138 (27)  10/21 (48)  .06   BM diagnosis, No. (%) diagnosed with LPLa  72/138 (52)  7/21 (33)  .10  Clinical symptoms, No. (%)   Vision changes  5/108 (5)  0/17 (0)  >.99   Headache  5/107 (5)  2/17 (12)  .25   Bleeding  10/108 (9)  0/17 (0)  .36  Clinical manifestations, No. (%)   Amyloid  2/113 (2)  2/17 (12)  .08   Neuropathy  12/107 (11)  2/17 (12)  >.99   Lymphadenopathy  32/107 (30)  5/17 (29)  .97   Organomegaly  13/113 (12)  3/17 (18)  .44  Laboratory parameters, median (range)   Hemoglobin, g/dL  10.8 (5.0-15.6)  10.2 (5.1-14.2)  .55   Platelet count, × 109/L  219 (0.3-2310)  191 (39-510)  .36   β2-Microglobulin, mg/L  3.4 (0-48)  3.1 (1.4-7.5)  .64   Serum viscosity, cp  1.7 (1.0-8.1)  2.0 (1.0-2.6)  .93   Lactate dehydrogenase, U/L  141 (2-848)  172 (88-498)  .20  BM, bone marrow; LPL, lymphoplasmacytic lymphoma. aOthers diagnosed with low-grade B-cell lymphoma with plasmacytic differentiation. View Large LPL was originally diagnosed in 52% of MYD88L265P cases compared with 33% of MYD88WT cases, while the remainder were generically classified as LGBLPD (P = .10; Table 1). One case diagnosed with LPL also had concurrent myelodysplastic syndrome. Twenty-two (17 MYD88L265P and five MYD88WT) patients also had biopsy-proven extramedullary tissue or body fluid involvement by their lymphoma, including two patients with transformation to diffuse large B-cell lymphoma. The remaining 20 were diagnosed with either LPL or LGBLPD, in keeping with the BM diagnosis. As described by the WHO, LPL in the BM is composed of small B lymphocytes admixed with a variable amount of plasma cells and plasmacytoid lymphocytes. A representative example of one of the cases in our series is shown in Image 1. Among cases with core biopsy slides available for review, 62 (42%) of 146 had Dutcher bodies and 55 (38%) of 146 had prominent, chunky hemosiderin deposition within macrophages in close association with the lymphoma Image 2A and Image 2B. Lymphomatous infiltrates demonstrated a purely interstitial pattern in 50 (34%) of 146, a nodular/aggregate pattern in 46 (32%) of 146, or both in 50 (34%) of 146. In total, 62 (42%) of 146 cases had paratrabecular aggregates Image 2C. Of cases with aspirate smears available for review, we assessed whether the mast cells were easily identifiable and present in greater than three consecutive high-power fields (×600). By these criteria, 93 (73%) of 127 had increased numbers of cytologically normal mast cells Image 2D. The cases were evaluated for whether the infiltrate was predominantly lymphocytic (112/148, 76%), predominantly mature plasma cells (15/148, 10%), or an approximately equal mixture of both (21/148, 14%) based on the aspirate smear (preferred, when available) or core biopsy specimen. Image 1 View largeDownload slide Bone marrow (BM) features of a representative lymphoplasmacytic lymphoma case from this series. BM biopsy specimen shows extensive involvement (A; H&E, x40) by a relatively monomorphic infiltrate of small lymphocytes and plasma cells (B; H&E, x400). It demonstrates a spectrum of small B lymphocytes, plasmacytoid lymphocytes, and plasma cells (C; Wright-Giemsa, x600). Image 1 View largeDownload slide Bone marrow (BM) features of a representative lymphoplasmacytic lymphoma case from this series. BM biopsy specimen shows extensive involvement (A; H&E, x40) by a relatively monomorphic infiltrate of small lymphocytes and plasma cells (B; H&E, x400). It demonstrates a spectrum of small B lymphocytes, plasmacytoid lymphocytes, and plasma cells (C; Wright-Giemsa, x600). Image 2 View largeDownload slide Commonly described features of lymphoplasmacytic lymphoma in bone marrow include Dutcher bodies (A; H&E, x600), hemosiderin deposition (B; H&E, x600), paratrabecular aggregates (C; H&E, x100), and reactive mast cells (D; Wright-Giemsa, x600). Image 2 View largeDownload slide Commonly described features of lymphoplasmacytic lymphoma in bone marrow include Dutcher bodies (A; H&E, x600), hemosiderin deposition (B; H&E, x600), paratrabecular aggregates (C; H&E, x100), and reactive mast cells (D; Wright-Giemsa, x600). The pathologic features of all cases with slides available for review, segregated by MYD88 status, are described in Table 2. Percentage of BM involvement by lymphoma was higher in MYD88L265P cases than MYD88WT cases (50% vs 20%, P = .0007 after adjustment for age, sex, and prior treatment). Mean (SD) percentage of BM involvement for all cases was 46% (26%; range, 10%-100%). By univariate analysis, histologic features associated with MYD88 status include increased mast cells in the aspirate (78% in MYD88L265P vs 44% in MYD88WT) and lymphocyte (not plasma cell)–predominant infiltrate (78% vs 58%), which remained significant after adjustment for age, sex, prior treatment, and percent BM involvement. Paratrabecular involvement showed a trend toward association with MYD88 status (adjusted P = .07). Presence of Dutcher bodies and prominent hemosiderin were not associated with MYD88 status (adjusted P = .56 and 0.32, respectively). Table 2 Histopathologic Features and Phenotyping by Flow Cytometry, Categorized by MYD88 Mutation Statusa Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  BM, bone marrow. aValues are presented as number (%) unless otherwise indicated. bAdjusted for age, sex, prior treatment (yes/no), and % BM infiltration, unless specified. cAdjusted for age, sex, and prior treatment (yes/no). View Large Table 2 Histopathologic Features and Phenotyping by Flow Cytometry, Categorized by MYD88 Mutation Statusa Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  Characteristic  MYD88L265P  MYD88WT  P Value  Adjusted P Valueb  Pathologic features   % BM infiltration, median (range)  50 (10-100)  20 (10-90)  .004  .0007c   Dutcher bodies  56/128 (44)  6/18 (33)  .46  .56   Hemosiderin  52/128 (41)  3/18 (17)  .07  .32   Mast cells in the aspirate  85/109 (78)  8/18 (44)  .007  .03   Aggregates/interstitial      .74  .74    Interstitial only  45/128 (35)  5/18 (28)        Aggregates only  39/128 (31)  7/18 (39)        Both  44/128 (34)  6/18 (33)       Paratrabecular  58/128 (45)  4/18 (22)  .08  .07   Distribution      .002  .02    Lymphocytes > plasma cells  101/129 (78)  11/19 (58)        Plasma cells > lymphocytes  8/129 (6)  7/19 (37)        Approximately equal  20/129 (16)  1/19 (5)      Phenotyping   Clonal B cells  115/118 (97)  16/17 (94)  .42  .71   Clonal plasma cells  98/113 (87)  15/18 (83)  .71  .78   Plasma cells CD19+ (at least partial)  76/82 (93)  10/12 (83)  .27  .40   Plasma cells CD45+ (at least partial)  81/84 (96)  8/12 (67)  .004  .007   B cells CD5+  19/101 (19)  1/12 (8)  .69  .56   B cells CD10+  3/97 (3)  0/12 (0)  >.99  .42  BM, bone marrow. aValues are presented as number (%) unless otherwise indicated. bAdjusted for age, sex, prior treatment (yes/no), and % BM infiltration, unless specified. cAdjusted for age, sex, and prior treatment (yes/no). View Large Flow cytometric analysis of plasma cells was done in 131 cases and of B cells in 135 cases. Of these, both panels were done in a total of 112 cases. Among the cases with B-cell flow cytometry, a clonal B-cell population was detected in 131 (97%). Among cases with plasma cell flow cytometry, a clonal plasma cell population was identified in 113 (86%). Among cases with both panels performed, clonal B cells and plasma cells were detected in 94 (84%). Of cases with clonal B cells detected by flow cytometry, CD5 was expressed in 20 (18%) of 113 cases and CD10 in three (3%) of 109. Plasma cells in most cases retained expression of CD19 (86/94, 91%) and/or CD45 (89/96, 93%). Of these markers, only CD45 expression by plasma cells was associated with MYD88L265P (adjusted P < .01) (Table 2). Eighty-eight patients had karyotyping and 50 had some type of fluorescent in situ hybridization (FISH) performed on the BM aspirate. Abnormal karyotype (except for isolated –Y) was seen in six (7%) of 88 cases, and an abnormal FISH result was seen in eight (16%) of 50 cases. Results were not specific for a given diagnosis in any case (Supplemental Table 1; all supplemental materials can be found at American Journal of Clinical Pathology online). However, several of the abnormalities seen have been specifically described in LPL, including trisomy 4, 6q deletion, 11q deletion, and gains of chromosome 3.19,20 One case had a t(11;14) identified by karyotype, although breakpoints were not compatible with CCND1-IGH. Other cases had abnormalities of chromosome 12, which have been described in a variety of lymphoid malignancies, albeit not commonly in LPL.21 None of the karyotypic abnormalities were felt to be diagnostic of an underlying myeloid neoplasm or to argue strongly against a diagnosis of LPL. Discussion The diagnosis of LPL is often challenging to pathologists due to morphologic and immunophenotypic overlap with MZL, as well as other LGBLs with plasmacytic differentiation and sometimes multiple myeloma (MM). The discovery of a highly recurrent point mutation, MYD88L265P, in WM has offered an appealing tool for pathologists and clinicians to aid in the differential diagnosis.8-14 However, although MYD88L265P is present in 70% to 100% of patients with WM in various series, at least 10% of MZLs may harbor the mutation, as well,10,14 thereby limiting its utility as a disease-defining mutation. While several studies have evaluated the prevalence of MYD88 mutations in pathologically and clinically well-defined cases of LPL/WM as well as MZL,8,10-14 our study aimed to evaluate the utility of this mutation in identifying a pathologically and clinically distinct group of patients when faced with a LGBLPD in the BM and an IgM paraprotein. In doing so, we sought to determine whether we could better define MYD88L265P-mutated LPL as a distinct pathologic entity. Secondarily, we sought to understand whether cases nonspecifically diagnosed in our practice as LGBLPD represented true LPL or a heterogeneous group. The prevalence of MYD88L265P (87%) in our cohort is similar to that observed in other published cohorts of LPL/WM, supporting our conclusion that most of these cases represent true LPL/WM.13,14,22 Virtually all of the patients studied, with the exception of one patient with IgM cold agglutinin disease (MYD88WT), were, on retrospective review, found to have been considered and treated as having WM clinically. Only 18.9% of these patients had any classic WM symptoms of hyperviscosity, vision changes, headache, bleeding, or neuropathy. Patients diagnosed with LPL (rather than LGBLPD) had a slightly lower median hemoglobin and slightly higher serum viscosity, both of which may be attributable to the slightly higher disease burden among those diagnosed with LPL. Overall, these findings suggest that, despite morphologic, phenotypic, and genetic heterogeneity, these cases seem to all represent a spectrum of true LPL with 158 presenting clinically as WM and one as cold agglutinin disease. In accordance with some prior studies, patients with MYD88L265P had higher disease burden than those with MYD88WT in our cohort.23,24 However, despite initial reports demonstrating adverse prognosis among MYD88WT patients,24 a recent publication from our group, including some overlapping patients with our study as well as others, demonstrated no difference in overall survival between MYD88WT and MYD88L265P patients with WM.25 The current WHO classification acknowledges a large amount of morphologic heterogeneity within the entity of LPL, which was primarily derived from descriptions of this disease process prior to the discovery of the MYD88 mutation.1 Other reports support the assertion that LPL is heterogeneous but indicate that LPL in the BM is identifiable by the presence of paratrabecular aggregates, presence of lymphoplasmacytoid cells, Dutcher bodies, and increased numbers of mast cells.7,26,27 A recent study found these features significantly more often in LPL than in MZL, suggesting their utility in distinguishing the two entities.7 Importantly, this study was done in a cohort of clinically well-defined LPL cases, most of which had MYD88L265P. This raises the question of whether some of the classic morphologic features of LPL are specific to those cases that harbor the mutation, thus supporting MYD88L265P LPL as a unique and identifiable pathologic entity. Our study, however, demonstrates that although some features, including mast cells (78% vs 44%, P = .007), are statistically associated with MYD88L265P, these are present frequently in MYD88WT cases as well. Features such as Dutcher bodies and hemosiderin deposition showed no correlation with MYD88 status in our cohort (Table 2). In addition, there were no significant differences in B-cell immunophenotype between the MYD88WT and MYD88L265P cohorts. Plasma cells in MYD88L265P cases more often showed at least partial CD45 expression compared with MYD88WT. Not surprisingly, most of our cases showed lymphocyte-predominant infiltrates, but a significant minority showed either plasma cell predominance (10%) or approximately equal numbers of plasma cells and lymphocytes (14%), again supporting the morphologic heterogeneity in this disease. In our study, a plasma cell–predominant infiltrate is significantly associated with MYD88WT. Importantly, these seven cases (MYD88WT/plasma cell predominant) did not have lytic bone lesions or other clinical features of MM, showed associated B-cell populations by immunohistochemistry or flow cytometry, and were negative for cyclin D1, making IgM MM unlikely. Although karyotype and FISH are not routinely performed at our institution in the workup of LGBCL in the BM, a subset of our cases did have data available. These results support other studies showing a preponderance of nonspecific abnormalities. Although clearly a useful addition to the routine workup of BM-based LGBLPD, our data suggest that MYD88L265P alone, even within a relatively narrow spectrum of patients with IgM paraproteins and BM-based disease, does not define a single uniform pathologic entity. As previously demonstrated in work by our group, the WHO entity of LPL is not limited to patients with IgM paraproteins and thus is not always synonymous with WM, at least by currently accepted definitions.1,5 The data here further assert that LPL is not limited to patients with MYD88 mutations or those with “typical” pathologic features such as paratrabecular infiltrates and reactive mast cells. There is no doubt that the discovery of MYD88L265P mutations in LPL has helped to clarify both diagnostic and therapeutic issues within the realm of this complex disease. However, as with any new diagnostic test, it has also raised many questions regarding whether histopathologic, molecular, or clinical features should take precedence in disease classification. As our data have shown, even when faced with difficult-to-classify, BM-based lymphoplasmacytic infiltrates, MYD88L265P does not define a distinct phenotypic or morphologic entity. Overall, our findings suggest that despite pathologic heterogeneity, these cases seem to all represent a spectrum of true LPL/WM. What our data most support is a clinicopathologic approach to the diagnosis of LPL and recognition that this intriguing disease may manifest with varying morphologies, phenotypes, and molecular features. From the pathologists’ perspective, our data support a more inclusive use of the WHO-defined diagnosis of LPL. Finally, while MYD88L265P itself may not define LPL as an entity, it clearly represents an exciting biomarker for therapeutic decisions and predicting response to certain therapies. 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American Journal of Clinical PathologyOxford University Press

Published: Jun 2, 2018

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