Lymphoplasmacytic Lymphoma Patients Research Articles

What differences in outcomes exist in Hispanic patients being treated for lymphoplasmacytic lymphoma and waldenstrom macroglobulinemia?

e19541 Background: Lymphoplasmacytic Lymphoma (LPL) and Waldenstrom Macroglobulinemia (WM) are both rare diseases with little known about their longterm outcomes in ethnic minorities. Overall, the 5- and 10-year survival of this disease has improved over the last 60 years (Blood, PMID: 24948623), but few studies have explored outcomes in Hispanic (HI) patients. We analyzed demographics, treatment patterns and outcomes in patients diagnosed with LPL and WM comparing HI and Non-Hispanics (NH) in the US. Methods: Data was analyzed on LPL and WM patients in the US reported to the Surveillance, Epidemiology, and End Results (SEER) analyzing patient characteristics, age-adjusted incidence rate, and survival rate among HI and NH patients. Results: HI were diagnosed at a younger median age on both LPL and WM cohorts compared to NH (p <0.001). There was a higher percentage of HI patients diagnosed with LPL and WM before the age of 60 compared to that of NH (p <0.001). Regarding race of patients with LPL, most of both HI and NH patients identified as whites (94% vs 86%), followed by blacks (2% vs 6%). Regarding stage, a majority of both HI (50%) and NH (60%) patients diagnosed with LPL were Stage IV. In terms of radiation, 7% of HI patients with LPL received it, compared to 4% of NH patients (p=0.027). On survival analysis for LPL patients, the survival probability at 2, 5 and 10 years (y) of HI vs NH were similar. There was not an OS difference favoring HI/NH (p=0.49). On survival analysis for WM patients, the survival probability at 2, 5 and 10 y of HI vs NH were similar. The median survival time for patients with WM was similar between HI and NH patients and there was not OS difference favoring HI vs NH (p=0.16). On multivariate analysis for LPL and WM, those diagnosed over the age of 80 had worse OS compared to those younger than 60 (95% CI; 4.8-8.1 and 2-3.4, respectively). No OS benefit when adjusting for stage was observed for either LPL or WM. Conclusions: In LPL and WM patients HI patients were diagnosed at younger ages compared to NH for both LPL and WM, and more likely to be diagnosed before the age of 60. The median survival rates were similar between HI and NH patients diagnosed with both LPL and WM. These findings may be confounded by earlier diagnosis in HI patients; however, further studies are needed exploring biological ethnic differences to explain early diagnosis in LPL and WM among HI.

Waldenström Macroglobulinemia and Non-IgM-Type Lymphoplasmacytic Lymphoma Are Genetically Similar

Introduction: Waldenström macroglobulinemia (WM) represents a subset of lymphoplasmacytic lymphoma (LPL) with the immunoglobulin (Ig)M paraprotein. MYD88 L265P and CXCR4 mutations are common mutations in WM patients, and mutations in ARID1A and KMT2D (MLL2) have also been reported. However, little information has been accumulated on genetic changes in LPL with other paraproteins like IgG. Methods: We therefore aimed to evaluate genetic differences between WM and LPL with non-IgM paraprotein (non-IgM-type LPL) using targeted next-generation sequencing (NGS) in 20 Japanese patients (10 with WM, 10 with non-IgM-type LPL). Results: Mutations were detected in ARID1A (10%), CXCR4 (20%), MYD88 (90%), and KMT2D (0%) for WM patients and in ARID1A (10%), CXCR4 (20%), MYD88 (70%), and KMT2D (10%) for non-IgM-type LPL patients. No significant differences were identified. No mutations were detected in NOTCH2, PRDM1, CD274 (PD-L1), PDCD1LG2 (PD-L2), RAG2, MYBBP1A, TP53, or CD79B. Discussion: Mutant allele frequency in MYD88 L265P did not differ significantly between WM and non-IgM-type LPL. Most mutations detected by NGS were subclonal following MYD88 L265P, although one non-IgM-type LPL patient harbored only CXCR4 S338X mutation. Our NGS analyses reveal genetic characteristics in LPL patients and suggest genetic similarities between these two subsets of LPL, WM and non-IgM-type.

Clinical and biological characteristics of non-IgM lymphoplasmacytic lymphoma

目的探索非IgM型淋巴浆细胞淋巴瘤（LPL）患者的临床及生物学特征。方法回顾性收集中国医学科学院血液病医院1993年7月至2020年8月收治的340例LPL患者的临床资料，其中23例为非IgM型LPL组，317例为华氏巨球蛋白血症（WM）组。比较两组患者的临床及生物学特征。结果23例非IgM型LPL患者中，2例分泌单克隆性IgA，14例分泌单克隆性IgG，7例不分泌单克隆性免疫球蛋白。非IgM型LPL和WM患者中位年龄均为62（35～81）岁。与WM组患者相比，非IgM型LPL组患者女性（56.5％对27.3％，P＝0.007）、脾大（60.1％对43.8％，P＝0.100）、结外侵犯（21.7％对12.3％，P＝0.672）比例更高。非IgM型LPL组18例患者进行了MYD88基因相关检测，阳性率55.6％。非IgM型LPL组17例患者接受了治疗，启动治疗的患者比例与WM组患者相当（94.4％对92.7％，P＝0.488）。非IgM型LPL组16例患者进行了疗效评价，一线治疗总体缓解率87.5％，中位随访时间33.9（3.5～125.1）个月，总体中位无进展生存（PFS）、总生存（OS）时间未达到，3年PFS率和OS率分别为71.4％和68.9％。WM组中位PFS、OS时间分别为66.2个月和78.1个月。两组PFS、OS的差异均无统计学意义（P值分别为0.340、0.544）。结论非IgM型LPL与WM患者的临床及生物学特征相似，但非IgM型LPL组女性、结外受累比例更高。非IgM型LPL患者的生存及预后与WM患者相似。

Lymphoplasmacytic Lymphoma with Only Lambda Light Chain Monoclonal Paraprotein Expression.

IntroductionLymphoplasmacytic lymphoma (LPL) is a rare low-grade B-cell neoplasm that accounts for approximately 2% of all haematological malignancies. Most patients have the clinical syndrome of Waldenstrom macroglobulinemia (WM), which is defined as LPL with an associated immunoglobulin M (IgM) serum monoclonal protein. Roughly 5% of LPL patients secrete non-IgM paraproteins (e.g., IgG, IgA, kappa, lambda) or are non-secretory.Case descriptionWe report the case of a 41-year-old woman who was diagnosed with non-IgM LPL with lambda light chain monoclonal paraprotein production and normal serum immunoglobulin levels. The MYD88 L265P mutation was detected on fluorescence in-situ hybridization (FISH) analysis of the bone marrow. The patient underwent treatment with a combination of ibrutinib and rituximab. There was an initial response but she died 8 months after diagnosis.DiscussionNon-IgM LPL poses diagnostic and therapeutic challenges to clinicians as it is an exceptionally rare malignancy with a heterogeneous clinicopathological presentation and scarce literature. Among non-IgM LPL cases, those with lambda light chain production are even more rare. To the best of our knowledge, none have been reported to date. The addition of MYD88 L265P testing to the diagnostic armamentarium of non-IgM LPL cases is advisable for potential therapeutic reasons.LEARNING POINTSOur case report and literature review provide insight into non-IgM lymphoplasmacytic lymphoma (LPL), an extremely rare malignancy.Our case report highlights the importance of the need for new treatments for non-IgM LPL.

Clinical characteristic and outcome of lymphoplasmacytic lymphoma of non-Waldenstrom macroglobulinemia type: A Swedish lymphoma registry study.

Lymphoplasmacytic lymphoma (LPL) not fulfilling the WHO diagnostic criteria (2017) for Waldenstrom's macroglobulinemia (WM) (named non-WM LPL) is a rare disease and only a few systematic studies have been published. Here, we present a population-based study of non-WM LPL focusing on diagnostic difficulties, patient characteristics, and outcome. From 1511 patients included in the Swedish Lymphoma Registry 1 Jan 2000 - 31 Dec 2014 with a diagnosis of WM/LPL, we could confirm the diagnosis of non-WM LP in only 33 patients. The median age at diagnosis was 69 years. A paraprotein was found in most (IgG in 54%, IgA in 15%) and 12% of the cases were non-secretory. Compared with the WM patients, the non-WM LPL patients were younger, had more adverse prognostic factors such as elevated LDH, anaemia, and lymphocytosis at diagnosis. In addition, the non-WM LPL patients more often were symptomatic and received treatment at diagnosis. The overall survival (OS) did not significantly differ between the non-WM LPL and WM groups (P= 0.247), with a median survival time of 71 and 96 months, respectively. To conclude, we found differences in clinical features between WM and non-WM LPL, but no difference in survival.

Lymphoplasmacytic Lymphoma Not Associated with an IgM Monoclonal Paraprotein Has Distinctive Clinical and Biological Features and a Similar Outcome As Compared with Waldenstrom's Macroglobulinemia: Multicentric Study of the Rete Ematologica Lombarda

Background. Lymphoplasmacytic lymphoma (LPL) is a low-grade B-cell neoplasm exhibiting a spectrum of B-cell differentiation ranging from small lymphocytes to plasma cells. In the majority of cases LPL is associated with an immunoglobulin M (IgM) paraprotein in the serum, corresponding to the clinico-pathologic entity of Waldenström's Macroglobulinemia (WM). In a minority of patients with LPL the paraprotein is absent or is of IgG or IgA isotype. These cases are very rare and their relationship with WM still unclear.Aim of the study. The aim of this study was to analyze clinical and biological features, treatment characteristics and outcome of patients with LPL not associated with an IgM paraprotein and to compare these cases with a control group of WM patients.Patients and methods. The study was conducted within the Rete Ematologica Lombarda (REL) and approved by the Ethic Committee of the six participating centers. Clinical characteristics, treatment and outcome of patients were retrieved from clinical archives. MYD88 and CXCR4 mutation status were evaluated by means of allele-specific PCR for MYD88 (L265P) and Sanger sequencing respectively. Five patients from the coordinating center were studied also with next-generation sequencing. DNA was extracted from bone marrow total mononuclear cells (n=20), bone marrow biopsy (n=8), lymph node (n=6), spleen (n=3) or extranodal sites (n=3).Results. We identified 45 cases of LPL not associated with a serum IgM paraprotein for whom clinical data were available. Twenty-nine patients out of 45 (64%) had a serum IgG (n= 22) or IgA (n=7) paraprotein, whereas serum immunofixation was negative in 16 cases (36%). Patients characteristics are reported in Table 1. Forty samples from 36 patients were available for molecular analyses. MYD88 and CXCR4 mutation status before treatment were evaluable in 34 and 35 cases respectively. Six patients of 34 (18%) were found to harbor the MYD88 L265P and one the S219C mutation. Five patients of 35 (14%) had a CXCR4 and three of them were MYD88-wild type. The median follow-up of LPL patients was 40.1 months (range 0.5-207). Thirty-five patients (78%) were treated after a median time of 2.5 months from diagnosis (range: 0-49). Immunochemotherapy represented the first line therapy in 25 of 35 patients (71%). Chemotherapy consisted of alkylating-containing regimens in 12/35 patients (34%), CHOP or CHOP-like regimens in 11 (31%), bendamustine in 9 (26%), fludarabine-based regimens in 2 (6%), other treatment in 1 patient (3%). Fifteen of 35 patients (43%) progressed after treatment. Progression-free survival (PFS) after first-line treatment was 55% at 5 years. Overall survival (OS) was 81% at 5 years.As compared with 142 WM controls with a median follow-up of 44.9 months (0-180), LPL patients were more frequently females (58% versus 39%, P=0.038), had more frequently lymphadenopathies (56% versus 15%, P<0.001), spleen enlargement (22% versus 8%, P=0.015) or extranodal involvement (20% versus 8%, P=0.05), while bone marrow infiltration was less common (84% versus 99%, P<0.001). The prevalence of the MYD88 (L265P) mutation was significantly lower in LPL as compared with WM patients (18% versus 91%, P<0.001) while the prevalence of CXCR4 mutations was similar in the two groups (14% versus 17%, P=0.794). A higher proportion of LPL patients were treated as compared to the control group (78% versus 50%, P=0.001). LPL received more frequently anthracycline-containing regimens (31% versus 4%, P<0.001) and less frequently alkylating-containing regimens (34% versus 65%, P=0.004) than WM patients. The use of bendamustine-containing regimens was not significantly different between the two groups (17% versus 26%, P=0.308). As compared with WM, LPL patients had a similar PFS (P=0.26) (Figure 1) and OS (P=0.62) (Figure 2).Conclusions. LPL not associated with an IgM monoclonal protein displays distinctive clinical and biological characteristics, in particular a higher prevalence of extramedullary disease and a lower rate of MYD88 mutations as compared to WM. The outcome of patients with LPL is similar to WM in terms of progression-free and overall survival. [Display omitted] DisclosuresVarettoni:Jannsen: Consultancy, Other: travel expenses. Tedeschi:Janssen: Consultancy, Other: travel expenses; AbbVie: Consultancy; Gilead: Consultancy, Other: travel expenses. Arcaini:Sandoz: Consultancy; Gilead: Research Funding; Pfizer: Membership on an entity's Board of Directors or advisory committees; Celgene: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bayer: Membership on an entity's Board of Directors or advisory committees; Roche: Consultancy, Membership on an entity's Board of Directors or advisory committees.

PD-1 Blockade with Pembrolizumab in Relapsed Low Grade Non-Hodgkin Lymphoma

Background: The natural history of low grade non-Hodgkin lymphoma (NHL) typically includes multiple relapses and short responses to therapy after each relapse. The development of novel therapies for NHL patients with multiple relapses is therefore clinically needed. In previous studies, we demonstrated the presence of PD-1 positive exhausted T cells in the nodal tissue of low grade NHL patients. We therefore hypothesized that blocking PD-1 will activate tumor infiltrating T cells to induce effector cell-mediated cytotoxicity. To test this hypothesis, we performed a phase 2 trial (MC1485) to test the efficacy of the PD-1 blocking antibody pembrolizumab in relapsed/refractory low grade NHL.Methods: Relapsed/refractory low grade NHL including follicular lymphoma (FL), marginal zone lymphoma (MZL) and lymphoplasmacytic lymphoma (LPL) who had ECOG performance 0-2 and normal organ function were included in arm B of the MC1485 trial. Patients with prior allogeneic stem cell transplant were excluded. Pembrolizumab 200 mg was administered intravenously every 3 weeks until progression, excessive toxicity, or completion of 2 years of therapy. The primary end point of this study was overall response rate. Tumor T cell infiltration and expression of PD-1 and PD-L1 were assessed with standard immunohistochemical (IHC) staining.Results: 23 patients including 18 FL, 2 MZL and 3 LPL were enrolled and the median age was 67 years (47-82). The median number of prior therapies was 4 (1-20). 17 out of 23 (74%) patients had received more than 2 prior therapies. 100% of patients had received an anti-CD20 antibody and 22/23 (96%) had received chemoimmunotherapy. 6 (26%) had a prior autologous stem cell transplant. 10 (43%) had received prior novel agents including lenalidomide or idelalisib or other experimental therapies. The median number of pembrolizumab doses received was 4 (range, 1 to 16), administered over a median treatment duration of 13.9 weeks (range, 2.6 to 50.8). Drug-related adverse events (AE) occurred in 17 patients (74%) with the common ones being thrombocytopenia in 6 (26%), neutropenia in 5 (22%), anemia in 5 (22%), nausea in 5 (22%), diarrhea in 4 (17%), lymphocyte decrease in 4 (17%), nausea, vomiting and cough each in 3 (13%), and dyspnea in 2 (8%) patients. Drug-related grade 3 or above AE occurred in 9 (39%) patients with most common ones being thrombocytopenia (3, 13%), anemia (3, 13%), neutropenia (2, 8%) and dyspnea (2, 8%). One possibly related death occurred in a patient who had baseline obstructive sleep apnea and developed respiratory failure after two doses of therapy without clear cause. He eventually refused to receive further supportive care. Based on the investigator assessment, 2 FL patients had partial responses (PR, 9%). One LPL patient had minor response using LPL/Waldenstrom macroglobulinemia response criteria (MR, 4%). 11 patients (48%) had stable disease (SD). These two responders had durations of response of 5.5 and 4.9 months respectively (Figure). One MZL patient had ~40% tumor reduction with ongoing therapy. 7 patients (30%) had progressive disease. With a median follow-up being 6.5 months (1.6-24.7), the median progression free survival is 3.4 months (95% CI: 2.1-5.7). The median overall survival has not been reached.Conclusion: Pembrolizumab had an acceptable safety profile in low grade NHL patients. In selected NHL patients, pembrolizumab has activity as single-agent therapy. Biomarker evaluation is needed to select patients who can benefit from this therapy. Further combination therapy will be required to improve efficacy. [Display omitted] DisclosuresDing:Merck: Research Funding. Witzig:Celgene: Membership on an entity's Board of Directors or advisory committees, Research Funding; Acerta: Research Funding; Kura: Research Funding; Novartis: Research Funding. Nowakowski:Bayer: Consultancy, Research Funding; Abbvie: Consultancy; celgene: Consultancy, Research Funding; Morphosys: Consultancy, Research Funding; Genetech: Consultancy, Research Funding; pharmacyclics: Consultancy; Nanostring: Research Funding. Reeder:Millennium: Research Funding; Celgene: Research Funding; Affimed: Research Funding; Bristol-Myers Squibb: Research Funding; Novartis: Research Funding. Kapoor:Takeda, Celgene and Amgen: Research Funding. Ansell:Bristol-Myers Squibb: Research Funding; Affimed: Research Funding; Merck: Research Funding; Celldex: Research Funding; Seattle Genetics: Research Funding.

Characterization of Chromosome 6q Deletions in MYD88 Mutant and Wild-Type Waldenström's Macroglobulinemia

Introduction: MYD88 L265P mutations have been described in more than 90% of Waldenström's Macroglobulinemia (WM) patients. MYD88 wild-type (wt) WM represent less than 10% of WM, and associate with shorter overall survival versus MYD88 mutated (mut) WM, as well as with lack of major responses and inferior progression-free survival to ibrutinib. Mutated MYD88 triggers WM cells growth and survival through NF-kB activation via IRAK1/4, BTK and HCK transactivation. CXCR4 mutations promote pro-survival AKT and ERK1/2 signaling and are associated with delayed response to ibrutinib. Deletions involving the long arm of chromosome 6 (Del6q) are highly recurrent in various B-cell malignancies, particularly those that are driven by mutated MYD88, and have been observed in up to half of WM patients. Chr6q genomic loss in WM affect important modulators of NFkB (TNFAIP3, HIVEP2), BCL2 protein family (BCLAF1), apoptosis and IGF1/PI3K/AKT signaling (FOXO3), and BTK (IBTK). The frequency and impact of Del6q on the different patient genotype remains to be clarified.Patients and Methods: Based on location and known regulation of key pathways in WM, copy number alterations (CNA) and gene expression level were assessed for IBTK, FOXO3, BCLAF1, TNFAIP3 and HIVEP2 genes, along with CXCR4 transcriptional levels. DNA and RNA from CD19+ sorted BM lymphoplasmacytic cells from 32 untreated WM and one IgM and IgG-secreting lymphoplasmacytic lymphoma patients were analyzed. Paired CD19-depleted peripheral-blood mononuclear cells (PBMCs) were used as germline controls. Paired CD19+ and CD19- PBMCs from 6 healthy donors were analyzed to rule out possible B-cell specific findings. CNA were measured in quadruplicate and gene expression in triplicate using TaqMan real-time polymerase chain reaction (RT-PCR) assays. Deletions affecting less than 20% of WM cells were considered to be beneath the RT-PCR detection threshold. The cohort included 33 patients (21 males, 12 females), with a median age of 62 (range 35-91) years, BM involvement of 60% (range 2.5-90%), serum IgM levels of 3010 (range 257-6910) mg/dl, and hemoglobin of 10.9 (range 8.4-14.4) g/dl. MYD88 L265P mutations were detectable in 25 (76%) patients, 11 of whom (44%) also carried CXCR4WHIM mutations. Eight patients (24%) were MYD88 and CXCR4 wt.Results: Somatic Del6q were observed in 4/8 (50%) and in 20/25 (80%) patients in the MYD88 wt and mut population, respectively (p=NS). In contrast, no CNA were present in the HDs CD19+/CD19- compartments. IBTK remained intact in all 8 MYD88 wt WM, whereas resulted deleted in 13/25 (52%) MYD88 mut patients (p=0.01). BCLAF1 deletion frequency was significantly different among the two subgroups, with only 2/8 deleted MYD88wt (25%) and 19/25 (76%) deleted MYD88 mut patients (p=0.02; Figure 1). Eight MYD88 mut WM (32%) showed contiguous fully clonal Del6q, that spanned all studied genes in 7/8 (87.5%) cases; in contrast, contiguous subclonal deletions occurred only in 3/12 (25%) cases (p=0.02). All 8 fully clonal Del6q patients were CXCR4 wt; in contrast, CXCR4 mut were detected in 9/12 (75%) patients with subclonal Del6q (p=0.001; Figure 2). Unlike the MYD88 mut population, all 4 MYD88 wt patients presented with non-contiguous subclonal deletions (p=NS). Fully clonal deletions of IBTK, BCLAF1 and HIVEP2 significantly reduced the respective gene transcriptional levels (p= 0.03, 0.01 and 0.01, respectively), while CNA in TNFAIP3 and FOXO3 did not affect their expression. The MYD88 wt patients showed significant decreases in the transcriptional levels of BCLAF1 related to gene subclonal deletions, and reduced CXCR4 levels compared to MYD88 mut WM (p=0.02 and p=0.02, respectively).Conclusions: Our findings demonstrate that gene loss of IBTK, FOXO3, BCLAF1, TNFAIP3 and HIVEP2 occurs in most WM patients, but at a lower frequency in MYD88 wt. Unlike the MYD88 mut patients, those MYD88 wt showed no contiguous fully clonal deletions. The latter correlate with CXCR4 wt and reduce only IBTK, BCLAF1 and HIVEP2 levels, suggesting that regulatory mechanisms may compensate for FOXO3 and TNFAIP3 gene loss. Significant variation in IBTK and BCLAF1 deletion rate suggest that BCR and BCL2 signaling may play a different role in MYD88 mut and wt WM. The findings provide valuable insights into MYD88 wt WM pathogenesis, and may be relevant to understanding therapeutic outcome with agents that target MYD88, CXCR4, BTK and BCL2. [Display omitted] DisclosuresCastillo:Millennium: Research Funding; Abbvie: Research Funding; Janssen: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding. Arcaini:Pfizer, Celgene, Bayer, Roche: Membership on an entity's Board of Directors or advisory committees; Celgene, Roche, Sandoz: Consultancy; Gilead: Research Funding. Varettoni:Jannsen: Consultancy, Other: travel expenses. Treon:Pharmacyclics: Consultancy, Research Funding.

Lymphoplasmacytic lymphoma patient with biclonal IgG and IgA: a case report

Lymphoplasmacytic lymphoma patient with biclonal IgG and IgA: a case report

Level of Coagulation Function Indexes and Its Clinical Significance in Patients with Lymphoplasmacytic Lymphoma

To analyze the level of coagulation function indexes in patients with lymphoplasmacytic lymphoma (LPL) and its clinical significance. The clinical data of 32 patients with initial LPL (LPL group) and physical examination data of 25 healthy persons (control group) who underwent physical examination in our hospital during the same period were collected. The differences of platelet (Plt), D-Dimer (D-D), fibrinogen (Fib), thrombin time (TT), prothrombin time (PT) and activated partial thrombin time (APTT) between the two groups were compared. The Plt count in LPL group [ (137.06±40.14)×109/L] was significantly lower than that in control group [ (215.07±33.25)×109/L], D-D [ (1.01±0.16) mg/L, PT [ (13.01±1.37) s] and APTT [ (40.96±7.24) s] in LPL group were significantly higher than those in control group [ (0.37±0.09) mg/L, (11.96±0.87) s, (25.07±5.13) s] (P＜0.01); there was no significant difference in TT and Fib levels between the two groups (P＞0.05). There was no significant difference in Plt, D-D, Fib, AP, TT and APTT among LPL patients secreting different types of immunoglobulin (Ig) (P＞0.05). After treatment, the coagulation function of LPL patients returned to normal, and no death cases occurred due to hemorrhage or thrombosis. LPL patients have hypercoagulable state blood and abnormal coagulation function, but which not closely relates to with the type of Ig secreted by patients.

MYD88 L265P mutation analysis is a useful diagnostic adjunct for lymphoplasmacytic lymphoma with pleural effusion.

Lymphoplasmacytic lymphoma (LPL) is a marrow-based lymphoma, rarely involving extramedullary sites, particularly the pleural cavities. The distinction of lymphomatous pleural effusion (PE) in LPL patients from benign effusion is challenging. We conducted this study to examine whether MYD88 L265P mutation analysis is useful in distinguishing benign from lymphomatous PE in four patients with LPL, in which the initial marrow specimens were all positive for MYD88 mutation. In one case each with plasma cell- or lymphocyte-predominant PE, MYD88 mutation was positive, confirming lymphomatous effusion. The other lymphocyte-predominant PE was negative for MYD88 mutation, but was clonally related to a previous nodal biopsy and this PE was also considered to have LPL involvement. The fourth case developed large B-cell lymphoma in the PE 30 months later. The PE specimen was negative for MYD88 mutation but was clonally related to the diagnostic marrow tissue, indicating large cell transformation. Four cases of small lymphocyte-predominant benign PE from patients without history of lymphoma were examined and were all negative for MYD88 L265P mutation. In conclusion, in this small case series we showed that MYD88 L265P mutation analysis could serve as a useful adjunct in distinguishing benign from lymphomatous PE in patients with LPL.

Clinical and molecular characteristics of lymphoplasmacytic lymphoma not associated with an IgM monoclonal protein: A multicentric study of the Rete Ematologica Lombarda (REL) network

Lymphoplasmacytic lymphoma (LPL) is usually associated with a serum IgM paraprotein, corresponding to Waldenström's Macroglobulinemia (WM). Cases presenting with IgG or IgA, or without a monoclonal protein are extremely rare. We analyzed clinical characteristics, frontline treatment, and the outcome of 45 patients with non-IgM LPL, and compared them with a control group of WM patients. The median age was similar, with significantly higher prevalence of females in non-IgM LPL, than in WM patients (60% vs 39%, P = .016). Patients with non-IgM LPL more frequently presented with lymphadenopathies (53% vs 15%, P < .001), splenomegaly (22% vs 8%, P = .015) or extranodal involvement (20% vs 8%, P = .05). In non-IgM LPL a serum monoclonal protein and bone marrow infiltration were less common than in WM patients (69% and 84% of cases respectively, P < .001 for both comparisons). The MYD88 (L265P) mutation was found in 8/19 patients using allele-specific polymerase chain reaction. A CXCR4 mutation was found in 4/17 cases using Sanger. In 16 patients we performed targeted next-generation sequencing of genes MYD88, CXCR4, ARID1-A, KMT2D, NOTCH2, TP53, PRDM1, CD79B, TRAF3, MYBBP1A, TNFAIP3. Seven patients (44%) had a MYD88 mutation (S219C in one), four (25%) a CXCR4 mutation, three (19%) a KMT2D mutation, one (6%) a TP53 mutation and one (6%) a TRAF3 mutation. With a median follow-up of 55.7 months, 36 non-IgM LPL patients (80%) were treated. Non-IgM LPL patients received more frequently anthracycline-containing regimens, as compared with WM patients, who mainly received alkylating-based therapies. Five-year overall survival (OS) was 84%, similar to that of WM patients.

Changes of blood coagulation indicators in patients with lymphoplasmacytic lymphoma and their clinical significances

Objective To observe the alteration and clinical significances of blood coagulation indicators in patients with lymphoplasmacytic lymphoma (LPL). Methods Twenty patients who were newly diagnosed LPL in the First People's Hospital of Changzhou from January 2008 to October 2017 and twenty healthy controls were studied. The patients were treated by chemotherapy, plasma exchange, supplement of coagulation factor or other supportive therapy. The parameters of prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), thrombin time (TT), D-dimer (D-D), and platelet count (Plt) were detected in LPL group and healthy controls. Results The levels of PT and APTT in LPL group were dramatically higher than those in control group [(12.9±1.2) s vs. (11.6±0.9) s, (41.7±9.8) s vs. (24.7±2.9) s], and the level of Plt in LPL group was lower than that in control group [112×109/L (3×109/L-379×109/L) vs. 210×109/L (170×109/L-271×109/L)], and the differences were statistically significant (all P 0.05). There were no statistical differences in PT, APTT, FIB, TT, D-D and Plt levels among LPL patients with different types of immunoglobins (all P > 0.05). After treatment, all the coagulation abnormalities got relieved and no patient died of hemorrhage or thrombosis. Conclusions The LPL patients have coagulation disorders and hypercoagulability, and this is independent of the type of immunoglobulin. Clinical attention should be paid to monitoring coagulation indicators to prevent the occurrence of adverse reactions. Key words: Lymphoplasmacytic lymphoma; Blood coagulation factors; Platelets

Phase I study of an active immunotherapy for asymptomatic phase Lymphoplasmacytic lymphoma with DNA vaccines encoding antigen-chemokine fusion: study protocol

BackgroundThere is now a renewed interest in cancer vaccines. Patients responding to immune checkpoint blockade usually bear tumors that are heavily infiltrated by T cells and express a high load of neoantigens, indicating that the immune system is involved in the therapeutic effect of these agents; this finding strongly supports the use of cancer vaccine strategies. Lymphoplasmacytic lymphoma (LPL) is a low grade, incurable disease featuring an abnormal proliferation of Immunoglobulin (Ig)-producing malignant cells. Asymptomatic patients are currently managed by a “watchful waiting” approach, as available therapies provide no survival advantage if started before symptoms develop. Idiotypic determinants of a lymphoma surface Ig, formed by the interaction of the variable regions of heavy and light chains, can be used as a tumor-specific marker and effective vaccination using idiotypes was demonstrated in a positive controlled phase III trial.MethodsThese variable region genes can be cloned and used as a DNA vaccine, a delivery system holding tremendous potential for streamlining vaccine production. To increase vaccination potency, we are targeting antigen-presenting cells (APCs) by fusing the antigen with a sequence encoding a chemokine (MIP-3α), which binds an endocytic surface receptor on APCs. Asymptomatic phase LPL is an excellent model to test our vaccine since patients have not received chemotherapeutics that interfere with innate immune function and have low tumor burden. We are evaluating the safety of this next-generation DNA vaccine in a first-in-human clinical trial currently enrolling asymptomatic LPL patients. To elucidate the mode of action of this vaccine, we will assess its ability to generate tumor-specific immune responses and examine changes in the immune profile of both the peripheral blood and bone marrow.DiscussionThis vaccine could shift the current paradigm of clinical management for patients with asymptomatic LPL and inform development of other personalized approaches.Trial registrationClinicalTrials.gov identifier NCT01209871; registered on September 24, 2010.

Detection of MYD88 L265P in patients with lymphoplasmacytic lymphoma/Waldenstrom macroglobulinemia and other B-cell non-Hodgkin lymphomas.

BackgroundRecent studies have identified a high prevalence of the MYD88 L265P mutation in lymphoplasmacytic lymphoma (LPL)/Waldenstrom macroglobulinemia (WM) cases, whereas low frequencies have been observed in other B cell non-Hodgkin lymphomas (NHLs).MethodsWe evaluated the sensitivity of the mutant enrichment 3'-modified oligonucleotide (MEMO)-PCR technique, a new detection method. We examined the MYD88 L265P mutation in a series of Korean patients with LPL/WM and other B cell NHLs in bone marrow aspirates, using the MEMO-PCR technique.ResultsThe sensitivity of MEMO-PCR was estimated to be approximately 10-16.7%. MYD88 L265P was detected in 21 of 28 LPL cases (75%) and only three of 69 B cell NHL cases (4.3%).ConclusionAlthough MEMO-PCR had relatively low sensitivity, we confirmed the high prevalence of the MYD88 L265P mutation in Korean LPL patients. Our study suggests the diagnostic value of MYD88 L265P for differentiating B-cell NHLs.

MYD88 L265P and CXCR4 mutations in lymphoplasmacytic lymphoma identify cases with high disease activity.

Recurrent mutations in MYD88 have been identified in >90% of lymphoplasmacytic lymphoma (LPL). Recently, WHIM (warts, hypogammaglobulinaemia, infections, myelokathexis) syndrome-like mutations in CXCR4 have been described in 28% of LPL cases, and seem to impact clinical presentation and response to therapy. We investigated the presence of the MYD88 L265P mutation in 90 decalcified, formalin-fixed, paraffin-embedded (FFPE) bone marrow (BM) biopsies, including 51 cases of LPL, 14 cases of B-cell chronic lymphocytic leukaemia (CLL), 13 cases of marginal zone lymphoma (MZL) and 12 normal controls. In addition, the C-terminal domain of CXCR4 was sequenced in LPL cases. MYD88 L265P was found in 49/51 (96%) LPL cases and in 1/13 (7·6%) MZL (splenic type), whereas all CLL samples remained negative. The two MYD88 wild type LPL cases were associated with cold agglutinin disease. Mutations in CXCR4 were detected in 17/47 (36·2%) LPL cases, which showed a higher extent of BM infiltration and lower leucocyte counts (P=0·02), haemoglobin (P=0·05) and platelet counts (P=0·01). In conclusion the detection of MYD88 L265P mutation in FFPE samples is reliable and useful for subtyping small B-cell lymphomas in BM biopsies. In addition, the presence of CXCR4 mutations identifies a subgroup of LPL patients with higher disease activity.

MYD88 L265P Mutation Detection: Analysis of Flow Cytometry Sorted Plasma and Lymphoid Cell Clones Improves Sensitivity and Specificity for WM/LPL Diagnosis

Background:Waldenström’s macroglobulinemia (WM) and lymphoplasmacytic lymphoma (LPL) are lymphoproliferative disorders with bone marrow infiltration by clonal lymphoplasmacytic cells (Treon et al., 2003, 2005). The somatic point mutation L265P in the myeloid differentiation primary response gene 88 (MYD88) has been reported in more than 90% of WM patients (Treon et al., 2012). Therefore MYD88 mutation analysis has been implemented in clinical practice to support the diagnosis of LPL/WM.After the implementation of MYD88 L265P assays with increased detection sensitivity, a substantial portion of patients with IgM monoclonal gammopathy of undetermined significance (IgM-MGUS) was also reported MYD88 L265P positive. Splenic marginal zone lymphoma (SMZL), B-cell chronic lymphoproliferative disorders (B-CLPD) and diffused large B-cell lymphoma (DLBCL) have been found positive with much lower incidence rates (Varettoni et al., 2013; Xu et al., 2013).Hence the remaining need to differentiate WM/LPL from other lymphoproliferative disorders with co-occurring plasma cells with high confidence.Patients and Methods:In this study, flow cytometric cell sorting was utilized to isolate clonal plasma and B lymphoid cell fractions as previously described (Zehentner et al., 2011). 69 patient specimen fractions with a clinical history of WM /LPL, multiple myeloma, CLL and lymphoma were analyzed for MYD88 L265P mutation using Sanger sequencing. Furthermore, the Biomed-2 primer sets for the immunoglobulin heavy (IgH) and/or the immunoglobulin kappa light chain region (IgK) were used to compare B cell clonality profiles in the lymphoid versus plasma cell compartments.Results:MYD88 L265P mutation was detected in all specimens with confirmed Waldenström’s macroglobulinemia (17/17, 100%). Of these 16/17 (94%) revealed MYD88 L265P as well as identical monoclonality profile by gene rearrangement analysis in both the plasma and the B lymphoid cell collections. In 47% (8/17), the mutation was only detected in the plasma and B cell fractions, but not in the whole bone marrow specimens.21 patient specimens with a known clinical history of LPL and co-occurring clonal plasma cells were tested. 9 of 21 (43%) were categorized with identical B cell clonality profile when comparing plasma and B lymphoid cells; whereas 12 (57%) had unrelated clonality profiles. 7 of the 9 (78%) specimens in the identical clonality group tested positive for MYD88 L265P in both the plasma and B lymphoid clone. None of the unrelated clonality group specimens carried the mutation in both cell fractions; for 7/12 (58%) MYD88 L265P was found in either the plasma (2) or the B-cell fraction (5) whereas 5/12 (42%) tested negative.11 bone marrow aspirate specimen with known presence of lymphoma (including splenic marginal zone (SMZL), mantle cell and marginal zone) were analyzed. 10/11 (90%) tested negative for MYD88 L265P, with the exception of one SMZL specimen. Furthermore, 12 known myeloma, 5 CLL and 3 healthy donor specimens were analyzed, all tested negative.Conclusions:In this study, we developed and tested a novel approach to assess MYD88 L265P mutation status in order to assist WM/LPL diagnosis. Flow cytometric cell sorting for clonal plasma and B cell populations was combined with molecular analysis. Subsequently, MYD88 L265P mutation as well as B-cell clonality profile was compared in both cell fractions.Our study postulates a significant improvement in sensitivity and most importantly specificity when applying MYD88 L265P mutation status in combination with cell sorting for WM/LPL diagnostic decisions. 47% WM patients (8/17) and 44% LPL patients (4/9) were positive for the MYD88 mutation exclusively in both flow cytometry sorted cell fractions but not in whole bone marrow specimens. 94% (16/17) WM as well as 78% (7/9) LPL specimens with identical plasma and B-cell clonality profile revealed the presence of the MYD88 L265P mutation in both the plasma cell and the B lymphoid cell clones. Whereas LPL specimens with unrelated clonality profile of the plasma and lymphoid cell fractions as well as other control specimens (lymphoma, myeloma, CLL and healthy) either tested negative or positive only in one of the sorted cell fractions.We therefore conclude that confirming the presence of MYD88 L265P in both B-lymphoid and plasma cell fraction is an important prerequisite to distinguish LPL/WM from related disorders with high confidence. DisclosuresWang:HematoLogics Inc.: Employment. Fritschle:HematoLogics Inc.: Employment. Bennington:HematoLogics Inc.: Employment. Burnworth:HematoLogics Inc.: Employment. Bennington:HematoLogics Inc.: Employment. Wentzel:HematoLogics Inc.: Employment. Verkamp:HematoLogics Inc.: Employment. Nguyen:HematoLogics Inc.: Employment. Ghirardelli:HematoLogics Inc.: Employment. Broderson:HematoLogics Inc.: Employment. Wells:HematoLogics Inc.: Employment, Equity Ownership. Loken:HematoLogics Inc.: Employment, Equity Ownership. Zehentner:HematoLogics Inc.: Employment, Equity Ownership.

Low-grade B-cell lymphoma presenting primarily in the bone marrow.

Cases of low-grade B-cell lymphoma presenting primarily in the bone marrow are rare, and its clinicopathology remains unclear. We retrospectively examined patients with low-grade B-cell lymphoma presenting primarily in the bone marrow. Fourteen patients met the inclusion criteria, including 5 with lymphoplasmacytic lymphoma (LPL), 3 with chronic lymphocytic leukemia/small lymphocytic lymphoma, 2 with follicular lymphoma (FL), and 4 with low-grade B-cell lymphoma not otherwise specified (LGBCL-NOS). The median age was 69.5 years (range, 42-89 years), and a slight male predominance was noted (9 men and 5 women, 1.8: 1). Immunohistochemically, all cases were positive for CD20. One case was positive for CD138. Both cases of FL were positive for CD10 and B-cell lymphoma 2 (BCL-2), and immunoglobulin heavy locus (IgH)/B-cell lymphoma 2 rearrangement was observed by fluorescence in situ hybridization. The myeloid differentiation primary response gene (88) leucine to proline mutation was observed in 3 of 5 LPL, 1 of 2 FL, and 2 of 4 LGBCL-NOS patients. Paraproteinemia was observed in 10 patients; IgM and IgG paraproteinemia were observed in 6 and 3 patients, respectively. In this patient series, 3 patients had died at a median follow-up of 36.5 months; the cause of death of 1 LPL patient was malignant lymphoma itself. Thus, low-grade B-cell lymphoma presenting primarily in the bone marrow has various subtypes, and approximately one-third of the patients had LGBCL-NOS. The immunophenotypic features and myeloid differentiation primary response gene (88) leucine to proline mutation data of LGBCL-NOS suggested that some cases present with characteristics similar to those of LPL or marginal zone lymphoma.

L265P Mutation of the MYD88 Gene Is Frequent in Waldenström’s Macroglobulinemia and Its Absence in Myeloma

L265P mutation in the MYD88 gene has recently been reported in Waldenström’s macroglobulinemia; however the incidence has been different according to the methods used. To determine the relevance and compare the incidence by different methods, we analyzed the L265P mutation in bone marrow mononuclear cells from lymphoid neoplasms. We first performed cloning and sequencing in 10 patients: 8 Waldenström’s macroglobulinemia; 1 non-IgM-secreting lymphoplasmacytic lymphoma; and 1 low grade B-cell lymphoma with monoclonal IgG protein. The L265P mutation was detected in only 1/8 Waldenström’s macroglobulinemia patients (2 of 9 clones). To confirm these results, direct sequencing was performed in the 10 patients and an additional 17 Waldenström’s macroglobulinemia patients and 1 lymphoplasmacytic lymphoma patient. Nine of 28 patients (7/25 Waldenström’s macroglobulinemia, 1/2 lymphoplasmacytic lymphoma, and B-cell lymphoma) harbored the mutation. We next tested for the mutation with BSiE1 digestion and allele-specific polymerase chain reaction in the 28 patients and 38 patients with myeloma. Aberrant bands corresponding to the mutation were detected by BSiE1 digestion in 19/25 patients with Waldenström’s macroglobulinemia (76%), 1/2 lymphoplasmacytic lymphoma and B-cell lymphoma, but not in the 38 myeloma patients. The L265P mutation was more frequent in patients with Waldenström’s macroglobulinemia than in those with myeloma (p=1.3x10-10). The mutation was detected by allele-specific polymerase chain reaction in 18/25 Waldenström’s macroglobulinemia patients (72%). In the 25 Waldenström’s macroglobulinemia patients, the L265P was more frequently detected by BSiE1 digestion than by direct sequencing (p=5.3x10-4), and in males (15/16, 94%) than in females (4/9, 44%) (p=1.2x10-2). No siginificant difference was observed in the incidence of the L265P mutation between BSiE1 digestion and allele-specific polymerase chain reaction (p=0.32). These results suggest that the L265P mutation is involved in the majority of Waldenström’s macroglobulinemia. BSiE1 digestion and allele-specific polymerase chain reaction may detect a small fraction of mutated cells in some cases.

Circulating B Cell Clones in Familial Waldenström Macroglobulinemia.

Abstract 2703 Background and Significance:Lymphoplasmacytic lymphoma (LPL) is an indolent but incurable B cell lymphoproliferative characterized by the clonal expansion of plasmacytoid lymphocytes in the lymph nodes and bone marrow. The majority of cases are associated with an IgM isotype paraprotein, and when present, is termed Waldenström macroglobulinemia (WM). WM shows familial aggregation suggesting an inherited risk for disease, and also co-aggregates with CLL in families. In our previous work, we have shown that clonal populations of peripherally circulating B cells can be identified in 18% of unaffected family members from CLL kindreds. Most of these clonal populations have a typical CLL immunophenotype and have been termed CLL-like monoclonal B cell lymphocytosis (MBL). Because CLL and WM have related gene expression profiles and appear to have shared genetic risk, we hypothesized that unaffected family members of WM kindreds would have detectable circulating clonal B cell populations. Further, we undertook systematic flow cytometric screening of familial WM and IgM MGUS cases to determine the prevalence, immunphenotype, and biologic characteristics of circulating malignant B cells. Methods:A diagnosis of LPL / WM or IgM MGUS was determined using standard WHO criteria. All patients and unaffected family members were ascertained at the National Cancer Institute and provided informed consent. Peripheral blood mononuclear cells were isolated using density centrifugation and viably frozen in DMSO containing media. We developed a two tube, nine color flow cytometric assay: the first tube allowed for detection of CLL-like clones based upon co-expression of CD5, CD20, and CD23; the second tube targeted WM populations based upon expression on CD19, CD20, CD25, CD38, and surface IgM. Cell populations were considered clonally restricted if the κ: λ was > 3.0 or < 0.3. Clonal populations were then isolated using fluorescence activated cell sorting (FACS). RNA and genomic DNA were extracted for genetic and genomic studies using phenol: chloroform purification. Results:A total of 155 individuals were analyzed: 54 WM / LPL, 17 IgM MGUS, 1 IgG MGUS, 1 IgA MGUS, 1 NHL, and 81 unaffected family members. Twenty of 54 WM patients had detectable peripherally circulating populations. Thirteen WM patients had no detectable B cells, of these, 11 patients had prior treatment. As such, among the 41 WM patients with a B cell compartment that was analyzable by flow cytometry, 49% (20 of 41) had peripherally circulating B cell clones detected. Four of these 20 cases showed two immunophenotypically distinct clonal B cell populations. The immunophenotype was somewhat heterogenenous: 18 cases expressed surface IgM, CD38 was variable but expressed in most cases, CD25 was not detected in any case, and 4 cases showed a CLL like (CD5+CD20dimCD23+) immunophenotype. Interestingly, we detected peripherally circulating B cell clones in 9 of 17 cases (53%) of IgM MGUS, a proportion nearly identical to that identified in WM / LPL. Three of 9 were “CLL-like” with co-expression of CD5 and CD23, while the majority of clones were CD5negIgM+CD38+. Among unaffected family members, we identified B cell clones in only 4 of 81 (5%). All 4 cases expressed CD5, and 3 showed a CLL-like phenotype, consistent with these individuals having MBL. Among all study subjects, 20 clonal B cell populations of > 104 B cells were FACS purified from 18 different cases: 12 WM, 4 IgM MGUS, 1 IgA MGUS, and 1 unaffected family member. Conclusions:Peripherally circulating B cell clones with an immunophenotype similar to that of LPL can be identified in approximately half of patients with WM / LPL. We observed for the first time that a similar proportion of patients with IgM MGUS have detectable clonal populations with an immunophenotype similar to that observed in WM patients. Genetic and genomic studies to determine the lineage of these populations are currently underway. The frequency of MBL among unaffected family members with WM is lower than that observed in CLL kindreds. CLL-like MBL can be detected at very low numbers because the cell population is immunophenotypically abnormal. The peripherally circulating clones identified in WM / LPL patients have an otherwise normal B cell immunophenotype and can only be detected by light chain restriction. This likely significantly limits the ability of flow cytometry to detect pre-clinical CD5neg IgM expressing clonal populations. Disclosures:No relevant conflicts of interest to declare.