IgM M-protein Research Articles

Non-Invasive MRD Monitoring in Multiple Myeloma Patients By Heavy/Light Chain Analysis

Introduction The majority of multiple myeloma (MM) patients are now achieving very deep responses. The most sensitive disease detection techniques are currently bone marrow assays. However, as bone marrow sampling is invasive and unpleasant for patients it is important to investigate whether blood-based assays could be equally or more informative of disease status. The heavy/light chain assay (Hevylite TM, The Binding Site, UK) evaluates serum heavy/light chains (HLC) for IgG, IgA and IgM M-proteins. This study assessed whether the HLC assay could act as an effective marker for disease in the bone marrow of treated MM patients and evaluated the prognostic utility of HLC measurements at best response and relapse time points. Methods 104 IgG and IgA MM patients were recruited either prior to transplant or during chemotherapy. Median follow up was 18 months. Routine M-protein tests were performed alongside HLC testing with parameters including involved HLC (iHLC), uninvolved HLC (uHLC) and kappa/lambda HLC ratio (HLCr). A bone marrow flow cytometry (NGF) assay was performed up to a sensitivity of 10 -6 and was considered to be the gold standard assay. Disease response was determined according to the IMWG criteria. Results and Discussion At pre- and post-transplant time points there was significant agreement between HLC measurements and NGF, with uHLC being the most sensitive measurement (86%) and HLCr the most specific (100%). Both uHLC and HLCr show substantial agreement with the bone marrow NGF method whilst iHLC cannot be used to indicate bone marrow status. Survival analysis showed significantly inferior progression free survival (PFS) in patients with an abnormal uHLC (p=0.01) (Figure A) or HLCr (p=0.02) result post-transplant but no survival difference based on iHLC status (p=0.78) (Figure B). Interestingly, performing survival analysis on these same samples using serum immunofixation (IFE) status (positive or negative) did not produce a significant PFS difference between participants (p=0.21). During long term follow up, HLC values showed a correlation with depth of response, with uHLC being abnormal in 60% of complete response (CR) patients. This suggests that those participants currently grouped as CR may be able to be further subdivided according to uHLC level and this could have a prognostic value in the monitoring and treatment of these patients. At best response uHLC was the only HLC or free light chain (FLC) measurement which gave a significant difference in PFS for all patients in partial response (PR) or better (p=0.005), remaining true when analysing patients in ≥CR only (p=0.002). The addition of free light chain measurements into the HLC analysis did not improve the significance of these survival differences. In relapsed patients, paired comparisons of M-protein and disease activity markers at best response vs. prior to clinical relapse were performed. uHLC was the only marker which was significantly different between the two time points in IgG and IgA patients. The significance of this was confirmed by showing that those patients in stable disease did not experience a significant change in uHLC levels. For other M-protein variables such as total immunoglobulin, IFE status and FLC values there was an increase in levels or abnormality between best response and pre-relapse but these were not significant. The paired comparison suggests that monitoring an individual using HLC could help to detect relapse earlier than conventional methods. In all analyses performed the HLC assay showed increased sensitivity and utility over the IFE technique. Conclusion This study suggests that the HLC assay can be used prior to bone marrow analysis to help influence decisions and improve the patient and laboratory experience. These results can be put into practice during patient follow up post initial treatment in the following ways: At deepest response an abnormal uHLC or HLCr suggests a positive MRD status, meaning a possible reduction or delay in the requirement for bone marrow assessment.All patients in ≥VGPR should have HLC performed alongside conventional assays during follow up.If uHLC values become abnormal during follow up then it should be assumed that the patient is about to undergo relapse. The measurement of HLC values have been shown to be a significant marker in this study, for sensitively detecting the M-protein, giving further clarification of response status and predicting early relapse.

Polyneuropathy Associated with IgM Monoclonal Gammopathy; Advances in Genetics and Treatment, Focusing on Anti-MAG Antibodies

With increasing age, the chances of developing either MGUS or polyneuropathy increase as well. In some cases, there is a causative relationship between the IgM M-protein and polyneuropathy. In approximately half of these cases, IgM targets the myelin-associated glycoprotein (MAG). This results in chronic polyneuropathy with slowly progressive, predominantly sensory neurological deficits and distally demyelinating features in nerve conduction studies. Despite the disease being chronic and developing slowly, it can cause considerable impairment. We reviewed English medical publications between 1980 and May 2022 on IgM gammopathy-associated polyneuropathy, with special attention to studies addressing the pathophysiology or treatment of anti-MAG polyneuropathy. Treatment options have been limited to a temporizing effect of intravenous immunoglobulins in some patients and a more sustained effect of rituximab but in only 30 to 55 percent of patients. An increase in our knowledge concerning genetic mutations, particularly the MYD88L265P mutation, led to the development of novel targeted treatment options such as BTK inhibitors. Similarly, due to the increasing knowledge of the pathophysiology of anti-MAG polyneuropathy, new treatment options are emerging. Since anti-MAG polyneuropathy is a rare disease with diverse symptomatology, large trials with good outcome measures are a challenge.

Dutch Physician's Perspectives on Diagnosis and Treatment of Waldenström's Macroglobulinemia Before and After the Implementation of a National Guideline.

Waldenström’s macroglobulinemia (WM), a rare low-grade B-cell non-Hodgkin lymphoma (NHL), has a distinct clinical presentation and different treatment-related side effects compared with other NHL. Currently, a wide variety of therapeutic agents are available for the treatment of WM but there is no consensus on optimal treatment in first line and/or at relapse. The aim of this survey was to evaluate the current knowledge and perspectives of hematologists on diagnosis and treatment of WM. Also, we compare these results to a similar survey done before the publication of the first Dutch national guideline, in order to evaluate the impact of the implementation of a national guideline. A link to an online survey was sent out to all registered hematologists and hemato-oncologists in the Netherlands with the request to participate. The survey contained questions regarding the preferred diagnostic and treatment methods in patients with WM as well as treatment goals. We also compared physicians preferred treatment goals to those of patients (as studied in a recent nationwide patient questionnaire). Ninety-five responses (30% response rate) were obtained, out of which 82 (86%) surveys were complete. The respondents most commonly used dexamethasone-rituximab-cyclophosphamide as first-line treatment. For second-line treatment, bendamustine with rituximab and ibrutinib monotherapy were the most frequently applied. Compared with the initial survey, serum IgM M-protein was determined in all cases, MYD88 mutation analysis was currently widely implemented, prevention of an IgM “flare” was uniformly managed by the respondents and use of rituximab-cyclophosphamide-vincristine-prednisone was entirely abandoned. Physicians differed somewhat from patients with regard to most important treatment goals. The approach to diagnostic methods and treatment options in WM was more consistent with international guidelines and was more homogeneous after implementation of the national guideline. These data indicate an increase in knowledge on WM diagnosis and treatment. This may have resulted from implementation of a local guideline or the global rise in awareness and attention for WM.

Study on the Mechanism of False Low Measurement of IgG-Binding (Affinity) IgM Type M Protein by Turbidimetric Immunoassay.

The purpose of this study was to investigate the immunological and physical characteristics of IgM-λ type M-protein from patients who were measured low in the turbidimetric immunoassay (TIA) IgM assay without error codes for high concentration to determine the cause of the false low levels and to clarify the mechanism of their occurrence. Materials were IgM patient samples and 8 serum samples from other IgM M-protein patients as controls. Patient samples were assayed by the TIA method, in which five manufacturers and six models (two reagent manufacturers) share the principle, and the BN ProSpec method (nephelometric method), which has a different principle. Dilution linearity tests, IgG addition experiments, isoelectric point electrophoresis, and hydrophobic chromatography were performed on patients and subjects. In addition, the binding capacity of γ-globulin by BIACORE was also examined. The reaction curve of the patient IgM curved downward when the concentration of IgM exceeded 20 g/L, and no error code was obtained. In the measurement by the TIA method of five manufacturers and six models, patient IgM was measured at a false low level with no error code obtained in undiluted dilution by any of the instruments and reagents, but could be measured without any problem by the nephelometric method. In addition, in the patient IgG addition experiment, only patient IgM showed a false low level under high IgG concentration. Furthermore, the binding capacity of patient IgM to γ-globulin (IgG) by BIACORE was significantly higher than that of the control IgM-type M protein. Patient IgM has an affinity (binding capacity) for IgG and forms an IgM-IgG complex under conditions of high IgG concentration. It was speculated that this complex inhibited the reaction with the anti-IgM antibody and the absorbance of the second reaction did not increase, suggesting a false low.

Comparative Outcomes of Immunochemotherapy with RCD (rituximab, cyclophosphamide, dexamethasone) and BCD (bortezomib, cyclophosphamide, dexamethasone) in Waldenstrom Macroglobulinaemia: NCT02971982

Objective: To compare the outcomes of immunochemotherapy with rituximab based treatment and bortezomib treatment in pantines with Waldenstrom macroglobulinaemiaMethod: The randomized study enrolling 11eligible patients at first affiliated hospital of Soochow university aged from 44 years old to 71 years old. All of them were diagnosed by bone marrow biopsy, immunofixed electroresis with IgM-M protein and MYD88L265P mutation, all of them had clinical symptoms such as fever, weak, loss of weigh, hemorrhage and so on. These patients with newly diagnosed WM were randomly to two groups to receive treatment. 4 patients were treated by bortezomib, cyclophosphamide and dexamethasone (PCD), the other 7 patients were treated by rituximab, cyclophosphamide and dexamethasone (RCD), the primary end point was overall response rate (ORR), and major response rate (MRR).Results: The median age of 11 patients was 63 years old (range: 44-71 years), with a male to female ratio was 4.5. ALL of them had clinical symptoms, most of them had non-specific symptoms such as weak, fever (36.6%, 4/11), and loss of weight (18.2%, 2/11).One of them had coagulation disorder with retroperitoneal hematoma, and one had rashes. 4 patients treated by PCD received 6 cycles and 7 patients treated by RCD received 3 to 6 cycles (average 5 cycles). The response evaluation is arcaded by the symptoms and M-protein in plasma. All of the clinical symptoms were remitted by treatment in each group after one to four cycles. 9 patients with anemia at first and 88.8% (8/9) were remitted. To evaluate the M-protein, the overall response rate (ORR) of Patients who received PCD treatment is 75% (3/4), and major reaction rate (MRR) (1 patient was PR, 1 patient was VGPR, and 1 patient was CR) is 75%. The ORR of patents received RCD treatment is 42.85% (3/7), and the MRR is 14.28(1 patient reached PR). The two group is not statistical significance in ORR (P=0.554), but the MRR is significantly better in the PCD arm compared with RCD arm (95%CI, 75% to 42.85%, P=0.044).The toxicity of the PCD arm was peripheral neuritis (100%), I to II grade, mostly accrued after 4 cycles of bortezomib, and one of them with dosage reduction to 1.0mg/m2 by bortezomib. The severe adverse event was hepatotoxicity by extending interval treatment of each cycle. The major toxicity of RCD arm was infusion reaction of rituximab (37.1%, 4/7), such as fever, chest congestion and hypoxemia, mostly at first to third cycle. All can be relieved by slow infusion and oxygen inhalation. The incidence rate of toxicity was similar in two groups (P=0.125).Conclusion: Rituximab and Bortezomib are new immunochemotherapy for Waldenstrom macroglobulinaemia, both (combination with other agent) are recommended as a first-line treatment for WM. In our preliminary statistics, both of them had effect, but the MRR is better in PCD group. Bortezomib may can act on the NF-κB cytokine pathway. But our samples were limited, we still need to be confirmed by expanding the samples quantity and extending the time of intervention. DisclosuresNo relevant conflicts of interest to declare.

Association of variability in antibody binding affinity with a clinical course of anti-MAG neuropathy

Anti-myelin-associated glycoprotein (MAG) neuropathy is mediated by the binding of IgM M-proteins to the human natural killer-1 epitope of several glycoconjugates, including MAG and phosphacan. We recently reported that IgM M-proteins with a higher ratio of anti-phosphacan titer to anti-MAG titer (P/M ratio) were associated with a progressive clinical course. Herein, we investigated the temporal variability of the P/M ratio. The results showed that P/M ratios in worsened cases were significantly increased relative to stable or improved cases. Thus, temporal variability in the specificity of IgM M-proteins may be related to the disease course of anti-MAG neuropathy.

Binding specificity of anti-HNK-1 IgM M-protein in anti-MAG neuropathy: Possible clinical relevance

Anti-myelin-associated-glycoprotein (MAG) neuropathy is an intractable autoimmune polyneuropathy. The antigenic region of MAG is the human natural killer-1 (HNK-1) carbohydrate. We and others previously suggested that the extension of antibody reactivities to HNK-1-bearing proteins other than MAG was associated with treatment resistance, without statistical analyses. In this study, we established an ELISA method with recombinant proteins to test binding specificities of currently available monoclonal antibodies to MAG and another HNK-1-bearing protein, phosphacan. Using this system, we found the distinct binding specificities of anti-MAG antibody in 19 patients with anti-MAG neuropathy. Their clinical relevance was then determined retrospectively with the adjusted 10-points INCAT disability score (0=normal and 10=highly disable). The results showed that strong reactivities of anti-MAG antibodies to phosphacan were significantly associated with treatment resistance or progressive clinical courses, indicating a possible clinical relevance of the binding specificities.

Carfilzomib, Rituximab and Dexamethasone (CaRD) Is Highly Active and Offers a Neuropathy Sparing Approach For Proteasome-Inhibitor Based Therapy In Waldenstrom's Macroglobulinemia

Bortezomib is active in Waldenstrom's macroglobulinemia (WM) but associated with considerable peripheral neuropathy (PN). The proteasome inhibitor carfilzomib (CFZ) is approved in the USA for relapsed/refractory myeloma. Herein, we examined the efficacy and safety of carfilzomib, rituximab, and dexamethasone (CaRD) in 31 proteasome inhibitor and rituximab naive WM patients with symptomatic disease. Median baseline characteristics: age 61, prior therapies 0 (range 0-1), hematocrit 32.3%, hemoglobin 10.7 g/dL, serum IgM 3375 mg/dL, serum M-protein 2.185 g/dL, B2M 3.6 mg/L, and bone marrow disease involvement 60%. Therapy consisted of IV CFZ 20 mg/m2 (cycle 1) then 36 mg/m2 (cycles 2 and beyond) with IV dexamethasone (dex) 20 mg given on days 1,2,8,9 and rituximab 375 mg/m2 on days 2,9 of each 21-day cycle. Treatment consisted of six induction cycles, then maintenance beginning 8 weeks after induction (given every 8 weeks for eight cycles; consisted of CFZ 36 mg/m2, and IV dex 20 mg on days 1,2 and rituximab 375 mg/m2 on day 2). Patients with IgM level >4000 mg/dL underwent plasmapheresis and/or had rituximab held until IgM <4000 mg/dL to prevent symptomatic IgM flare. Patients received oral acyclovir (400 mg twice daily) and famotidine (20 mg twice daily) as concomitant medications. For all 31 patients, median serum IgM levels and M-protein declined to 749 mg/dL and 0.7 g/dL, respectively (p<0.00001). Median hematocrit and hemoglobin rose to 40.9% and 13.7 g/dL, respectively (p<0.00001). A total of 30 patients concluded induction therapy with bone marrow tumor involvement reduced to a median of 7.5% (p=0.0003). The best overall response rate using criteria adapted from the 3rd International Workshop on WM was 81% (1 CR, 8 VGPR, 12 PR, 4 Minor Responses). With a median follow-up of 8 cycles, 22 patients remain on study, including 20 currently on maintenance therapy. Median time to response (for minor responders or better) was 2.1 months. Grade >2 treatment related toxicities included asymptomatic elevation in lipase (12.9%), dex-related hyperglycemia (6.45%), reversible neutropenia (9.67%), and cardiomyopathy (3.22%). There were no grade 2 or greater PN events. Treatment discontinuation occurred for non-response (n=8), cardiomyopathy in a patient with multiple cardiac risk factors (n=1), and progressive disease (n=1). CaRD is highly active and offers a neuropathy sparing approach for proteasome-inhibitor based therapy in WM. Disclosures:Off Label Use: Carfilzomib is a novel proteasome inhibitor, which offers a neuropathic sparing approach on waldenstrom macroglobulinemia disease when used in a combination therapy with rituximab and dexamethasome.

A Prospective Multicenter Study Of The Bruton's Tyrosine Kinase Inhibitor Ibrutinib In Patients With Relapsed Or Refractory Waldenstrom's Macroglobulinemia

IntroductionWhole genome sequencing has revealed highly prevalent somatic mutations in WM (Hunter et al, ICML-12, 2013). MYD88 L265P is present in >90% of patients with Waldenstrom's Macroglobulinemia (WM), and supports malignant growth via signaling involving Bruton's Tyrosine Kinase (BTK). Ibrutinib inhibits BTK, and in vitro induces apoptosis of WM cells bearing MYD88 L265P (Yang et al, Blood 2013). WHIM-like mutations in CXCR4 are present in 1/3 of patients with WM, and their expression induces BTK activity and confers decreased sensitivity to ibrutinib mediated growth suppression in WM cells (Cao et al, ASH 2013, submitted). We therefore evaluated the efficacy and tolerability of ibrutinib in relapsed or refractory WM, and examined the impact of MYD88 L265P and WHIM-like CXCR4 mutations on ibrutinib response given our laboratory findings. Patients and MethodsSymptomatic WM patients who received at least 1 prior treatment were enrolled on this prospective clinical trial. Intended therapy consisted of 420 mg of oral ibrutinib daily for 2 years or until progression, or unacceptable toxicity. Sanger sequencing was used to determine MYD88 and CXCR4 mutations in sorted bone marrow lymphoplasmacytic cells (BM LPC) from 43 and 40 patients, respectively. Forty of 43 (93%) and 10/40 (25%) patients had MYD88 L265P and WHIM-like CXCR4 mutations, respectively. Results63 patients including 17 with refractory disease were enrolled; all 63 are evaluable for response and toxicity. Median baseline characteristics: Age 63 (range 44-86) ; Prior therapies 2 (range 1-6); Hematocrit 30.8% (range 24.5-41.5); Hemoglobin 10.5 g/dL (8.2-13.8 g/dL); serum IgM 3,610 mg/dL (range 735-8390 mg/dL); serum M-protein 2.14 g/dL (range 0.5-5.4 g/dL); B2M 3.9 mg/L (range 1.3-14.2 mg/L); BM disease involvement 65% (range 3.2-95%). At best response, median serum IgM levels and M-protein declined to 1,340 mg/dL and 0.84 g/dL, respectively (p<0.00001). Median hematocrit and hemoglobin rose to 38.1% and 12.6 g/dL, respectively (p<0.00001). Post-treatment bone marrow assessment at 6 months is available for 34 patients at this time, and showed a reduction from 70% to 45% in WM disease involvement for these patients (p=0.0006). With a median follow-up of 6 (range 2-15 cycles), the best overall response rate i.e. minor response (MR) or better using consensus criteria adapted from the 3rd International Workshop on WM is 81% (4 VGPR; 32 PR, 15 MR), with a major response rate (PR or better) of 57.1% and a median time to response of 4 weeks. 11 patients have stable disease, and 1 patient was a non-responder. Independent disease and response assessments are planned. Grade >2 treatment related toxicities include thrombocytopenia (n=9; 14.3%); neutropenia (n=12; 19.1%); stomatitis (n=1; 1.6%); atrial fibrillation (n=1; 1.6%); diarrhea (n=1; 1.6%); herpes zoster (n=1; 1.6%); hematoma (n=1; 1.6%); hypertension (n=1; 1.6%) and epistaxis (n=1; 1.6%). 59 patients remain on study with 7 on reduced doses of ibrutinib. Reasons for discontinuation include non-response (n=1) in a patient with wild type MYD88; MDS/RAEB (n=1) in a heavily pre-treated patient who attained PR, but who had 5q deletions pre-dating protocol therapy; thrombocytopenia (n=1) in a patient with splenic entrapment; and patient decision (n=1). In patients who underwent tumor sequencing, attainment of major responses was impacted by mutations in CXCR4, but not MYD88 L265P. The major response rate was 77% for patients with wild-type CXCR4 vs. 30% in those with WHIM-like CXCR4 mutations (p=0.018). Decreases in serum IgM (p=0.047) and IgM M-spike (p=0.012), as well as improvements in hemoglobin (p=0.058) were greater in patients with wild-type CXCR4. Patients with wild-type CXCR4 also had increased peripheral lymphocytosis following ibrutinib treatment versus those with WHIM-like CXCR4 mutations (p=0.001). ConclusionsIbrutinib is highly active, and well-tolerated in patients with relapsed or refractory WM. Rapid reductions in serum IgM and improved hematocrit occur in most patients receiving ibrutinib. The presence of WHIM-like CXCR4 mutations impacts responses and peripheral lymphocytosis in WM patients undergoing ibrutinib treatment. Disclosures:Off Label Use: Ibrutinib is a Bruton's Tyrosine Kinase (BTK)inhibitor that, in vitro, induces apoptosis of Waldenstrom's macroglobulinemia cells bearing somatic mutations of MYD88 L265P.

Clinical and laboratory features of neuropathies with serum IgM binding to TS‐HDS

In this investigation we studied clinical and laboratory features of polyneuropathies in patients with serum IgM binding to the trisulfated disaccharide IdoA2S-GlcNS-6S (TS-HDS). We retrospectively compared 58 patients with selective IgM binding to TS-HDS to 41 consecutive patients with polyneuropathies without TS-HDS binding. Patients with IgM vs. TS-HDS commonly had distal, sensory, axonal neuropathies. Weakness was associated with IgM M-proteins. Hand pain and serum IgM M-proteins were more common than in control neuropathy patients. TS-HDS antibody binding was often selectively κ class. Biopsies showed capillary pathology with thickened basal lamina and C5b9 complement deposition. IgM in sera with TS-HDS antibodies often bound to capillaries. Serum IgM binding to TS-HDS is associated with painful, sensory > motor, polyneuropathies with an increased frequency of persistent hand discomfort, serum IgM M-proteins, and capillary pathology. Serum IgM binding to TS-HDS suggests a possible immune etiology underlying some otherwise idiopathic sensory polyneuropathies.

The role of sulfoglucuronosyl glycosphingolipids in the pathogenesis of monoclonal IgM paraproteinemia and peripheral neuropathy

In IgM paraproteinemia and peripheral neuropathy, IgM M-protein secretion by B cells leads to a T helper cell response, suggesting that it is antibody-mediated autoimmune disease involving carbohydrate epitopes in myelin sheaths. An immune response against sulfoglucuronosyl glycosphingolipids (SGGLs) is presumed to participate in demyelination or axonal degeneration in the peripheral nervous system (PNS). SGGLs contain a 3-sulfoglucuronic acid residue that interacts with anti-myelin-associated glycoprotein (MAG) and the monoclonal antibody anti-HNK-1. Immunization of animals with sulfoglucuronosyl paragloboside (SGPG) induced anti-SGPG antibodies and sensory neuropathy, which closely resembles the human disease. These animal models might help to understand the disease mechanism and lead to more specific therapeutic strategies. In an in vitro study, destruction or malfunction of the blood-nerve barrier (BNB) was found, resulting in the leakage of circulating antibodies into the PNS parenchyma, which may be considered as the initial key step for development of disease.

Hevylite®, a New Marker of Tumor Measurement In Waldenstrom Macroglobulinemia

AbstractAbstract 5076 Background.Waldenstrom macroglobulinemia (WM) is a low grade B cell lymphoma characterized by bone marrow infiltration of lymphoplasmacytic tumor cells that secrete monoclonal IgM (M-protein) into the serum. Measurement of the serum M-protein [using serum protein electrophoresis (SPEP)] and measurement of total IgM (using nephelometry) are used to diagnose and monitor WM. There are, however, many limitations with these techniques and new markers are needed. IgG and IgA Hevylite® immunoassays have been reported to be more sensitive than SPEP and nephelometry for identifying monoclonal immunoglobulins in multiple myeloma and unlike immunofixation, provide quantitative information. We hypothesized that serum IgM Hevylite assays (specifically measuring IgMkappa and IgMlambda, separately) would accurately identify serum IgM M-proteins. We also evaluated the association between known tumor burden markers and prognostic factors with IgM Hevylite results in patients with WM. Method.We retrospectively measured IgMkappa and IgMlambda in sera from 59 WM patients: 44 patients were at diagnosis and 15 had relapsed disease. The diagnosis of WM was made according to the current guidelines. All serum samples were kept frozen in the Lille serum bank since collection. All patients gave informed consent prior to the collection and none were treated at time of collection of the serum. Approval of this protocol was obtained from the CHRU Lille and was in accordance with the Declaration of Helsinki.Hevylite measurements were made at The Binding Site Ltd, Birmingham, UK. A normal range was produced from normal (blood donor) sera (n=120), median (and 95%ile ranges) were; IgMkappa 0.634g/L (0.29-1.82), IgMlambda 0.42g/L (0.17-0.94), IgMkappa/IgMlambda ratio 1.6 (0.95-2.3). For ease of comparison IgM hevylite ratios were expressed as the involved monoclonal immunoglobulin/uninvolved polyclonal immunoglobulin (IgMi). To describe the distribution of IgMi Hevylite levels in patients with WM, the median and range (min-max) were reported. Median values were compared using the Wilcoxon rank-sum test and ANOVA. Fisher's exact test was used to compare proportions. All statistical tests were two-sided. All analyses were conducted using SPSSv12 software. Results.The baseline characteristics of the patients were as follows: the median (range) age was 68 years (41-86), male/female 38/21, serum b2M 3.0mg/L (1.2-9.0), hemoglobin 11.8g/dL (7.6-15.4), platelet count 267 ×109/mm3 (55-741), serum M-spike 19g/L (3.0-52.7). In our series, 18 (31%), 22 (37%) and 19 (32%) patients had low, intermediate and high risk disease respectively, in the WM-IPSS scoring system. The median (min-max) IgMkappa ratio was 134 (8.7-2850) and IgMlambda ratio was 0.03 (0.0007-0.39). IgMi Hevylite ratio was 98.07 (2.59-2850). The IgMi Hevylite correlated well with the M-spike measured using SPEP (r=0.601, p<0.0001).In our study, high IgMi Hevylite levels correlated well with markers of high tumor burden and of poor prognosis. The median (range) IgMi Hevylite level was higher in patients with hemoglobin <10g/dL versus ≥10 g/dL, 267 (8.1-1722) and 76 (2.6-2850) respectively (p=0.013). The median (range) IgMi Hevylite ratio level was significantly (p=0.033) higher in the high risk IPSS group 208 (2.6-2850) than in the intermediate 75 (15-1033) and low risk groups, 98 (8-571).The IgMi Hevylite levels also separated WM patients with progressive disease who required therapy. Twenty seven pts were symptomatic and required specific treatment for WM, and 32 pts were left untreated. The median IgMi Hevylite ratio was significantly higher in progressing patients, 210 (8.1-2850) and 60 (2.6-571) in the 2 groups, respectively (p=0.014). Conclusion.In this study we demonstrated that IgM Hevylite measurement is a new and reliable marker for monitoring WM disease. It is related to poor prognostic markers that separate WM patients with progressive disease who require therapy. We are currently expanding the cohort to confirm these observations. These findings have implications in the management of patients with WM. Disclosures:Leblond:Roche: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Mundipharma: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Genzyme: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Janssen: Membership on an entity's Board of Directors or advisory committees. Leleu:Celgene: Consultancy, Research Funding; Janssen Cilag: Consultancy, Research Funding; Leo Pharma: Consultancy; Amgen: Consultancy; Chugai: Research Funding; Roche: Consultancy, Research Funding; Novartis: Consultancy, Research Funding.

Central nervous system involvement in patients with monoclonal gammopathy and polyneuropathy

To evaluate clinical presentation of patients with the clinical triad of monoclonal gammopathy, polyneuropathy and signs of CNS involvement. Nineteen patients with monoclonal protein (M-protein, 9 IgM, 10 IgG) were studied. Clinical examination, MRI, cerebrospinal fluid analysis and immune reactivity against myelin-associated glycoprotein and gangliosides in serum were obtained. By immunohistochemistry, different binding patterns of M-proteins to human CNS tissue were investigated. Nine out of 19 patients (four IgM, five IgG) showed one or more clinical signs of CNS involvement. Clinical features associated with signs of CNS pathology were disease duration and greater concentration of IgM paraprotein. The IgM M-protein of two patients strongly stained the cortex/cerebellar neurons in human brain sections. Our results complement previous reports that some patients with monoclonal gammopathy and polyneuropathy can develop solitary or disseminated signs of CNS involvement. It indicates that pathological effects of M-proteins are not necessarily restricted to the peripheral nervous system. The specificity and affinity of circulating M-protein to antigens in the CNS might be critical for the development of different clinical phenotypes.

Two Distinct Syndromes of Lymphoma-Associated Amyloidosis: Findings From a Case Series.

Abstract 2938 Poster Board II-914 Introduction: Light chain (AL) amyloidosis has a rare but recognized association with non-Hodgkin lymphoma; however, limited information regarding this association exists. We present the clinical features and outcomes of patients with lymphoma -associated AL amyloidosis identified at Princess Margaret Hospital (PMH). We describe two distinct syndromes of lymphoma-associated amyloidosis: 1) peritumoral amyloidosis in which amyloid deposition occurs in the immediate vicinity of the lymphoma and 2) systemic amyloidosis where deposition is in sites remote from the underlying lymphoma. Methods: Cases were identified retrospectively at PMH by survey of hematologists and a pathologist. The International Consensus Criteria (ICC) (Gertz et al., 2005) were used to determine organ involvement by amyloid. Results: We identified 10 patients with lymphoma -associated AL amyloidosis seen between 1997 and 2008. The median age was 69 (52-81) and 50% were female. Five patients had peritumoral amyloidosis. All had extranodal marginal zone lymphoma, MALT subtype (MALT lymphoma).A monoclonal immunoglobulin (M-protein), as determined by serum protein electrophoresis (SPEP) and free light chain (FLC) analysis, was undetectable in two patients, was present in trace amount in two patients and detected but not quantified in a fifth. Of the patients with detectable M-protein, the isotype was IgM in two and IgG in one. Four had lambda light chain restriction while biclonality was questioned in the fifth. All but one patient had single organ involvement by amyloid with lung and soft tissues being the only sites affected. Rectal biopsies and fat pad aspirates, when performed, were negative. Four patients received systemic therapy: two patients were treated with chlorambucil, one received melphalan and prednisone, and one received rituximab, cyclophosphamide, and prednisone followed by rituximab maintenance. Radiographic follow-up of pulmonary lesions in two patients did not identify significant change in lymphoma/amyloid related abnormality. No complete responses were seen. Despite therapy, three patients experienced continuing local symptoms at the site of lymphoma/amyloid involvement while one remained asymptomatic. A fifth patient, untreated at last follow up, was asymptomatic. No patients with peritumoral amyloidosis were found to develop systemic amyloidosis even after a median follow up of 11 months (3-60 mo). The other five patients had lymphoma with systemic amyloidosis. These patients had lymphoma restricted to the bone marrow with histology showing lymphoplasmacytic lymphoma (LPL) in three and small B cell lymphoma with plasmacytic differentiation in two (differential diagnosis LPL vs marginal zone lymphoma). An IgM M-protein was detectable in all five patients. Waldenstrom's macroglobulinemia was therefore present in at least three patients. High levels of M-protein were detected by SPEP in four patients (mean 31 g/L) and FLC analysis in one. Kappa was more common than lambda light chain restriction (3:2). Amyloid was detected in a median of three organs (1-5) including cardiac involvement in four patients. Four patients received multiple lines of therapy with first line being R-CVP in three and ASCT in a fourth. Organ responses, as defined by the ICC, in sites affected by amyloid were not observed. Three of the four treated patients had &gt;50% reductions in M-protein levels, but none had complete haematological response. Response of the underlying lymphoma, which would have required repeat biopsy, was not assessed. Median follow up was 30 months (4-100) with one patient dying of sepsis, one developing therapy-related AML and three surviving with ongoing symptoms of congestive heart failure, dyspnea and/or neuropathy. Conclusions: Lymphoma-associated amyloidosis presents as two distinct syndromes which are associated with characteristic clinical, laboratory and pathologic features. Patients with peritumoural amyloidosis have low level M-protein expression with predominant lambda light chain restriction, symptoms confined to the site of their MALT lymphoma and poor radiological responses to systemic therapy. Patients with systemic amyloidosis have high M-protein levels with a uniform IgM isotype, lymphoma localized to the bone marrow, multiorgan involvement by amyloid and worse outcomes. Future studies aimed at better defining prognosis and optimal treatment in these patients are warranted. Disclosures: Kukreti: Celgene: Honoraria.

Peripheral neuropathy in monoclonal gammopathy of undetermined significance: prevalence and immunopathogenetic studies.

In an unselected series of patients with monoclonal gammopathy of undetermined significance (MGUS) we found neuropathy in 2 of 34 patients with IgG (6%), 2 of 14 with IgA (14%), and 8 of 26 with IgM MGUS (31%). The neuropathy was subclinical in 6 patients (1 IgG, 1 IgA, and 4 IgM). Patients with IgG or IgA MGUS had a prominent motor impairment with electrophysiologic and morphologic findings suggestive of predominant axonal degeneration. No deposit of the M-protein in sural nerve and no reactivity of the M-protein with nerve was detected in these patients. Patients with IgM MGUS had a prominent sensory impairment with evidence of predominant demyelination. In 6 of these patients the M-protein reacted with the myelin-associated glycoprotein (MAG). The higher prevalence of neuropathy in patients with IgM MGUS may be related to the frequent reactivity of IgM M-proteins with MAG.

Anti-D Antibody in the Serum of an Rh-Positive Vietnamese Woman

© symptoms.5 It is speculated that the therapeutic benefit of IFN-α is related to its combined antilymphoproliferative and antiviral actions.5 Given the availability of a specific therapy, it is important that HCV-associated type II cryoglobulinemia be recognized and diagnosed as early as possible. It should be part of the differential diagnosis in patients with an unexplained IgM M-protein, signs/symptoms of cryoglobulinemia, a positive cryoglobulin assay, and/or an unexplained positive rheumatoid factor assay.

2-Mercaptoethanol Treatment Improves Measurement of an IgMκ M-Protein by Capillary Electrophoresis

A recent consensus conference on monoclonal gammopathies recommended the use of high-resolution electrophoresis, which achieves crisp separation of the major β-1 (transferrin) and β-2 (C3) bands (1)(2). Capillary electrophoresis (CE) provides this degree of resolution and has an additional advantage of automating both the analysis and the presentation of serum protein electrophoresis results (3)(4)(5). We report a rare discrepancy in the CE measurement of an IgM M-protein in the serum. The patient, a 59-year-old man, was first referred in August 1999 for evaluation of alcohol dependence, thrombocytopenia, and splenomegaly. Hemoglobin was 140 g/L, the white blood cell count was 4.7 × 109/L, and the platelet count was 54 × 109/L. A bone marrow aspirate was almost entirely plasmacytoid lymphocytes consistent with Waldenstrom macroglobulinemia. In addition to the lymphoproliferative process, the patient suffered from alcoholic hepatitis. On the day the sample …

Variability in immunohistochemistries of IgM M-proteins binding to sulfated glucuronyl paragloboside

Serum IgMs from 4 of 12 patients with polyneuropathy and IgM M-proteins that bind to sulfated glucuronyl paragloboside (SGPG) strongly immunostained the human peripheral nerve myelin (group A), whereas those from the other eight patients strongly immunostained the cytoplasm of the Schwann cells surrounding the myelin sheath with only weak staining of the myelin (group B). Strong immunostaining of peripheral myelin by IgMs from group A patients may be due to the strong cross-reactivities against P0 and peripheral myelin protein-22 (PMP-22), which are localized in compact myelin. Only three patients (all in group B) showed some response to the immunotherapies. Weak reactivities to P0 and and PMP-22 might indicate the possibility of improvement after the immunotherapies.

Anti-SGPG antibody in CIDP: nosological position of IgM anti-MAG/SGPG antibody-associated neuropathy.

Polyneuropathy with monoclonal gammopathy usually is considered a nosological entity different from chronic inflammatory demyelinating polyneuropathy (CIDP). Criteria proposed by the American Academy of Neurology AIDS Task Force (1991), however, show monoclonal gammopathy to be a condition concurrent with CIDP. The purpose of this study was to clarify the nosological relationship between CIDP and IgM anti-myelin-associated glycoprotein (MAG)/sulfated glucuronyl paragloboside (SGPG)-associated polyneuropathy. We investigated IgM anti-MAG/SGPG antibody in 85 CIDP patients by various methods, then examined the relation of M-protein to the presence of IgM anti-MAG/SGPG antibody. In our large study, 17 (20%) of 85 CIDP patients had high IgM anti-SGPG antibody titers in the enzyme-linked immunosorbent assay. This was confirmed by thin-layer chromatography-immunostaining for IgM anti-SGPG antibody and immunoblotting for IgM anti-MAG antibody. Immunoelectrophoresis and immunofixation, respectively, detected IgM M-protein in 6 (35%) and 13 (76%) of the 17 CIDP patients. We conclude that some patients with IgM anti-MAG/SGPG antibody with or without monoclonal gammopathy may be diagnosed as having CIDP, when patients are diagnosed according to the current CIDP criteria.

Long-term prognosis of neuropathy associated with anti-MAG IgM M-proteins and its relationship to immune therapies.

Many data point to a pathogenetic role for IgM antibodies to the myelin-associated glycoprotein (MAG) in the neuropathy associated with IgM monoclonal gammopathy, supporting the use of immune therapies in affected patients. Almost 50% of patients have been reported to improve with these therapies, but the effect of treatment on the long-term prognosis of the neuropathy remains unclear. We analysed the outcome of 25 of the 26 patients (mean age at entry 65 years, range 45-85 years) with neuropathy and high anti-MAG IgM, first examined by us between 1984 and 1994. By January 1999, after a mean follow-up of 8.5 years (range 2-13 years) and a mean duration of neuropathy symptoms of 11.8 years (range 3-18, >10 years in 16), 17 patients (68%) (aged 58-84 years, mean 73.4) were alive, while eight (32%) (aged 69-78 years, mean 73.1) had died 3-15 years (mean 10.6) after neuropathy onset; in none of them was death caused by the neuropathy, although in three it was possibly related to the therapy for the neuropathy. By the time of last follow-up or patients' death, 11 patients (44%) were disabled by severe hand tremor, gait ataxia or both. The disability rates at 5, 10 and 15 years from neuropathy onset were 16, 24 and 50%, respectively. Of the 19 patients treated during the follow-up for 0.5-11 years (mean 4 years) with various immune therapies, five reported a consistent and four a slight improvement in the neuropathy (total 47%) after one treatment or more, but in only one patient was improvement persistent throughout, to the end of follow-up. In 10 patients (53%), severe adverse events, possibly related to therapy, occurred during treatment and were considered responsible for the patient's death in three. The neurological impairment did not differ between treated and untreated patients at the end of a similar follow-up. Our findings indicate that (i) the majority of patients with neuropathy and anti-MAG IgM have a favourable prognosis even after several years, and (ii) current immune therapies, though temporarily effective in half of the patients, are associated with considerable side effects which limit their prolonged use and efficacy, suggesting that until more effective or safer therapies become available, they should probably be reserved for patients impaired in their daily life or in a progressive phase of the disease.