Serum Monoclonal Immunoglobulin Research Articles

Serum monoclonal immunoglobulin light-chain detection differs between immunofixation electrophoresis methods in patients with AL amyloidosis.

Serum immunofixation electrophoresis (IFE) is often performed for screening monoclonal proteins (M proteins) in immunoglobulin light-chain amyloidosis (AL amyloidosis). However, the performance of serum IFE for detecting M protein in AL amyloidosis patients is often insufficient. In this study, we examined the detection rate of serum M protein in newly diagnosed AL amyloidosis patients and analyzed differences in M protein detection between IFE methods. Among 60 patients newly diagnosed with AL amyloidosis, 22 had undetectable serum M protein by IFE with the Epalyzer2 system. Samples with undetectable M protein had significantly lower involved serum-free light-chain (iFLC) and a smaller difference between involved and uninvolved serum-free light-chain (dFLC) values than samples with IFE-detectable monoclonal light chains. When samples that tested negative for M protein by the Epalyzer2 system were retested by IFE with the HYDRASYS 2 system, 50% had IFE-detectable monoclonal light chains. The IFE system and reagents used may affect serum monoclonal immunoglobulin light-chain detection in AL amyloidosis patients, especially those with low iFLC or low dFLC samples. More attention should be paid to the performance of IFE systems, since it may affect the diagnostic and therapeutic evaluation of AL amyloidosis patients.

Clinical Characteristics and Treatment Outcome of Waldenstrom Macroglobulinemia: Experience from a Tertiary Cancer Centre

Background: Waldenstroms macroglobulinemia (WM) or Lymphoplasmacytic lymphoma is a rare B cell lymphoproliferative malignancy characterized by serum monoclonal immunoglobulin M (IgM) protein and lymphoplasmacytic infiltration in the bone marrow. Patients usually present in their seventh decade with symptoms related to the infiltration of the marrow or the effects of monoclonal IgM in the blood. Objective: Was to study the clinical characteristics and treatment outcome of patients diagnosed with WM. Materials and Methods: This is a retrospective analysis of 26 cases of WM treated at a tertiary cancer centre. Results: WM constituted 0.68% of our nonHodgkin’s lymphoma. The median age at presentation was 67 years with a male to female ratio of 2:1. Four patients had lymphadenopathy, six had splenomegaly and four patients had evidence of hyper viscosity. All patients had IgM paraproteinemia, the M band was IgM-kappa in 19 patients and IgM-lambda in seven patients. All patients had a histopathological confirmation. According to IPSS-WM, 12 patients each were in the intermediate and high risk group and 2 in the low risk group. Among the 26 patients, 20 patients received upfront treatment and four patients were kept under observation. The indications for treatment were cytopenia in ten patients, constitutional symptoms in five, hyper viscosity in four and symptomatic lymphadenopathy in one patient. Sixteen patients received rituximab-based chemotherapy. Four patients with features with hyper viscosity underwent plasmapheresis. The three year progression free survival and overall survival were 69.4% and 78 % respectivel. Conclusions: WM is a low grade B cell lymphoproliferative disorder having an indolent course and do not require treatment for prolonged period. Diagnosis is challenging due to lack of distinct diagnostic features. Rituximab containing regimens are the standard of care. Newer targeted treatment options may improve the outcome of this incurable disease.

Endogenous monoclonal immunoglobulins analyzed using the EXENT® solution and LC-MS

IntroductionThe EXENT® Solution, a fully automated system, is a recent advancement for identifying and quantifying monoclonal immunoglobulins in serum. It combines immunoprecipitation with MALDI-TOF mass spectrometry. Compared to gel-based methods, like SPEP and IFE, it has demonstrated the ability to detect monoclonal immunoglobulins in serum at lower levels. In this study, samples that tested negative using EXENT® were reflexed to LC-MS to determine if the more sensitive LC-MS method could identify monoclonal immunoglobulins missed by EXENT®. ObjectivesTo assess whether monoclonal immunoglobulins that are not detected by EXENT® can be detected by LC-MS using a low flow LC system coupled to a Q-TOF mass spectrometer. MethodsSamples obtained from patients confirmed to have multiple myeloma (MM) were diluted with pooled polyclonal human serum and analyzed using EXENT®. If a specific monoclonal immunoglobulin was not detected by EXENT®, the sample was then subjected to analysis by LC-MS. For the LC-MS analysis, the sample eluate, obtained after the MALDI-TOF MS spotting step, was collected and transferred to an autosampler tray for subsequent analysis using LC-MS. ConclusionLC-MS has the capability to detect monoclonal immunoglobulins that are no longer detected by EXENT®. Reflexing samples to LC-MS for analysis does not involve additional sample handling, allowing for a faster time-to-result compared to current approaches, such as Next-Generation Sequencing, Next-Generation Flow, and clonotypic peptide methods. Notably, LC-MS offers equivalent sensitivity in detecting these specific monoclonal immunoglobulins.

Mutational Landscape of Bone Marrow CD19 and CD138 Cells in Waldenström Macroglobulinemia (WM) and IgM Monoclonal Gammopathy of Undetermined Significance (IgM MGUS)

Background Waldenström's Macroglobulinemia (WM) is an incurable B-cell neoplasm characterized by serum monoclonal immunoglobulin M (IgM) and clonal lymphoplasmacytic cells infiltrating the bone marrow (BM). Smoldering WM (sWM) is the asymptomatic/indolent form with a high risk of progressing to symptomatic WM requiring treatment whereas IgM monoclonal gammopathy of undetermined significance (IgM MGUS) is an early precursor stage of WM. Despite recurrent and activating mutations, including MYD88, CXCR4, ARID1A, KMT2D, and CD79B have been identified, the genetic basis for WM and the risk of progression of IgM MGUS to WM remain to be fully elucidated. Methods Based on the mutational profiling of WM and our previous wide transcriptome analyses on BM CD19+ and CD138+ cells of patients with WM and IgM MGUS, we decided to investigate the mutation status of these patients as follows: WM (n=10), sWM (n=7), and IgM MGUS (n=5), by performing high throughput targeted AmpliSeq next generation sequencing on 117 target genes potentially involved in B-cell lymphomagenesis and in the risk of progression of IgM MGUS to WM (Trojani A. et al. Cancers, 2021). Results We identified genetic aberrations in multiple genes, including previously described classic mutations in MYD88 (L260P alias L265P) found in 82.3% (14/17) of WM/sWM patients and in 60% (3/5) of IgM MGUS subjects. The CXCR4 mutation (S338Ter) was detected in 23.5% of CD19+ cells of WM patients, whereas it was absent in IgM MGUS subjects. Interestingly, we also identified new mutated genes, including WNK2 somatic mutations affecting 47% of WM patients, for which a recurrent allelic variant (V1635Ter) was observed, and BCL9 with a recurrent frameshift variant (P516Lfs) identified in 29.4% of WM patients exclusively in the CD19 cell fraction. The BCL9 (P516Lfs) variant was also detected in the B cells of one IgM MGUS patient. Moreover, sequencing evaluation revealed recurrently frameshift or missense mutations involving NFKB2 (L473Afs), PTPN13 (P1546Tfs), CARD11 (S622del), KMT2C (I823T), and ATM in WM and IgM MGUS patients (Figure 1). Of interest, KMT2C (also known as MLL3) exhibited a somatic hypermutated phenotype in both CD19+ and CD138+ cells of WM and IgM MGUS samples. The KMT2C mutations were not mutually exclusive with those involving KMT2D in our cohort of patients. Conclusion The present genomic analysis revealed new insights into the mutational landscape of Waldenström Macroglobulinemia, providing evidence of the recurrence of some variants in subjects with IgM MGUS as well. Notably, we uncovered new genetic mutations with somatic variants recurrently observed (n≥3) in KMT2C, WNK2, BCL9, PTPN13, CARD11 and NFKB2 genes. These recurring somatic events provide a genomic basis for future research and a better understanding of the WM pathogenesis as well as the risk of transformation of IgM MGUS to WM.

Outcomes of Septuagenarians and Octogenarians with Monoclonal Gammopathy of Undetermined Significance Admitted for Acute Pulmonary Embolism

Introduction: Monoclonal gammopathy of undetermined significance (MGUS) is clinically defined with a serum monoclonal immunoglobulin of less than 3 g/dL and the presence of less than 10% of plasma cells in the bone marrow, in the absence of any end-organ damage that is typically seen in patients with multiple myeloma. The incidence of MGUS mostly involves the elderly, with a median age estimated at 70 years. While some studies have linked MGUS with a hypercoagulable state, there are no large studies investigating the impact and outcomes of MGUS in the elderly with pulmonary embolism(PE). We thus propose a national analysis to investigate. Methods: Our retrospective study via the National Inpatient Sample (NIS) focused on adults admitted with a primary diagnosis of Acute Pulmonary Embolism. We excluded all cases of ages <70 years and >90 years. Patient characteristics in acute PE cases with and without MGUS such as age, sex, race, and presence of various comorbidities such as diabetes, hypertension, obesity, among many, were explored via Chi-Square tests, t-tests, and Mann-Whitney U tests as appropriate. Differences in outcomes for PE in MGUS and non-MGUS patients were estimated via logistic regression models on SPSS 29.0. A p-value of <0.05 was considered as statistically significant. To comply with the database provider's (HCUP) guidelines, we did not report values <11. Results: Among the 327090 septuagenarians and octogenarians admitted primarily for acute PE in our study, there were 1095 (0.3%) patients with a diagnosis for MGUS that also reported a higher mean Charlson Comorbidity Index (CCI) score (4.26 vs. 2.90, p<0.01) (Table 1). While 51.1% of MGUS cases with acute PE were males, the non-MGUS group mainly consisted of females (56.1%, p<0.01). Medicare covered 91.3% and 90.8% of all MGUS and non-MGUS cases in our study, and 68.9% and 66.3% of all admissions involved Urban-teaching centers, respectively. Around 11.0% of MGUS cases also had a history of prior pulmonary embolism, while it was 8.0% in the non-MGUS cohort (p<0.01). The MGUS cohort also showed a lower prevalence of hypertension (41.1% vs.46.7%, p<0.01) while having a higher prevalence of peripheral vascular disease (11.4% vs. 5.1%, p<0.01), smoking status (40.6% vs. 37.8%, p=0.049), lipid disorders (56.6% vs. 50.6%, p<0.01), and chronic kidney disease (35.6% vs. 20.0%, p<0.01) than the non-MGUS group. MGUS patients showed lower odds of experiencing in-patient events such as myocardial infarction (aOR 0.345, 95% CI 0.221-0.538, p<0.01), acute kidney injury (aOR 0.843, 95% CI 0.719-0.990, p=0.037), and respiratory compromise (aOR 0.749, 95% CI 0.652-0.860, p<0.01). While a higher odds of cardiogenic shock (aOR 1.903, 95% CI 1.316-2.753, p<0.01) was also noted, patients with and without MGUS had no differences related to in-hospital mortality (aOR0.721, 95% CI 0.513-1.013, p=0.060) or the need for IVC filters (aOR 1.156, 95% CI 0.924-1.446, p=0.206). MGUS patients also experienced a longer stay (mean length of stay 4.91 vs. 4.62 days, p<0.01). Conclusion: This nationwide data analysis suggests a higher prevalence of a history of previous PE in MGUS cases admitted for PE than those without MGUS. Furthermore, while odds of events involving in-patient myocardial infarction, acute kidney injury, or respiratory compromise in MGUS patients were lower, they had a higher risk of cardiogenic shock. It is thus vital for physicians to be on the watch-out of such complications and provide adequate testing and care in a timely manner to improve their prognosis and outcomes.

The Synthetic Collagen-Binding Peptide NIPEP-OSS Delays Mouse Myeloma Progression.

Multiple myeloma (MM) is the second most common hematological malignancy. It is a clonal B-cell disorder characterized by the proliferation of malignant plasma cells in the bone marrow, the presence of monoclonal serum immunoglobulin, and osteolytic lesions. An increasing amount of evidence shows that the interactions of MM cells and the bone microenvironment play a significant role, suggesting that these interactions may be good targets for therapy. The osteopontin-derived collagen-binding motif-bearing peptide NIPEP-OSS stimulates biomineralization and enhances bone remodeling dynamics. Due to its unique targeted osteogenic activity with a broad safety margin, we evaluated the potential of NIPEP-OSS for anti-myeloma activity using MM bone disease (MMBD) animal models. In a 5TGM1-engrafted NSG model, the survival rates of the control and treated groups were significantly different (p = 0.0014), with median survival times of 45 and 57 days, respectively. The bioluminescence analyses showed that myeloma slowly developed in the treated mice compared to the control mice in both models. NIPEP-OSS enhanced bone formation by increasing biomineralization in the bone. We also tested NIPEP-OSS in a well-established 5TGM1-engrafted C57BL/KaLwRij model. Similar to the previous model, the median survival times of the control and treated groups were significantly different (p = 0.0057), with 46 and 63 days, respectively. In comparison with the control, an increase in p1NP was found in the treated mice. We concluded that NIPEP-OSS delays mouse myeloma progression via bone formation in MMBD mouse models.

Isolated bone marrow amyloid light chain amyloidosis coexisting with multiple myeloma.

A 62-year-old woman was diagnosed with Bence-Jones protein multiple myeloma in another hospital. She referred sternum pain and a clavicle fracture. Nuclear magnetic resonance showed multiple lytic lesions in clavicle, rib, vertebra, humerus and sternum. She was referred to our hospital because of an inadequate response to treatment. Complete blood count, kidney function, calcium, lactate dehydrogenase and beta-2-microglobulin did not show abnormalities. Monoclonal immunoglobulins in serum were not detected. Immunofixation in serum and urine was positive for kappa light chain. A bone marrow aspirate was performed in the sternum. The sternal aspirate showed a patchy infiltration by plasma cells (CD19–, CD56+, CD117+) of variable size, some of them binucleated with a cytoplasm with a flammig appearance. Nebulous amorphous basophilic protein material suggestive of amyloid was also observed (Figure 1a). Congo red stain was positive, and the microscope fluorescence showed apple-green birefringence under polarized light and confirmed the suspicion (Figure 1b). Fluorescence in situ hybridisation (FISH) showed the t (11;14) (q13;q32); the TP53 was normal. The study in the fat tissue did not show amyloid and the renal and heart functions were normal. The patient was diagnosed of amyloid light chain amyloidosis associated with multiple myeloma and started treatment with bortezomib, lenalidomide and dexamethasone. The response was suboptimal, and daratumumab was added to the treatment. This case demonstrates the importance of recognizing the presence of amyloid in bone marrow aspiration. This diagnosis is relevant for the prognostic and therapeutic implication. All authors have contributed equally to the manuscript. None The authors declare they have no conflicts of interest. The authors received no specific funding for this work. A written informed consent was obtained from the patient.

Pathological characteristics of light chain crystalline podocytopathy

Monoclonal immunoglobulin light chain (LC) crystalline inclusions within podocytes are rare, poorly characterized entities. To provide more insight, we now present the first clinicopathologic series of LC crystalline podocytopathy (LCCP) encompassing 25 patients (68% male, median age 56 years). Most (80%) patients presented with proteinuria and chronic kidney disease, with nephrotic syndrome in 28%. Crystalline keratopathy and Fanconi syndrome were present in 22% and 10%, respectively. The hematologic condition was monoclonal gammopathy of renal significance (MGRS) in 55% and multiple myeloma in 45%. The serum monoclonal immunoglobulin was IgG κappa in 86%. Histologically, 60% exhibited focal segmental glomerulosclerosis (FSGS), often collapsing. Ultrastructurally, podocyte LC crystals were numerous with variable effacement of foot processes. Crystals were also present in proximal tubular cells as light chain proximal tubulopathy (LCPT) in 80% and in interstitial histiocytes in 36%. Significantly, frozen-section immunofluorescence failed to reveal the LC composition of crystals in 88%, requiring paraffin-immunofluorescence or immunohistochemistry, with identification of kappa LC in 87%. The LC variable region gene segment, determined by mass spectrometry of glomeruli or bone marrow plasma cell sequencing, was IGKV1-33 in four and IGKV3-20 in one. Among 21 patients who received anti-plasma cell-directed chemotherapy, 50% achieved a kidney response, which depended on a deep hematologic response. After a median follow-up of 36 months, 26% progressed to kidney failure and 17% died. The mean kidney failure-free survival was 57.6 months and was worse in those with FSGS. In sum, LCCP is rare, mostly associates with IgG κappa MGRS, and frequently has concurrent LCPT, although Fanconi syndrome is uncommon. Paraffin-immunofluorescence and electron microscopy are essential to prevent misdiagnosis as primary FSGS since kidney survival depends on early diagnosis and subsequent clone-directed therapy.

A series of heterogeneous lymphoproliferative diseases with CD3 and MUM1 co-expressed in cats and dogs.

Lymphoma diagnosis in dogs and cats is continually evolving as new subtypes and human correlates are being recognized. In humans, T-cell lymphomas with MUM1 expressed and plasma cell neoplasia or B-cell lymphomas with CD3 expressed aberrantly are reported only rarely. We report here a case series of tumors in dogs and cats with CD3 and MUM1 co-expressed as determined by immunocytochemistry or immunohistochemistry. Lineage was assigned for these tumors by 3 board-certified pathologists and a veterinary immunologist based on review of clinical and cellular features and the results of ancillary testing including PCR for antigen receptor rearrangements, flow cytometry, and serum protein electrophoresis with immunofixation. In cats, 7 of 7 tumors, and in dogs, 3 of 6 tumors with CD3 and MUM1 co-expressed had clonal rearrangement of the immunoglobulin gene or serum monoclonal immunoglobulin, consistent with a diagnosis of a plasma cell neoplasia or myeloma-related disorder with CD3 expressed aberrantly. Disease was often disseminated; notably, 3 of 7 feline cases had cutaneous and/or subcutaneous involvement in the tarsal area. In dogs, 3 of 6 cases had a clonal T-cell receptor gamma result and no clonal immunoglobulin gene rearrangement and were diagnosed as a T-cell tumor with MUM1 expressed. The use of multiple testing modalities in our series of tumors with plasma-cell and T-cell antigens in dogs and cats aided in the comprehensive identification of the lymphoproliferative disease subtype.

The deubiquitinase OTUD1 regulates immunoglobulin production and proteasome inhibitor sensitivity in multiple myeloma

Serum monoclonal immunoglobulin (Ig) is the main diagnostic factor for patients with multiple myeloma (MM), however its prognostic potential remains unclear. On a large MM patient cohort (n = 4146), we observe no correlation between serum Ig levels and patient survival, while amount of intracellular Ig has a strong predictive effect. Focused CRISPR screen, transcriptional and proteomic analysis identify deubiquitinase OTUD1 as a critical mediator of Ig synthesis, proteasome inhibitor sensitivity and tumor burden in MM. Mechanistically, OTUD1 deubiquitinates peroxiredoxin 4 (PRDX4), protecting it from endoplasmic reticulum (ER)-associated degradation. In turn, PRDX4 facilitates Ig production which coincides with the accumulation of unfolded proteins and higher ER stress. The elevated load on proteasome ultimately potentiates myeloma response to proteasome inhibitors providing a window for a rational therapy. Collectively, our findings support the significance of the Ig production machinery as a biomarker and target in the combinatory treatment of MM patients.

Retinal oxygen saturation in monoclonal gammopathies patients: Apilot study.

The aim of this pilot study was to assess oxygen saturation in retinal blood vessels in patients with monoclonal gammopathies (MGs). Thirty-one patients with MGs (11 women and 20 men, mean age 65.9± 8.9 years) were enrolled during 2016-2020. The patients were diagnosed at the Haemato-Oncology Department and subsequently examined at the Ophthalmology Department before initiating systemic therapy. All patients were subjected to automatic retinal oximetry (Oxymap ehf.) and had their fundus photographed (Topcon TRC-50DX retinal camera). We assessed the association between retinal oxygen saturation (SatO2 ) - arterial SatO2 , venous SatO2 and arterio-venous (AV) difference-and MGs parameters: serum monoclonal immunoglobulin (M-protein) level and serum immunoglobulin-free light chains (FLC kappa and lambda), total protein, serum viscosity, haemoglobin, albumin, lactate dehydrogenase, C-reactive protein, creatinine and serum calcium level. Hyperviscosity-related retinopathy was also evaluated. Statistical analysis showed a significant positive correlation (r=0.462; p=0.009) between the AV difference and the haemoglobin level. A significant, medium strong negative correlation was found between the AV difference and the serum levels of the monoclonal light lambda chains (r=-0.450; p=0.011). Contrary to expectations, no statistically significant correlation was found between retinal oxygen saturation and the total protein or viscosity. This study found correlation between retinal oxygen saturation and certain parameters in the blood of patients with MGs. Increasing levels of monoclonal immunoglobulin seem to reduce oxygen absorption in retinal arterioles, resulting in a lower AV difference, particularly in patients with a high free light chain level.

Does Waldenstrom’s macroglobulinemia also cause bone destruction? A rare case report

Waldenstrom’s macroglobulinemia (WM) is a rare type of malignant B-cell lymphoma. The main feature of WM is elevated serum monoclonal immunoglobulin M, similar to multiple myeloma (MM). Unlike in MM, the rarity of destructive bone lesions in WM has been repeatedly emphasized. We report a unique case of WM with a vertebral compression fracture as the first symptom. This case highlights that the presence or absence of bone destruction may not clearly distinguish between WM and MM. The possibility of WM should be considered in patients with vertebral fracture and destruction as the first presentation. Performing vertebral bone marrow aspiration biopsy during percutaneous vertebroplasty is a convenient and effective method to assist in the diagnosis of WM.

Clinicopathological Spectrum of Cryoglobulinemic Glomerulonephritis without Evidence of Autoimmunity Disorders: A Retrospective Study from a Single Institute of China

Background: Cryoglobulinemic glomerulonephritis (Cryo-GN), caused by circulating cryoglobulins, has varied etiology and clinical-pathologic manifestations. This study aimed to investigate the clinicopathological spectrum and outcome of patients with various Cryo-GN in China. Methods: A retrospective review of 74 Chinese patients with biopsy-proven cryoglobulin-related renal lesions in Peking University First Hospital from 2010 to 2020 was performed. Results: The mean age at diagnosis was 52.9 ± 15.0 years, and the female-to-male ratio was about 2/5. For the etiology screening, serum/urine monoclonal immunoglobulin could be detected on immunofixation electrophoresis in 34% of patients, including 6 patients who had hematological malignancies. Fifty-seven percent of patients had HBV infection, far more than HCV infection (5%). Ten percent of patients had other infections, and 27% of patients were classified as essential or idiopathic. Eleven out of the 15 patients with type II cryoglobulinemia had a consistent monotype of serum monoclonal immunoglobulins and monoclonal cryoprecipitate. The clinical manifestations were similar between various types of cryoglobulinemia. Hematuria, proteinuria, hypertension, anemia, and chronic renal insufficiency were the most common features. Fifty-three percent of patients presented with nephrotic syndrome, and 32% experienced acute kidney injury. Hypocomplementemia, serum-positive rheumatoid factor activity, and skin lesions were reported in 45%, 29%, and 28% of patients, respectively. After a median of 24 months follow-up, 18 patients reached end-stage kidney disease. The clone-targeted treatment could retard the renal deterioration compared with immunosuppressive therapy. Conclusions: This was the largest single-center, clinicopathological retrospective study of Cryo-GN in China. Our data strongly support the association between monoclonal gammopathy and type II Cryo-GN. The renal responsive rate of immunosuppressant therapy is still suboptimal. The clone-targeted treatment shows promising effects in patients with type I or II Cryo-GN.

Waldenström macroglobulinemia.

Waldenström macroglobulinemia (WM) is a hematological malignancy; it is a monoclonal gammopathy, a disease characterized by presence of a monoclonal immunoglobulin in serum and/or urine. The median age at dia-gnosis is 71 years. WM is not an aggres-sive disease and patients with this dia-gnosis can live for several years. Infiltration of the bone marrow with lymphoplasmacytoid cells causes anemia, leading to various problems, mainly fatigue. Hepatomegaly, splenomegaly and lymphadenopathy can also occur. Hyperviscosity syndrome can appear and is caused by excessive production of immunoglobulin M. A mutation in MYD88 gene is detected in almost every WM patient, and in almost one third of them, a mutation in CXCR4 gene is detected. The detection of MYD88 mutation is important for a correct therapeutic strategy, since a Brutons tyrosine kinase inhibitor, ibrutinib, is most effective in patients with mutated MYD88 and wt CXCR4. The therapy is started when first symptoms occur. The aim of this study is to summarize current knowledge about this disease, its dia-g-nostics, molecular basis and treatment.

RNA-based immunoglobulin repertoire sequencing is a new tool for the management of monoclonal gammopathy of renal (kidney) significance

The diagnostic approach of monoclonal gammopathy of renalsignificance is based on the detection of a monoclonal immunoglobulin in the blood and urine, and the identification of the underlying clone through bone marrow and/or peripheral blood cytologic and flow cytometry analysis. However, the monoclonal component and its corresponding clone may be undetectable using these routine techniques. Since clone identification is the cornerstone for guiding therapy and assessing disease response, more sensitive methods are required. We recently developed a high-throughput sequencing assay from bone marrow mRNA encoding immunoglobulins (RACE-RepSeq). This technique provides both full-length V(D)J region (variable, diversity andjoining genes that generate unique receptors as antigen receptors) of the monoclonal immunoglobulin and the dominant immunoglobulin repertoire. This allows analysis ofmutational patterns, immunoglobulin variable gene frequencies and diversity due to somatic hypermutation. Here, we evaluated the diagnostic performance of RACE-RepSeq in 16 patients with monoclonal-associated kidney lesions, and low serum monoclonal immunoglobulin and free light chain levels at diagnosis. Bone marrow immunohistochemical analysis was negative in all 11 patients so tested and 7 of 12 patients had no detectable clone matching the kidney deposits using flow cytometry analysis. By contrast, RACE-RepSeq detected a dominant clonal light chain sequence of matched isotype with respect to kidney deposits in all patients. Thus, high throughput mRNA sequencing appears highly sensitive to detect subtle clonal disorders in monoclonal gammopathy of renal significance and suggest this novel approach could help improve the management of this kidney disease.

Animal Models of Multiple Myeloma Bone Disease.

Multiple myeloma (MM) is a clonal B-cell disorder characterized by the proliferation of malignant plasma cells (PCs) in the bone marrow, the presence of monoclonal serum immunoglobulin, and osteolytic lesions. It is the second most common hematological malignancy and considered an incurable disease despite significant treatment improvements. MM bone disease (MMBD) is defined as the presence of one or more osteolytic bone lesions or diffused osteoporosis with compression fracture attributable to the underlying clonal PC disorder. MMBD causes severe morbidity and increases mortality. Cumulative evidence shows that the interaction of MM cells and bone microenvironment plays a significant role in MM progression, suggesting that these interactions may be good targets for therapy. MM animal models have been developed and studied in various aspects of MM tumorigenesis. In particular, MMBD has been studied in various models, and each model has unique features. As the general features of MM animal models have been reviewed elsewhere, the current review will focus on the features of MMBD animal models.

Immunofluorescence staining for immunoglobulin heavy chain/light chain on kidney biopsies is a valuable ancillary technique for the diagnosis of monoclonal gammopathy-associated kidney diseases

Heavy chain/light chain (HLC) antibodies target conformational epitopes at the junctions of the heavy chain and light chain constant regions (CH1 and CL) of serum IgGκ, IgGλ, IgAκ, IgAλ, IgMκ, and IgMλ to provide quantitation of intact HLC pairs. Here, we developed an HLC tissue immunofluorescence protocol to test if it can complement conventional immunofluorescence in the diagnosis of monoclonal gammopathy-associated kidney diseases. HLC immunofluorescence was performed on archived frozen tissue of 104 kidney biopsies. The sensitivity and specificity of HLC immunofluorescence was confirmed by testing cases of lupus nephritis, other polyclonal immunoglobulin nephropathies, and light chain nephropathies (light chain amyloidosis and deposition disease). Testing of ten cases of the IgG variant of proliferative glomerulonephritis with monoclonal immunoglobulin deposits excluded monoclonal deposits in two by revealing positivity for IgGκ and IgGλ. Testing of 12 cases of monotypic IgA nephropathy excluded monoclonal deposits in six by revealing staining for IgAκ and IgAλ. Testing of six cases of monotypic fibrillary glomerulonephritis excluded monoclonal deposits in three by revealing positivity for IgGκ and IgGλ. None of 14 cases of glomerulonephritis in which HLC immunofluorescence unmasked polytypic deposits were associated with a serum or urine monoclonal immunoglobulins matching the conventional immunofluorescence results. HLC immunofluorescence outperformed paraffin immunofluorescence and IgG subclass staining in 10/13 (77%) of cases. Testing of 18 cases of cryoglobulinemic glomerulonephritis showed better correlation with serum cryoprecipitate immunofixation than conventional immunofluorescence with regards to the type of cryoglobulin in 47% of cases. Thus, HLC immunofluorescence is a valuable ancillary technique in kidney pathology for the diagnosis of monoclonal gammopathy-associated nephropathies, and could be utilized to confirm or exclude the monoclonal nature of deposits.

C3 glomerulonephritis associated with monoclonal gammopathy: a retrospective case series study from a single institute in China

Objective To investigate the demographic and clinicopathological features and renal outcomes of Chinese patients with C3 glomerulonephritis in the setting of monoclonal gammopathy. Methods Patients with renal biopsy-proven C3 glomerulonephritis and detectable serum and/or urine monoclonal immunoglobulin from 2006 to 2018 in Peking University First Hospital were included, their clinical data, renal pathology type, treatment, and prognosis were collected and analyzed. Results Nineteen patients were enrolled, accounting for 24% of C3GN patients in the study period. The mean age of onset was 55 years old and the gender ratio was 4/15 (female/male). The mean eGFR at biopsy was 49.55 ± 29.81 ml/min/1.73m2. The prominent clinical manifestations included nephrotic syndrome (58%), anemia (68%), microscopic hematuria and leukocyturia (58%), and hypocomplementemia (13, 68%). The IgG was the most common isotype of monoclonal Ig on immunofixation electrophoresis. Kidney biopsies revealed a relatively prominent MPGN pattern. Only two patients had direct evidence of monocle immunoglobulins acting as C3GN pathogenic factors. Two patients had concurrent TMA-like renal injuries. The median renal survival was 12 and 15 months, respectively in patients receiving conservative therapy and immunosuppressant therapy, without statistical significance. The efficacy of clone-targeted therapy needed further investigation. Plasma exchange therapy only improved one patient’s renal outcome. Conclusions This is the first case series report of C3GN combined with monoclonal Ig in northern China. The renal prognosis of these patients is poor, and immunosuppressant therapies show no advantage over supportive therapy in renal prognosis, while the benefit of clone-targeted chemotherapy is still requiring investigation.

Efficacy and Safety of Regimens Used for the Treatment of POEMS Syndrome- a Systematic Review

Introduction: POEMS syndrome (polyneuropathy, organomegaly, endocrinopathy/edema, myeloma protein, skin changes) is a rare disorder associated with plasma cell neoplasia where an overproduction of proinflammatory cytokines such as vascular endothelial growth factor (VEGF) upregulates vasculogenesis and increases vascular permeability resulting in multiple symptoms of the disease e.g. edema and hemangioma. Although the limited-stage disease can be treated with radiotherapy, treatment for the more advanced disease remains unclear. Here we offer a systematic review of the efficacy and safety of treatment regimens used to treat POEMS syndrome in the adult population. Methods: Databases i.e. PubMed, Embase, Web of Science, and clinicaltrials.gov were searched since inception through May 15th, 2020, following the PRISMA guidelines. Out of 421 studies, 8 articles (5 prospective clinical trials, and 3 retrospective studies) were included. Phase I clinical trials, abstracts, case reports, case series, and review articles were excluded. Results: A total of 171 patients (21-79 years of age) were evaluated in 8 clinical studies. Immunomodulator Based Regimens: In the 5 clinical studies (N=96) where lenalidomide (Len) plus dexamethasone (Dex) were used, the hematological (heme) complete response (CR) ranged from 20% to 46%. Heme CR was defined as the absence of monoclonal immunoglobulins in serum or urine. VEGF CR defined as normalization of serum VEGF levels ranged from 35% to 80%. Neurological response defined as the improvement in the overall neuropathy limitations scale (ONLS) by at least one score ranged from 0% to 95%. These studies reported 2-3 year progression-free survival (PFS) and overall survival (OS) of 59% to 92% and 90% to 100%, respectively. In the study by Suichi et al. (N=5), VEGF CR was reported to be 80%. However, no neurological improvement was seen in these patients. Proteasome Inhibitor Based Regimens: In the study by He et al. (N=20) bortezomib in combination with cyclophosphamide and Dex yielded heme, VEGF, and neurological responses in 76%, 88%, and 95% of the patients, respectively. No grade ≥3 treatment-related adverse effects (TRAE) were seen. Conventional Chemotherapy Based Regimens: In the phase II trial conducted by Misawa et al. (N=25) the reduction in serum VEGF levels, as well as clinical response as measured by improvement in muscle strength, was higher in the thalidomide arm as compared to the placebo arm. Grade ≥3 TRAE reported in the treatment arm were cardiac arrest, heart failure, and dehydration in 1 (8%) patient each. In the study by Li et al. (N=31) melphalan (Mel) in combination with Dex achieved heme, VEGF, and neurological response rates of 81%, 100%, and 100%, respectively. Twenty percent of the patients experienced grade ≥3 TRAE. Conclusion: Immunomodulator and chemotherapy based regimens appear to have a reasonable safety and efficacy profile. Lenalidomide and melphalan based therapies were most effective. Proteasome inhibitors based regimens yielded efficacious results without significant toxicity. More studies with existing and newer combinations with autologous stem cell transplantation are warranted. Disclosures Anwer: Incyte, Seattle Genetics, Acetylon Pharmaceuticals, AbbVie Pharma, Astellas Pharma, Celegene, Millennium Pharmaceuticals.: Honoraria, Research Funding, Speakers Bureau.

Severe but reversible pulmonary hypertension in scleromyxedema and multiple myeloma: a case report

BackgroundScleromyxedema is a progressive, systemic connective tissue disorder characterized by fibro-mucous skin lesions and increased serum monoclonal immunoglobulin levels. Pulmonary involvement occurs in a subset of patients, though the overall prevalence of pulmonary lesions in scleromyxedema is unknown. Since pulmonary hypertension presumably occurs in these patients due to disease progression and development of additional conditions, treatment of the underlying plasma cell dyscrasia and connective tissue disorder may improve pulmonary hypertension symptoms.Case presentationAn elderly patient with scleromyxedema developed pulmonary hypertension refractory to vasodilator and diuretic therapy and subsequently multiple myeloma that responded to a combination therapy of bortezomib, cyclophosphamide, and dexamethasone treatment.ConclusionsTreatment of the underlying disease(s) that contributed to pulmonary hypertension development with anti-neoplastic agents like bortezomib may improve cardiopulmonary symptoms secondary to reducing abnormal blood cell counts and paraprotein levels.