Lambda Light Chain Type Research Articles

#6394 ACUTE KIDNEY INJURY IN MYELOMA WITH HIGH LIGHT CHAIN PRODUCTION: A 10-YEAR EXPERIENCE

Abstract Background and Aims In multiple myeloma (MM), a high serum light chain (LC) concentration significantly increases the probability of developing cast nephropathy. The use of high cut-off (HCO) hemodialysis (HD) membranes has not been clearly shown to be effective in 2 large European studies (MYRE and EuLITE) in terms of improved prognosis. Therefore, their use is currently controversial. We want to review our experience, in the last 10 years, of patients with Acute Kidney Injury (AKI) in the context of MM in progression with high circulating LC: whether or not any HD technique has been performed and what factors may have had an impact on kidney and patient survival. Method We reviewed all cases of MM flare or progression, with LC >1000 mg/L and AKI, evaluated by our service between May 2012 and May 2022 (N = 20). We collected data on baseline renal function, laboratory tests, type and amount of light chain and monoclonal component, whether or not HD was performed, type of technique, evolution of kidney function and mortality. Results We counted 60% males, median age of 76.5 years (69.5-83.0), and baseline creatinine of 1.30 (0.94-1.66) with an estimated glomerular filtration rate (eGFR) of 52 mL/min (35-58). At the time of AKI, the median obtained data were: peak creatinine 4.6 mg/dL (4.1-6.0), Hb 8.8 g/dL (8.3-11.6), albumin 3.6 mg/dL (3.1-4.1), calcemia 10.1 mg/dL (9.5-11.5), blood monoclonal component of 0.80 mg/dL (0.36-2.22) and serum LC of 5. 480 mg/L (2,143-11,825). In 9 cases the myeloma was CL kappa-producing and in 11 cases CL lambda-producing. In 9 of the 20 cases (45%) the patient died within 6 months. HD was performed in 11 cases: 7 with HCO membrane, 2 with polymethylmethacrylate (PMMA) and 2 with conventional technique. Of the 7 patients dialyzed with HCO, 3 died in <6 months (43%) and 2 of them (29%) were also HD-dependent at the time of death. The 2 patients dialyzed with PMMA had eGFR>30 mL/min at 6 months after the episode. In contrast, the 2 patients who underwent conventional HD died in <6 months without recovery of renal function. Of the 9 patients who did not undergo HD, 4 had optimal recovery of renal function (eGFR>45 mL/min) but 2 of them died in <6 months (50%). Of the remaining 5: 2 died in <6 months (40%), progression to end-stage CKD was observed in 2 cases (HD was not performed and one of them died in <1 year), and the remaining patient remains alive with eGFR>20 mL/min at 2 years of follow-up. Concerning chemotherapy (QT), 13 patients were treated with bortezomib (12 in 1st line), 4 with lenalidomide and only 1 case was not treated. Of 8 patients who achieved remission with chemotherapy (40% of the total), 7 remained alive at 2 years of follow-up (87.5% of them) and did not require HD, and only 1 of them died at 1 year after a new recurrence (despite HD being implemented). Although statistical significance was not reached due to the small sample size, we did observe a statistical trend towards a higher risk of death at 6 months in patients with peak creatinine >5 mg/dL (p = 0.078), albuminemia<3.5 mg/dL (p = 0.078) and lambda-type LC (p = 0.064). Conclusions The use of HCO membranes has not seemed to influence kidney and patient prognosis. In contrast, remission after chemotherapy appears to be the most important determinant of patient survival. Recovery of kidney function was not related in this review to increased patient survival. Analytical parameters such as peak creatinine, hypoalbuminemia and type and amount of circulating LC could be studied as prognostic factors in this entity.

Abstract 6073: Distinct subtype of multiple myeloma revealed by whole genome and transcriptome sequencing

Abstract Background: Advanced sequencing technologies have contributed to identifying genomic and transcriptomic features of various types of tumors. Despite several sequencing data from multiple myeloma (MM) being generated, understanding various properties of MM remained insufficient. Methods: The patients confirmed with MM by bone marrow examination were analyzed in the study. Myeloma cells were enriched from the bone marrow aspirates of the patients using CD138. Whole genome sequencing (WGS) and whole transcriptome sequencing (WTS) were performed for isolated myeloma cells. Mutational signatures were analyzed using somatic mutations detected by WGS. Results: A total of 37 MM patients (median age 67) were enrolled in this study. WGS revealed somatic mutations of an average of 8872 single nucleotide variants (SNVs) and 934 insertions and deletions (Indels) from the patients, respectively. By the analysis of structural variants, 7 chromothripsis and 4 chromoplexy patients were identified. Mutational signature analysis showed that single base substitution (SBS) 9 signature largely varied between patients. SBS9 signature high patients were mostly diagnosed with non-IgG/non-IgA heavy chain and lambda light chain type myeloma. Notably, IgD myeloma patients were solely detected in the SBS9 signature high group compared to the low group (p = 0.001). Gene expression patterns obtained from WTS were well-divided into SBS9 signature high and low groups, and analysis of differentially expressed genes (DEG) between SBS9 signature high and low groups implied IgD myeloma features including high expression of IGHD. As a validation test, we tried mutational signature analysis on whole exome sequencing (WES) data of 784 MM patients from the MMRF database. Of the 38 patients with more than 500 exonic mutations, only the SBS9 signature high group had IgD myeloma candidates, whereas the SBS9 signature low group had no IgD myeloma candidate (p = 0.014). Conclusions: DNA and RNA sequencing of myeloma patients could classify specific subtypes of disease categories. Further study for SBS 9 signature high and IgD myeloma is needed to discover the underlying mechanism of distinct features and to find out their clinical implication. Citation Format: Sheehyun Kim, Hyundong Yoon, Youngil Koh, Sung-Soo Yoon. Distinct subtype of multiple myeloma revealed by whole genome and transcriptome sequencing. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6073.

Preliminary exploration of the metabolic profile and metabolic pathways in newly diagnosed multiple myeloma

Objective: To explore the metabolite profile and metabolic pathways of newly diagnosed multiple myeloma (MM). Methods: Gas chromatography-mass spectrometry (GC-MS) was employed for the high-throughput detection and identification of serum samples from 55 patients with MM and 37 healthy controls matched for age and sex from 2016 to 2017 collected at the First Affiliated Hospital of Soochow University. The relative standard deviation (RSD) of quality control (QC) samples was employed to validate the reproducibility of GC-MS approach. The differential metabolites between patients with MM and healthy controls were detected by partial least squares discrimination analysis (PLS-DA), and t-test with false discovery rate (FDR) correction. Metabolomics pathway analysis (MetPA) was employed to construct metabolic pathways. Results: There were 55 MM patients, including 34 males and 21 females. The median age was 60 years old (42-73 years old). There were 30 cases of IgG type, 9 cases of IgA type, 1 case of IgM type, 2 cases of non-secreted type, 1 case of double clone type and 12 cases of light chain type, including 3 cases of kappa light chain type and 9 cases of lambda light chain type. The result of QC sample test showed that the proportion of compounds with the RSD of the relative content of metabolites < 15% was 70.21% obtained by the reproducibility of GC-MS experimental data, which implied that the experimental data were reliable. A total of 17 metabolites were screened differently with the healthy control group, including myristic acid, hydroxyproline, cysteine, palmitic acid, L-leucine, stearic acid, methionine, phenylalanine, glycerin, serine, isoleucine, tyrosine, valine, citric acid, inositol, threonine, and oxalic acid (VIP>1, P<0.05). Metabolic pathway analysis suggested that metabolic disorders in MM patients comprised mainly phenylalanine metabolism, glyoxylic acid and dicarboxylic acid metabolism, phosphoinositide metabolism, cysteine and methionine metabolism, glycerolipid metabolism, glycine, serine, and threonine metabolism. Conclusion: Compared with normal people, patients with newly diagnosed MM have obvious differences in metabolic profiles and metabolic pathways.

Prognostic role of the ratio of natural killer cells to regulatory T cells in patients with multiple myeloma treated with lenalidomide and dexamethasone

Despite advances in treating newly diagnosed multiple myeloma (NDMM), a biomarker-driven personalized approach remains an unmet need. A combination of lenalidomide and dexamethasone (RD) is a widely available chemotherapeutic option for NDMM. We aimed to find a circulating immune cell-based biomarker to predict prognosis following RD in patients with NDMM. Clinical data and peripheral blood samples of 71 consecutive NDMM patients treated with RD were retrospectively analyzed. Peripheral blood samples were taken at the time of diagnosis. Immune cell populations, including natural killer (NK) cells, T cells, and their subpopulations, were identified by flow cytometry. In univariable analysis, four variables, including low expression (third or lower quartile) of NK cells, high expression (first or greater quartile) of regulatory T (Treg) cells, female sex, and lambda light chain type, could be plausible factors in predicting poor progression-free survival (PFS). With use of the ratio of NK cells to Treg cells (NK/Treg) as a biomarker, the median PFS of patients with low NK/Treg (less than first quartile, n=18) was significantly inferior to that of patients with high NK/Treg (first or greater quartile, n=53): 19.8 months versus 57.3 months, p=0.047. In multivariable analysis, low NK/Treg was significantly associated with poor PFS (hazard ratio: 2.877, 95% confidence interval: 0.001-1.009, p=0.048), even after adjusting for other confounding factors. NK/Treg at the time of diagnosis might be a useful immune cell biomarker for clinical decision-making for the use of RD in NDMM. Further investigations are needed to improve outcomes of NDMM patients based on the understanding of the role of NK/Treg.

IgG1 Subclass Restriction and Biochemical Peculiarities of Monoclonal Immunoglobulins in Scleromyxedema.

Scleromyxedema (SME) is a rare mucinosis associated with monoclonal gammopathy. Several biochemical peculiarities of monoclonal immunoglobulins (Ig) in SME patients were reported in case reports or short series, such as IgGλ over-representation, cationic migration, and partial deletion. Monoclonal immunoglobulins (Ig) from the serum of 12 consecutive patients diagnosed with scleromyxedema (SME) were analyzed using electrophoretic and immunoblotting techniques. All monoclonal Ig from 12 SME were of IgG1 subclass, with an overrepresentation of the lambda-type light chain and a cationic mobility on standard zone electrophoresis, as compared with 21 cases of monoclonal gammopathy of undetermined significance (MGUS) of IgG1 subclass. Reactivity with specific monoclonal antibodies demonstrated no evident deletion of the heavy chain constant domains, which was also confirmed by analysis of Ig heavy chain molecular weight on a purified monoclonal component from one case. Significant isotype restriction of both heavy and light chains, and peculiar biochemical properties suggest that monoclonal Ig might be involved in pathophysiological events of SME.

Prognostic Impact of Beta 2 Microglobulin in Patients with Immunoglobulin Light-Chain Amyloidosis Undergoing Autologous Hematopoietic Stem Cell Transplantation

Prognostic Impact of Beta 2 Microglobulin in Patients with Immunoglobulin Light-Chain Amyloidosis Undergoing Autologous Hematopoietic Stem Cell Transplantation

Brain Amyloidoma: Case Report and Literature Review

Introduction: Primary deposition of amyloid can take several forms including amyloidoma organ restricted among which is brain. Brain amyloidoma can mimic several lesions and offers a diagnostic challenge to the physician. The present work reports an additional case and summarizes the salient characteristics of previously published cases. Case Report: A 61-year-old, woman experienced paroxystic auditory hallucinations followed at times by generalized tonico-clonic seizures. Examination revealed a mild left pyramidal weakness. Head CT scan and MRI revealed a lobular lesion of the right atrium with contrast enhancement. The diagnosis of amyloidoma Lambda light chain (λLC) type was performed by means of a stereotactic biopsy. The patient was treated by 3 cures of high dose methotrexate (15 mg/m2). At 24 months follow up the patient remains stable clinically and biologically. The 2 years follow-up MRI showed the same findings. Conclusion: No guidelines can be suggested for brain amyloidoma treatment; the “wait and see” was the mind-set in the literature. However, a non-tumour like pattern of a cerebral mass developing near the ventricle with a fine, irregular radiating margin enhancement and lack of oedema, and mass effect are important MRI clues for brain amyloidoma diagnosis. The diagnosis is histological after biopsy or excision.

The diagnostic and prognostic values of serum free light chain in patients with primary light chain amyloidosis

目的研究血清游离轻链（sFLC）检测在原发性轻链型淀粉样变（pAL）中的诊断和预后价值。方法回顾性分析2009年1月至2015年6月确诊且有完整sFLC数据的126例pAL患者的临床资料，探讨sFLC的诊断价值，并采用Cox回归法分析sFLC差值（dFLC）的预后价值。结果126例患者中男女比例为1.57∶1，中位年龄为57 (37~81）岁，λ轻链型者80例（63.5%），肾脏受累者87例（69.0%），心脏受累者79例（62.7%）。126例患者的中位dFLC为99 (1~4 263）mg/L。血清蛋白电泳、血清免疫固定电泳、尿免疫固定电泳和sFLC比值异常法检测单克隆免疫球蛋白（M蛋白）的阳性检出率分别为34.9%（44/126）、63.5%（80/126）、77.0%（97/126）和81.0%（102/126）。联合上述四种方法可将M蛋白的检出率提高至98.4%（124/126）。中位随诊16个月后，37例患者死亡，所有患者的中位生存时间尚未达到。多因素分析发现dFLC ≥130 mg/L是影响pAL患者预后的独立危险因素（HR=3.272, 95% CI 1.384~7.739，P=0.007）。结论sFLC检测可显著提高pAL患者M蛋白的检出率；高水平dFLC是影响患者预后的独立危险因素。

The clinical spectrum of IgM monoclonal gammopathy: A single center retrospective study of 377 patients

ObjectivesWe retrospectively evaluated the clinical features, serum levels of IgM, and prevalence of IgM related diseases in patients with serum immunofixation electrophoresis (sIFE) confirmed IgM monoclonal gammopathy at our center. MethodsWe included patients with sIFE confirmed IgM monoclonal gammopathy between January 2008 and December 2014 in this retrospective study. We evaluated clinical data, sIFE, serum IgM levels, and diagnosis. ResultsIn total, 7107 patients had sIFE confirmed monoclonal gammopathy, with 377 (5.3%) patients having the IgM type. The median age was 62 years (range, 19–105 years). The median level of serum IgM is 8.3g/L (range, 0.24–150g/L). The diagnosis included monoclonal gammopathy of undetermined significance (MGUS, 157 patients, 41.6%), Waldenstrom macroglobulinemia (WM, 105 patients, 27.9%), B cell non-Hodgkin’s lymphoma (69 patients, 18.3%), primary cold agglutinin disease (pCAD, 16 patients, 4.2%), primary amyloidosis (14 patients, 3.7%), cryoglobulinaemia (six patients, 1.6%), IgM MGUS associated neuropathy (five patients, 1.3%), multiple myeloma (three patients, 0.8%), and POEMS syndrome (two patients, 0.5%). Levels of serum IgM>15.5g/L were 80.6% sensitive and 89.2% specific for the diagnosis of WM. Kappa type light chain indicated the diagnosis of WM, pCAD, IgM MGUS associated neuropathy and cryoglobulinaemia, while lambda type light chain indicated POEMS and amyloidosis. There were 41/157 (26.1%) MGUS patients diagnosed with complications due to IgM-unrelated autoimmune diseases. ConclusionIgM monoclonal gammopathy contains a broad spectrum of diseases. Levels of serum IgM and the type of light chain can be used to help with differential diagnosis. The association between MGUS and some autoimmune diseases requires further investigation.

Characterization of monoclonal gammopathy in patients with amyotrophic lateral sclerosis.

Recently, new guidelines for the monitoring and the risk evaluation of monoclonal gammopathy with undetermined significance (MGUS) became available and the light chain MGUS subtype was defined. To characterize the type, risk and diagnostic implications of MGUS in patients with amyotrophic lateral sclerosis. We screened 97 consecutive patients with ALS and 97 age- and gender-matched controls for MGUS by serum electrophoresis and immunofixation and compared the characteristics of MGUS with population-based data. MGUS was identified in 8.2% of ALS patients and 6.2% of controls (mean age: 62.5years both). Seven of eight ALS patients with MGUS had 'low-risk MGUS'. M-protein was moderately increased in one ALS patient. The immunoglobulin distribution in ALS patients with MGUS was IgG kappa (n=2), IgM lambda (n=1) and light chains of lambda type (n=3). No differences in demographic and clinical parameters were found between patients with and without MGUS. The percentage of patients with MGUS is increased in ALS, but the immunoglobulin distribution is similar to that reported in the general population. MGUS in ALS mostly represents 'low-risk MGUS'; therefore, unnecessary diagnostic procedures should be avoided in ALS patients.

Iron Overload Pattern in Multiple Myeloma at Diagnosis its Important Clinical Associations

As a plasmaproliferative disorder, multiple myeloma (MM) is not expected to cause IO and actually there are no reports on this condition in MM. In this retrospective study, we evaluated serum iron test results in MM patients at diagnosis in order to find the frequency of IO. Iron overload pattern (IOP) was described as increased transferring saturation (> 45%) in the presence of normal to increased serum iron and ferritin levels. Among 90 newly diagnosed MM patients (54 male, 36 female, and the median age: 61 years, range: 31-86) IOP was found in 17 cases (17/90, 19%). There was a close association between lambda light chain type paraproteinemia and IOP (13/17 in IOP cohort vs 26/73 lambda paraproteinemia, p= 0.003). Half of the MM patients (7/14) with free lambda light chain paraproteinemia had IOP. The main reason of the elevated TS was increased serum iron level in IOP patients, 144 (±71) vs 57 (±24) pg/ml (p< .001). Four out of 17 IOP patients had extramedullary involvement and this was significantly more frequent than the other patients. Although high International Staging System risk disease (stage 2 or 3) was more frequent in IOP patients (15/15 vs 51/68, p= 0.03), we could not observe any difference regarding treatment response, and survival between these cases and others. In conclusion, IO is also not infrequent MM at diagnosis and it has some clinical associations.

Diagnosis Of Amyloidosis Subtype By Laser-Capture Microdissection (LCM) and Tandem Mass Spectrometry (MS) Proteomic Analysis

AimCorrect identification of the protein that is causing amyloidosis is crucial for clinical management. Current standard diagnostic methods have limited ability to detect the full range of amyloid forming proteins. We assessed combining specific sampling of amyloid deposits by LCM and analysis of tryptic digests by tandem MS proteomic analysis to determine whether the majority of amyloidosis clinical samples can be correctly classified as reported by the Mayo Clinic pathology department. MethodsWe studied 58 cases of well characterised amyloid deposition and 10 cases in which the amyloid subtype was unable to be diagnosed with confidence. For all specimens, 10µm sections of formalin-fixed paraffin embedded tissue were stained with Congo Red using a standard technique. LCM was performed using an Arcturus XT instrument with an infrared capture laser. Proteins were digested with trypsin and peptides were analysed by nano-liquid chromatography-coupled tandem mass spectrometry using a Chip CUBE-QTOF. Database searching was performed using Spectrum Mill (Agilent) with the NCBInr human protein database. Protein identification cut-offs were protein score > 11, peptide score > 10 and % scored peak intensity >60. ResultsBiopsy sites included: GIT (n=16), cardiac (n=12), soft tissue (n=8), renal (n=9), liver (n=3) and other (n=20). The amyloid subtype was able to be determined in 64 cases analysed. In 7 of these cases a second sample or second LCM was required as the first analysis was non-diagnostic. In 4 cases the amyloidogenic protein was not identified mostly due to the amyloid deposits being very small. Proteins identified included immunoglobulin light chain (localised amyloid n=6, systemic AL n=32), transthyretin (senile amyloid n=17, hereditary ATTR n=2), serum amyloid A (AA n=4), fibrinogen (AFib n=1), TGFb (corneal lattice amyloid n=1) and semenogelin (seminal vesicle amyloid n=1).Three diagnostically challenging cases are detailed as examples of the utility of LCM and tandem MS. The first case had extensive gastrointestinal amyloidosis and no evidence of clonal light chain disease; negative kappa, lambda, SAA and transthyretin immunohistochemistry; and negative genetic studies. Tandem MS revealed immunoglobulin lambda light chain type. The second diagnostically challenging case had: isolated renal amyloidosis with a positive AA stain and kappa restricted serum free light chains. Tandem MS revealed serum amyloid A2 protein. The third case had: cardiac, neurological and gastrointestinal involvement; and equivocal immunohistochemistry. Tandem MS demonstrated transthyretin and genetic studies showed a A97S ATTR mutation.Various other proteins were identified by tandem MS in amyloid extracts. Of particular interest is the presence of proteins typically known to be co-located in amyloid deposits which helps confirm that the microdissected tissue is amyloid. Typical amyloid-associated proteins were identified in the following number of cases: SAP (n=36), apolipoprotein A4 (n=42), vitronectin (n=44), apolipoprotein E (n=40) and clusterin (n=21). Various types of collagen were frequently present (n=29) and various, presumably contaminating, keratins were identified (n=24). ConclusionLCM and tandem MS allows correct typing of amyloid deposits in the majority of clinical biopsy samples. Disclosures:No relevant conflicts of interest to declare.

Multiple myeloma with pleural involvement after pelvic radiotherapy for endometrial carcinoma

Although ionizing radiation is strongest factor linked to multiple myeloma, increased myeloma risk has not been fully explained after pelvic radiation. Pleural involvement of MM is also rare. We present a MM case with pleural involvement as an unusual presentation diagnosed in fifth years of pelvic radiotherapy. A sixty-two-year-old woman with dyspnea and a mass in forehead was admitted to our clinic. Before five year, the patient had received pelvic external beam radiotherapy (RT) with dose of 40 Gy for endometrial adenocarcinoma. PET/CT scan detected FDG uptakes in frontal bone, right pleura, and sacrum. Lambda light chain type multiple myeloma with pleural involvement was diagnosed with histopathological examinations of frontal bone mass, bone marrow, and pleural fluid and with serum/urine electroforesis. The patient died in second course of VAD chemotherapy. Although relation between increased myeloma risk and pelvic radiation is not clear and pleural involvement is rare, multiple myeloma should be included to the differential diagnosis in patients received pelvic radiotherapy or in unexplained pleural effusion.

Immunoglobulin Heavy/Light Chain Ratios as An Alternative to Immunofixation In the Identification of Monoclonal Gammopathies

AbstractAbstract 5018 IntroductionHistorically, serum protein electrophoresis (SPE), urine protein electrophoresis (UPE) and immunofixation (IFE) have been used to identify and quantify monoclonal proteins (M-proteins). Whilst this approach is adequate for the identification of intact immunoglobulin multiple myeloma (MM), it is not sensitive enough to detect free light chain MM (LCMM). Therefore, an algorithm which utilises SPE, serum free light chain (FLC) immunoassays and IFE for the identification of M-proteins has been suggested. Assays have now been developed which utilise polyclonal antisera raised against the kappa and lambda light chain types of IgG, IgA and IgM immunoglobulins (HLC). Here we report the use of these assays as an alternative to IFE and propose a screening algorithm which utilises SPE, FLC and HLC Materials and MethodsSerum FLC measurement was added to 1063 requests for SPE, from primary care or from a hospital source. Samples from patients with previously diagnosed MM, Waldenstrom's Macroglobulinemia and lymphoma were, where possible, removed. Sera showing monoclonal proteins or hypogammaglobulinemia (by SPE) or an abnormal FLC ratio were tested further by IFE and IgG, IgA, IgM HLC assays. SPE and IFE were performed on a SEBIA Hydrasys system, and gels were interpreted by experienced clinical chemists. FLC and HLC measurements were performed on a Siemens BN™II nephelometer. HLC results were compared with IFE results and clinical diagnoses. The study was approved by the Wolverhampton, New Cross, NHS Trust Review Board Results80/1063 patients were identified as having abnormal SPE or abnormal FLC results. 42/80 patients had positive IFE results. 24/42 of these patients were positive by HLC (Table 1), 11/42 had light chain only myeloma/MGUS, the remaining 7/42 were MGUS patients. The 7 MGUS patients (6 IgG and 1 IgM) with normal HLC ratios and positive IFE all had less than 2g/L monoclonal protein measured by SPE densitometry and a normal FLC ratio. Of the 38/80 with normal IFE's all had been investigated further because of abnormal FLC results. 9/38 had abnormal HLC ratios of which 3/9 had confirmed hematological malignancies (1× chronic lymphocytic leukemia (CLL), 1× small lymphocytic lymphoma (SLL) and 1× asymptomatic MM (ASMM)).The use of FLC immunoassays alongside SPE as part of the primary screening protocol identified 10 additional hematological malignancies (1× ASMM, 6×CLL, 2× non-Hodgkin lymphoma, 1× SLL). DiscussionHLC ratio analysis matched IFE for the identification of all symptomatic haematological malignancies. Abnormal FLC ratios identified 10 additional haematological malignancies of which 3 also had abnormal HLC ratios, which would have been missed using SPE/ IFE. In 7/19 MGUS (6×IgG, 1×IgM) patients there were normal HLC ratios. In all cases the monoclonal protein load was below 2g/L and the FLC ratio was normal; identifying the IgG patients as having a low risk and the IgM patient as having a low/intermediate risk of progression. It may be beneficial not to identify these patients, who do not require therapeutic intervention or justify close monitoring. Another advantage of using HLC analysis instead of IFE is that the HLC ratio has been found to be a prognostic indicator in myeloma and MGUS. It would also form a useful “baseline” comparison if HLC assays were used in monitoring or for the detection of residual disease. ConclusionsHLC analysis identified all symptomatic patients who were IFE positive and in an additional 3 hematological malignancies. Low risk MGUS patients may not be identified using these tests but this might be beneficial to patients and physicians alikeTable 1IFE positive patients with SPE densitometry and heavy/light chain immunoglobulin results.DiseasenRange SPE(g/L)HLCr abnormalIgA MM20–122/2IgA MGUS505/5IgGMM40–274/4IgG MGUS110–65/11IgG Amyloid101/1IgG Plasmacytoma1231/1Waldenstrom's101/1IgM MGUS32–202/3IgM lymphoma30–123/3 Disclosures:Harding:Binding Site Group Ltd: Employment.

Autologous Transplantation Is the Optimum Approach to the Management of Myeloma Patients with Extramedullary Disease at Presentation

Although usually confined to the bone marrow, extramedullary (EM) involvement has been reported in up to 15%–20% of patients with myeloma at diagnosis and develops in an additional 15% during the course of the disease. To date clinical observations on EM myeloma are based on small series of relapsed patients and results suggest that the disease course is often aggressive associated with both early progression and short survival. The purpose of this study was to compare the presenting clinical/laboratory features and outcome of newly presenting myeloma patients with and without extramedullary manifestations and to determine the optimum treatment for this group of patients. EM involvement was defined as the presence of extramedullary plasmacytomas clinically or radiologically; patients with solitary or multiple bony plasmacytomas were excluded. Among 459 newly-diagnosed symptomatic myeloma patients prospectively recorded in the Royal Marsden database, 75 (16.3%) had EM involvement. The main sites of involvement were paravertebral (56%), chest wall (15%) and subcutaneous (15%). The presence of EM disease was associated with higher haemoglobin, albumin, lower β2-M level, and consequently a more favorable ISS staging at diagnosis (p<0.05), which could be explained by the early presentation of these patients caused by the tumors. The EM group also had less IgG heavy chain type (36% vs 61.2%, p<0.001), more Bence-Jones (30.7% vs 13.8%, p<0.001) and non-secretory (9.3% vs 4.4%, p=0.09) type, and higher percentage of Lambda light chain type (47% vs 35%, p=0.05). The overall response rates to frontline treatment were similar for the two groups of patients; patients with EM disease having 45% CR, 31% PR and 24% NR compared to 47% CR, 34% PR and 19% NR in the other group. (P=0.5). In a univariate analysis for the whole group of patients, lower Hb, Alb, higher Ca, Cr, β2-M, older age, no high dose therapy (HDT) exposure and no thalidomide exposure were associated with shorter survival. There was no difference in progression free survival (PFS) and overall survival (OS) between the two groups. The OS was 62 months in patients with EM disease and 67 months in the other group (p=0.88), and the PFS was 24 vs 28 months (p=0.73). When adjusted according to independent risk factors (age, Ca, β2M, Alb, exposure to HDT and thalidomide), patients with EM disease who received conventional chemotherapy with no HDT had significantly shorter OS compared to those without EM disease who received similar treatment (p=0.046). HDT significantly improved the OS in both groups (p<0.001), however, it had more impact on OS among patients with EM disease, overcoming the negative prognostic impact of presenting extramedullary disease on the outcome of myeloma. A similar pattern was seen with PFS. In conclusion when treated optimally patients with EM disease at presentation have a similar outcome to patients without EM manifestations. Our results suggest that HDT should form an integral component of first line treatment for patients with extrameduallary disease.

Rapid Progressive Extramedullary Plasmacytoma in the Orbit

BACKGROUND: Plasma cell tumors of the orbit are uncommon and mostly the first manifestation of a multiple myeloma (MM). Primary extramedullary plasmacytoma (EMP) of the orbit is exceedingly rare and only a few cases have been reported so far. Differentiation of an EMP from a MM is important, since 9.5 years have been reported as the mean survival of EMP, compared with a mean survival of 3 to 4 years for multiple myeloma. HISTORY AND SIGNS: A 68-year-old man presented with a history of rapid progressive extramedullary plasmacytoma in the right orbit, increasing in size within a few weeks. MR imaging revealed an intraorbital mass measuring 5.2 × 4.7 × 3 cm. The earlier performed biopsy was positive for plasmacytoma of the lambda light chain type. Systemic work-up was negative for further tumor spread. THERAPY AND OUTCOME: Irradiation (a maximum of 80 Gy) and several chemotherapy cycles showed no effect. Exenteration with sparing of the upper and lower lid skin was performed. The residual defect was covered by a Thiersch graft. Although the orbit had been irradiated with the maximal dose, the Thiersch graft healed well. CONCLUSION: Normally, EMP is slow to increase in size. In our patient, marked growth of tumor was noted within one week. One underlying reason is the poorly differentiated, blastic morphology leading to this rapid tumor growth. However, particular molecular features, like a t(4;14), could not be demonstrated.

Rapid Progressive Extramedullary Plasmacytoma in the Orbit

Plasma cell tumors of the orbit are uncommon and mostly the first manifestation of a multiple myeloma (MM). Primary extramedullary plasmacytoma (EMP) of the orbit is exceedingly rare and only a few cases have been reported so far. Differentiation of an EMP from a MM is important, since 9.5 years have been reported as the mean survival of EMP, compared with a mean survival of 3 to 4 years for multiple myeloma. A 68-year-old man presented with a history of rapid progressive extramedullary plasmacytoma in the right orbit, increasing in size within a few weeks. MR imaging revealed an intraorbital mass measuring 5.2 x 4.7 x 3 cm. The earlier performed biopsy was positive for plasmacytoma of the lambda light chain type. Systemic work-up was negative for further tumor spread. Irradiation (a maximum of 80 Gy) and several chemotherapy cycles showed no effect. Exenteration with sparing of the upper and lower lid skin was performed. The residual defect was covered by a Thiersch graft. Although the orbit had been irradiated with the maximal dose, the Thiersch graft healed well. Normally, EMP is slow to increase in size. In our patient, marked growth of tumor was noted within one week. One underlying reason is the poorly differentiated, blastic morphology leading to this rapid tumor growth. However, particular molecular features, like a t(4;14), could not be demonstrated.

Systemic lupus Erythematosus and IgA multiple myeloma: a rare association?

The coexistence of systemic lupus Erythematosus (SLE) and multiple myeloma (MM) is uncommon and the pathogenetic mechanisms underlying this association remain unclear. We report the case of a woman who was diagnosed with SLE in 1993 aged 57, then developing IgA lambda type MM in the IIB clinical stage 7 years later. The SLE was treated successfully with methylprednisolone and chloroquine, and low dose maintenance steroid was continued with bisphosphonate protection until December 1994 when she suffered multiple vertebral fractures. She continued to receive 4 mg alternate day methylprednisolone and calcitonin until she decided to discontinue her own treatment 2 years later. In 2000, while still in stable SLE remission, she was diagnosed with MM. Protein electrophoresis revealed the IgA lambda paraprotein (40.5 g/l) and she had a Bence Jones (BJ) proteinuria of the lambda light chain type. Bone marrow trephine biopsy revealed a massive patchy infiltrate of abnormal plasmocytes (70%), while an extensive x-ray skeletal survey did not show any new fractures or osteolysis. The patient was treated according to the VMCP protocol without attaining a plateau phase. There was a similar poor clinical response to second and third line treatments (VAD, Thalidomide, Melphalan, and high dose dexamethasone). After 4 years of refractory disease the patient died from severe bilateral pneumonia. This case is discussed with reference to the literature.

Cloning of Idiotype Immunoglobulin Genes in B Cell Lymphomas by Anchored PCR and Production of Individual Recombinant Idiotype Vaccines in E. coli.

Abstract The hypervariable regions (idiotypes) of the immunoglobulins (Ig) expressed by lymphoma cells represent individual and specific tumor antigens. Purified or recombinant lymphoma-derived idiotype (Id) may be formulated into an individual vaccine and administered to lymphoma patients to elicit a therapeutic tumor-specific host immune response. We have developed an expression strategy based on anchored PCR amplification of lymphoma-derived cDNA. The Id variable (V) gene segments are inserted into a prokaryotic expression vector, and recombinant Fab fragment corresponding to the native Id protein is expressed in E.coli. This approach was applied to 106 biopsies with confirmed lymphoma involvement (47 follicular, 14 mantle cell, 6 diffuse large cell NHL, 22 myelomas, 6 CLL, and 6 other NHL types) and 8 non-lymphomatous control samples with the aim to investigate the feasibility of this production strategy in a large cohort of NHL patients, and to assess the yield and purity of the recombinant Id vaccines under large-scale production. According to clonality criteria defined to minimize the likelihood of mistakenly interpreting occasional repeat sequences (observed with low frequency in the control biopsies) as lymphoma-derived Id, cloning of Id VH and VL segments was successful in 95% of lymphoma biopsies. Expression of recombinant Fab fragment was successful (&amp;gt; 1 mg) in 93% of lymphomas. Fab yield was variable and ranged from 1.2 to 250 mg, whereas Fab purity was &amp;gt;95% in 95% of the cases after affinity chromatography purification. One Fab preparation only was contaminated with E.coli protein by western blot. MM had a trend towards superior Fab yields but this finding may merely reflect a selection for cases with secreted paraprotein, i.e. Ids with favorable folding properties and solubility. No other NHL type was associated with inferior expression levels. Fabs with lambda-type light chain had a trend towards lesser yield (p=0.0016) and purity (p=0.029) than kappa-Fabs. As expected, 25 of 38 follicular lymphoma cases had acquired novel potential glycosylation sites within their Id V segments (preferentially in CDR regions), whereas 34 of 40 other lymphomas lacked such sites (p&amp;lt;0.0001). Nevertheless, such sites did not influence the Fab yield (p=0.97). All other structural Id features (isotype, V gene usage, hypermutation status, CDRIII charge) also had no adverse influence on Fab expression. In summary, A-PCR cloning of Ig transcripts has a high success rate for the identification and analysis of Id genes in all major types of B-cell lymphoma. A-PCR with expression of idiotype protein in E. coli as a recombinant Fab fragment provides a solid basis for production of individual Id vaccines of all major IgH and IgL classes. The short culture time required for Id expression in bacteria in comparison to eukaryotic cell cultures may facilitate clinical trials and reduce production costs.

Characterization of amyloidogenic immunoglobulin light chains directly from serum by on‐line immunoaffinity isolation

Primary systemic amyloidosis (AL) is characterized by the overproduction of immunoglobulin light chain proteins by a monoclonal, terminally differentiated B-lymphocyte or plasma cell clone. The free immunoglobulin light chains are deposited in an abnormal conformation as amyloid in a variety of organs in the body. The mechanism of amyloid formation is not well understood, but appears to be associated with some form of cleavage of the immunoglobulin light chain with subsequent aggregate formation. In an attempt to characterize the structure of amyloid-forming light chain proteins we developed an on-line immunoaffinity purification and subsequent characterization of free kappa and free lambda immunoglobulin light chains by electrospray ionization mass spectrometry. The methodology is totally automated and requires 20 micro L of serum. Mass spectral analysis of Bence Jones proteins under non-denaturing conditions was also utilized to examine the tertiary and quaternary structure of light chain proteins and clearly shows covalent dimer formation of lambda type light chain. This type of on-line assay may prove helpful in elucidating distinguishing features capable of discriminating AL from benign monoclonal gammopathies of undetermined significance as well as diagnosing AL.