Serum Free Light Chain Research Articles

A novel and robust approach to defining population-based reference ranges for a serum free light chain assay in a clinical laboratory

Abstract Serum protein electrophoresis and immunofixation are used to detect the presence of monoclonal heavy and light immunoglobulin chains for the diagnosis of plasma cell neoplasms (PCN). In conjunction, the Freelite® assay has been used to detect excess free kappa and lambda light chains produced by PCN, where the kappa:lambda free light chain ratio (KLR) serves as a surrogate marker of clonality and disease. The manufacturer-defined KLR reference range of 0.26-1.65 is used in internationally recognized criteria for PCN diagnosis and monitoring, and KLR results affect clinical decision making, disease categorization, and response assessment. Our laboratory has verified the manufacturer-defined Freelite® KLR reference range three times using healthy control sera since assay implementation in 2006. Despite this, high percentages of abnormal KLR have consistently been observed among immunofixation-defined monoclonal-negative specimens. We therefore retrospectively interrogated 48,012 serum free light chain clinical results reported at our institution between 2010-2020 by extracting results from our laboratory information system and analyzing population distributions using the R programming language. For all specimens, the distribution of KLR values was shifted towards the towards the upper limit of the manufacturer-defined reference range, indicating that it poorly describes both our monoclonal-negative and monoclonal-positive patient populations. Furthermore, >80% of monoclonal-negative specimens exhibited KLR above the median of the manufacturer-defined reference range (0.26-1.65), while 24% (1,226/5,057) unexpectedly exhibited KLR >1.65. We found that this phenomenon has been present consistently since 2010 and is not affected by patient sex or age, impaired renal function, lot-to-lot variation, or assay drift. We therefore sought to establish an institution-appropriate KLR reference range by defining a reference population from within our existing dataset. We leveraged International Classification of Diseases codes, estimated glomerular function data, and serum/urine immunofixation results to exclude specimens from individuals with PCN- or renal-related diagnoses, impaired renal function, and prior, current, or subsequent monoclonal components. The resultant 1,536 monoclonal-negative reference specimens were then classified according to the manufacturer-defined KLR reference range, where 13% remained abnormally elevated, consistent with the published false positive rate in healthy volunteers. Lastly, we determined a new KLR reference range of 0.66-2.21 using the central 95% of KLR values from these reference specimens. When applied to our dataset, this institution-specific KLR reference range improved the accurate classification of specimens containing monoclonal free light chains by immunofixation, while reducing the false-positive rate in monoclonal-negative specimens. Together, our findings support the use of historical KLR and immunofixation data from routine clinical testing to derive institution-appropriate KLR reference ranges, which can then be used for continuous quality improvement. Implementation of this more-accurate KLR reference range should improve subsequent test utilization by non-specialist providers, reduce concern for/over-diagnosis of kappa-involved disease, and improve the detection and monitoring of lambda-involved disease.

The Clinical Significance of Light Chain Predominant Multiple Myeloma

Abstract Multiple Myeloma (MM) is a neoplastic clonal proliferation of terminally differentiated B- lymphocytes. It accounts for 1.6% of all cancers and about 10-15% of all hematological malignancies in the United States. Monoclonal immunoglobulins provide an indication of tumor burden in patients with plasma cell neoplasm. Recent development has shown that a proportion of intact immunoglobulin producing multiple myeloma secrete much higher amounts of free light chains than usual; i.e. light chain predominant multiple myeloma (LCPMM). About 18% of multiple myeloma secreting intact immunoglobulins secrete higher amount of free light compared to other intact immunoglobulin secreting myelomas and are associated with significantly poorer clinical outcomes. In this study, a retrospective investigation comparing the laboratory and clinical findings in conventional MM and LCPMM was undertaken. Three hundred and sixteen patients with conventional MM and seventy-one patients with LCPMM were studied. The results were remarkable for marked increase in death rate in the LCPMM group compared to the conventional MM group, 35.2% and 13.6% respectively; P-value .00001. The occurrence of dialysis was also markedly increased in the LCPMM group compared with the conventional MM; 18.3% vs. 3.5% P-value .00001. A markedly lower eGFR was observed in patients with LCPMM group compared to patients with conventional MM with an average value of 46.3 versus 69.4 ml respectively P value .0000003. In conclusion, light chain predominant plasma cell myeloma confers a significant disease burden, poor clinical outcomes and shorter survival. Further prospective trial will be helpful in understanding the pathogenesis, the course, and management of the disease. An assay for serum free monoclonal light chains has been described and could serve as a marker for minimal residual disease in such patients. We aim to further study the clinical impact of light chain predominant plasma cell myeloma (LCPMM) to increase the current knowledge of the disease and investigate therapeutic interventions.

A strategy for detecting an anomalous batch of kappa serum free light chains measurements using nonparametric analysis of patient deltas

Abstract Patient-based quality control (PBQC) involves monitoring for shifts in patient results to detect analytical problems. Established methods perform well, but only for assays with high volumes and approximately normal result distributions. Recent publications have described monitoring the average of patient deltas (AOD) as a way generalize PBQC methods more broadly. Here we propose a nonparametric analysis of patient deltas (NPAOD). To illustrate the concept we present a case of an analytical issue in the kappa serum free light chain assay (kappa sFLC) caused by a bad reagent pack and how it could have been detected using NPAOD. We were alerted to a possible issue by the ordering clinicians who noticed a series of unexpectedly high results. Significantly lower values were observed when the samples were repeated with a new reagent pack (median [IQR]: 5.3 [3.12-13.52] versus 2.52 [0.95-4.25], p < 0.5 e-7 by paired Wilcoxon test). To evaluate the use of PBQC in detecting this error, we analyzed one year of retrospective data comprising 7657 results from 3061 patients. The distribution of kappa sFLC values was positively skewed (mean = 14.5 g/dL, median = 2.2 g/dL, skew = 26.6). Both volumes and distributions varied significantly by day of the week following the scheduling of subspecialty clinics. These properties make the conventional approach of simple moving averages poorly suited for monitoring kappa sFLC at our institution. We computed delta values as the difference between the current result and the most recent previous result and produced delta values for 4596/7657 (60%) of the samples. The distribution of deltas was zero centered and symmetric (mean = 0.03 g/dL, median = 0.01 g/dL). Using the historical distribution as a reference, we found that the average of deltas was not significantly different for the results of the bad reagent pack (p = 0.78 by Student’s T test). In contrast, a nonparametric approach yielded a significant difference whether by comparison of medians using a Wilcoxon test (p = 0.0004) or comparison of empirical distributions using a Kolmogorov-Smirnov test (p = 1e-7). The latter approach may offer several benefits. First, the Komogorov-Smirnov test has greater statistical power because it considers both the position and the shape of the distribution. Second, an algorithm based on comparison of empirical distributions may have fewer hyperparameters to tune. We continue this work with an in-silico characterization of detection and false alarm rates at different magnitudes of analytical bias. In conclusion, this case study provides initial evidence that a nonparametric analysis of patient deltas may help to extend PBQC to additional assays that have unfavorable distributions and volumes for the simple moving average algorithm.

Management of Atrial Fibrillation Using Immunoglobulin Free Light Chains, Novel Biomarkers of Inflammation.

AF is the most common cardiac arrhythmia. There is growing evidence that inflammatory mechanisms play an important role in its pathogenesis; inflammasome activation contributes to the onset and progression of AF. An increase in NOD-like-receptor-pyrin domain-containing-3 (NLRP3) inflammasome activation releases proinflammatory cytokines that activate nuclear factor (NF)-κB, which regulates the production of immunoglobulin free light chains (FLCs). Serum FLC levels are increased in patients with AF, and FLCs are biomarkers of inflammation. Inflammasomes and NF-κB may be targets for anti-inflammatory strategies to prevent and treat AF when FLC levels are elevated. This review discusses the role of inflammation in the pathogenesis of AF, as well as FLCs as novel inflammatory biomarkers for the management of AF.

Abstract 10594: Cardiac Tamponade in a Covid-19 Positive Patient With Idiopathic Capillary Leak Syndrome: A Perilous Combination

Introduction: Clarkson’s disease, more known as idiopathic systemic capillary leak syndrome (SCLS) is a rare condition characterized by hypotensive episodes due to capillary hyperpermeability and can manifest as fluid accumulation in any organ or potential space. This may cause a range of complications: respiratory failure (from pulmonary edema or pleural effusion), mesenteric hypoperfusion (from hepatic congestion and severe intestinal edema), compartment syndrome and rhabdomyolysis, and even shock from hypovolemia or cardiac tamponade. Very few cases of cardiac tamponade in Clarkson's disease and COVID-19 have been reported, and when present, is certainly a serious and life-threatening complication. Case Summary: A 44-year-old female - with prior episodes of angioedema, pleural and pericardial effusions aggravated by menstruation and infection - presented with hypotension and diaphoresis. She was exposed to a COVID patient and eventually tested positive for COVID-19 infection. Persistent hypotension warranted triple inotropic support. Initial laboratories revealed hemoconcentration and hypoalbuminemia. Urinalysis was normal. 2D echocardiography showed normal left ventricular ejection fraction but with large pericardial effusion and signs of cardiac tamponade, hence, she underwent emergency pericardial windowing. She then developed pleural effusion which was drained surgically. Hepatitis, mycoplasmal and Epstein-Barr virus were negative. Direct and indirect Coomb’s test, and Lupus panel were unremarkable. Serum free light chain panel, immunopathology and C1 inhibitor results were non-diagnostic. Patient was managed as a case of idiopathic SCLS or Clarkson's disease. Methylprednisolone pulse therapy (MPPT) and intravenous immunoglobulin (IVIg) were administered. Patient was discharged with tapering doses of oral steroids and scheduled monthly IVIg infusion. Conclusions: Cardiac tamponade is a very rare but life-threatening complication of Clarkson’s disease, and on top of a COVID-19 infection, warrants emergent intervention. The pathogenesis is not yet fully elucidated. Diagnosis of SCLS is challenging thus recurrent pericardial and pleural effusions should raise the clinician’s index of suspicion.

X-LINKED LYMPHOPROLIFERATIVE DISEASE: DIAGNOSTIC CHALLENGE IN A MIDDLE-AGED MALE

<h3>Introduction</h3> X-Linked Lymphoproliferative Disease Type 1 (XLP1) is a rare disease of males caused by mutation in the gene <i>SH2D1A</i> and is characterized by vulnerability to EBV infection, lymphoma and dysgammaglobulinemia. Fulminant infectious mononucleosis in early childhood is the most common clinical manifestation, but on rare occasions XLP1 does not present until adulthood. <h3>Case Description</h3> We present a 43-year-old male with a history of recurrent sinusitis and bronchitis and leukopenia since childhood. He had EBV infection when he was 18 years old complicated by jaundice and hepatitis that required treatment with corticosteroids. At the age of 42, he developed ITP and was found to have mild splenomegaly. Serum immunoglobulins were mildly abnormal with IgG of 575, IgM of 762 and IgA of 145 mg/dL. Lymphocyte immunophenotyping was within normal limits and serum protein electrophoresis and serum free light chains were normal. Bone marrow biopsy was unremarkable. Invitate PID panel was performed, which revealed a pathogenic variant in SH2D1A gene (c.2T>C). SAP expression by flow cytometry revealed absence in NK cells but present in 17% of CD8+ T cell population suggestive of a somatic reversion in this population. He had mildly decreased NK cell function and positive EBV serology. <h3>Discussion</h3> We present a case of XLP1 diagnosis in adulthood with a history of EBV infection of moderate severity and mild initial immunologic abnormalities. In patients with somatic reversion in CD8+ T cells, milder disease may be more common and genetic testing should be considered early. The management options including bone marrow transplant should be carefully considered.

Polyneuropathy, Organomegaly, Endocrinopathy, Monoclonal gammopathy and Skin abnormalities (POEMS) Syndrome in a Filipino Nurse Returning to Work Following Bortezomib Treatment: A Case Report

POEMS syndrome (polyneuropathy, organomegaly, endocrinopathy, M-protein, skin changes) also known as Crow-Fukase syndrome or Takatsuki syndrome is a rare and disabling paraneoplastic syndrome that frequently occurs in the fifth or sixth decade of life without a known standard first-line therapy. A 34-year-old Filipino male nurse, who presented with gradually progressive distal lower extremity weakness and sharp shooting pain in bilateral legs accompanied by mild joint effusion on the left knee, hypertrichosis, bilateral conjunctival injection, and gynecomastia underwent extensive workup and was diagnosed with POEMS syndrome. Complete blood count revealed erythrocytosis and thrombocytosis with elevated serum VEGF (vascular endothelial growth factor) and elevated monoclonal serum free lambda light chains. The electrophysiologic studies revealed chronic demyelinating sensorimotor polyneuropathy while bone marrow core biopsy and bone marrow aspirate smear immunohistochemical staining showed it to be positive for lambda and CD138. He had an initial unsuccessful treatment course with melphalan and prednisone. Hence, bortezomib and dexamethasone were given which gave significant improvement in symptoms from the overall neuropathy limitation score of 5 to 1.

Clinic-pathological characteristics of rare tubulointerstitial diseases.

Tubulointerstitial diseases is one of the common causes of renal dysfunction. Some rare pathological types are easy to be misdiagnosed and missedly diagnosed because of their low prevalence and relatively insufficient understanding, which affects the treatment and prognosis of patients. This study aims to explore clinical manifestations and pathological characteristics of several rare tubulointerstitial diseases, and therefore to improve their diagnosis and treatment. A total of 9 363 patients diagnosed by renal biopsy in the Department of Nephrology, Second Xiangya Hospital, Central South University from November 2011 to September 2021 were selected. Six cases of light chain cast nephropathy (LCCN), 2 cases of light chain proximal tubulopathy (LCPT), 1 case of LCCN with LCPT, 4 cases of genetic tubulointerstitial disease, and 6 cases of non-genetic related tubulointerstitial lesion were screened out, and their clinical manifestations and renal biopsy pathological results were collected, compared, and analyzed. Patients with LCCN presented with mild to moderate anemia, microscopic hematuria, and mild to moderate proteinuria. Compared with patients with LCPT, proteinuria and anemia were more prominent in patients with LCCN. Five patients with LCCN and 2 patients with LCPT had elevated serum free kappa light chain. Five patients with LCCN presented clinically with acute kidney injury (AKI). Two patients with LCPT and 1 patient with LCCN and LCPT showed CKD combined with AKI, and 1 LCPT patient presented with typical Fanconi syndrome (FS). Five patients with LCCN, 2 patients with LCPT, and 1 patient with LCCN and LCPT were diagnosed with multiple myeloma. Five patients with LCCN had kappa light chain restriction in tubules on immunofluorescence and a "fractured" protein casts with pale periodic acid-Schiff (PAS) staining on light microscopy. Immunohistochemical staining of 2 LCPT patients showed strongly positive kappa light chain staining in the proximal tubular epithelial cells. And monoclonal light chain crystals in crystalline LCPT and abnormal lysosomes and different morphological inclusion bodies in noncrystalline LCPT were observed under the electron microscope. Six patients with LCCN were mainly treated by chemotherapy. Renal function was deteriorated in 1 patient, was stable in 4 patients, and was improved in 1 patient. Two patients with LCPT improved their renal function after chemotherapy. Four patients with genetic tubulointerstitial disease were clinically presented as CKD, mostly mild proteinuria, with or without microscopic hematuria, and also presented with hyperuricemia, urine glucose under normal blood glucose, anemia, polycystic kidneys. Only 1 case had a clear family history, and the diagnosis was mainly based on renal pathological characteristics and genetic testing. Compared with patients with non-genetic related tubulointerstitial lesion, patients with genetic tubulointerstitial disease had an earlier age of onset, higher blood uric acid, lower Hb and estiated glomemlar fitration (eGFR), and less edema and hypertension. Renal pathology of genetic tubulointerstitial disease presented tubular atrophy and interstitial fibrosis, abnormal tubular dilation, glomerular capsuledilation, and glomerular capillary loop shrinkage. Glomerular dysplasia and varying degrees of glomerular sclerosis were observed. Genetic tubulointerstitial disease patients were mainly treated with enteral dialysis, hypouricemic and hypoglycemic treatment. Two genetic tubulointerstitial disease patients had significantly deteriorated renal function, and 2 patients had stable renal function. Patients with AKI or FS, who present serum immunofixation electrophoresis and/or serum free kappa light chain abnormalities, should be alert to LCCN or LCPT. Renal biopsy is a critical detection for diagnosis of LCCN and LCPT. Chemotherapy and stem cell transplantation could delay progression of renal function in patients with LCCN and LCPT. If the non-atrophic area of the renal interstitium presents glomerular capsule dilatation, glomerular capillary loop shrinkage, and abnormal tubular dilatation under the light microscopy, genetic tubulointerstitial disease might be considered, which should be traced to family history and can be diagnosed by genetic testing.

Role of serum-free light chain assay for defining response and progression in immunoglobulin secretory multiple myeloma.

The International Myeloma Working Group (IMWG) guidelines recommend using electrophoresis and immunofixation to define response and progressive disease (PD) in immunoglobulin (Ig) secretory multiple myeloma (Ig-MM), whereas the role of serum-free light chain (sFLC) is controversial. We retrospectively analyzed the value of adding sFLC assays in the definition of response and PD according to IMWG criteria in 339 Ig-MM patients treated with a first-line novel agent-based therapy (median follow-up 54 months). sFLC PD was defined according to conventional criteria plus increased sFLC levels, or sFLC escape (sFLCe); progression/sFLCe-free survival (ePFS) was the time from the start of treatment to the date of first PD or sFLCe, or death; overall survival after PD/sFLCe (OS after Pe) was the time from first PD or sFLCe to the date of death. 148 (44%) patients achieved a complete response and 198 (60%) a normal sFLC ratio (sFLCR). sFLCR normalization was an independent prognostic factor for extended PFS (HR=0.46, p=0.001) and OS (HR=0.47, p=0.006) by multivariable analysis. 175 (52%) patients experienced PD according to the IMWG criteria, whereas 180 (53%) experienced PD or sFLCe. Overall, a sFLCe was observed in 31 (9%) patients. Median PFS and ePFS were both equal to 36 (95% CI=32-42, and 32-40, respectively) months. sFLC PD adversely affected the OS after Pe compared to PD with increasing monoclonal Ig only (HR=0.52, p=0.012). Our results support the inclusion of the sFLC assay for defining response and PD in Ig-MM.

Progression, regression and redefining the treatment response – cardiac magnetic resonance with T1 and extracellular volume mapping in cardiac light-chain amyloidosis

Abstract Background The presence and severity of cardiac involvement in AL amyloidosis is the main driver of prognosis [1]; patients with symptomatic heart failure frequently die within 6 months [1] but median survival has nearly doubled over the past decade, mainly due to significant improvements in chemotherapy. The haematological response to chemotherapy is principally evaluated with serial measurements of serum-free light-chains (FLC) [2]. The cardiac response to chemotherapy is assessed through changes in serum concentrations of brain natriuretic peptides (including NT-proBNP) and echocardiographic parameters [3–5]. Neither are able to directly measure cardiac amyloid burden. Cardiovascular magnetic resonance (CMR) with extra-cellular volume (ECV) mapping can measure the extent cardiac amyloid infiltration [6]. Aims We investigated the ability of CMR to: 1) measure changes in response to chemotherapy; 2) assess the correlation between haematological response (HMR) and changes in cardiac amyloid; 3) assess the association between changes in cardiac amyloid and prognosis over and above existing predictors. Methods In total, 176 patients with cardiac light-chain amyloidosis treated with chemotherapy were assessed with FLC, NT-proBNP and CMR with ECV mapping at baseline (before chemotherapy), 6-months, 12-months &amp; 24-months after commencing chemotherapy. Haematological response was categorized by reductions in FLC as: complete response (CR), very good partial response (VGPR), partial response (PR) or no response (NR). CMR response was categorized by changes in ECV as: progression (≥0.05 increase), stable (&amp;lt;0.05 change) or regression (≥0.05 decrease). Results A progressive increase in patients achieving either CR or VGPR was observed at each time point (61% of patients at 6-months, 71% at 12-months and 80% at 24-months). At 6-months, CMR regression was observed in 3% (all had either CR or VGPR) and progression in 32% (61% had either PR or NR; 39% had either CR or VGPR). At 1-year, CMR regression was observed in 22% (all had either CR or VGPR); progression in 22% (63% had either PR or NR; 37% had either CR or VGPR). At 2-years, CMR regression was observed in 38% (all had CR/VGPR); progression in 14% (80% had either PR or NR; 20% had either CR or VGPR). During follow-up (40±15 months), 36 (25%) patients died. CMR response at 6-months predicted death (progression HR 3.821; 95% CI 1.950–7.487; p&amp;lt;0.001) and remained independently associated with prognosis after adjusting for haematological response, NT-proBNP and longitudinal strain on echocardiography (p&amp;lt;0.01). Conclusions CMR demonstrates that cardiac amyloid deposits frequently regress following chemotherapy, but only in patients who achieve CR or VGPR, highlighting the need for deep haematological response. Changes in amyloid burden (ECV) predict outcomes after adjusting for known predictors, showing the crucial role of CMR in redefining treatment response. Funding Acknowledgement Type of funding sources: Foundation.

Correlation between Serum Free Light Chain and Blood Routine Parameters in Patients with Multiple Myeloma

To investigate the correlation between serum free light chain (sFLC) and blood routine parameters in patients with multiple myeloma (MM). 347 patients with multiple myeloma diagnosed in Sichuan people's Hospital from April 2019 to July 2021 were selected. sFLC, serum total light chain (sTLC), peripheral blood routine, coagulation and biochemical parameters were analyzed retrospectively. The correlation analysis between sFLC and blood routine parameters were calculated by Pearson or Spearman correlation coefficients. Multiple stepwise linear regression was used to screen the combined blood routine parameters related to sFLC. The efficacy of the selected blood routine parameters eflecting sFLC level was evaluated, the Kruskal Wallis test of independent samples was used for inter group comparison, and the receiver operating characteristic (ROC) curve was drawn at the same time. In MM patients, sFLC was positively related with sTLCκ, sTLCλ, sTLCκ/λ, Cr, Urea, Cr and Cys_C significantly, while negatively correlated with eGFR markedly (｜r｜≥0.3). Multivariate stepwise linear regression showed that the influence factors of sFLCκ were Cr and sTLCκ (P=0.000, P=0.003), the influence factors of sFLCλ were eGFR and sTLCλ(P=0.000, P=0.000), the sFLCκ/λ influence factor was sTLCκ/λ (P=0.032). Kruskal Wallis test of independent samples showed that Cr and sTLCκ, eGFR and sTLCλ, TLCκ/λ were good or better parameters to reflect the level of sFLCκ, sFLCλ and sFLCκ/λ in MM patients(P＜0.05), respectively. ROC curve analysis shows that Cr, sTLCκ, eGFR, sTLCλ and sTLCκ/λ had the ability to judge the abnormality of sFLC in MM patients (AUC=0.684-0.875, P＜0.05). In MM patients, sTLCκ and sTLCλ with renal function parameters could evaluate sFLCκ or sFLCλ level respectively, while sFLC κ/λ was estimated by sTLCκ/λ.

A CASE OF TYPE 1 CRYOGLOBULINEMIA PRESENTING WITH DIFFUSE ALVEOLAR HEMORRHAGE

A CASE OF TYPE 1 CRYOGLOBULINEMIA PRESENTING WITH DIFFUSE ALVEOLAR HEMORRHAGE

The role of serum-free light chain ratios in the prediction of poor prognosis in multiple myeloma patients: a systematic review and meta-analysis

ABSTRACT Background The association between the serum free light chain (sFLC) ratio and the prognosis of multiple myeloma (MM) patients is controversial. Aim The purpose of this study is to explore the relationship between the sFLC ratio and the prognosis of MM patients through meta-analysis. Methods Online public databases were searched to find relevant studies. The retrieval time is limited from the establishment of the database to July 2021. The overall survival (OS) and progression-free survival (PFS) rates were compared. The results were described using hazard ratio (HR) and a 95% confidence interval (CI). Qualitative studies were also included. Results A total of 9 studies involving 2864 participants were included. A pooled analysis based on four studies including newly-diagnosed MM patients, demonstrated that an abnormal sFLC ratio was associated with poor outcomes of OS (HR = 1.82, 95% CI: 1.15–2.90) and PFS (HR = 1.87, 95% CI: 1.20–2.90). Three qualitative studies showed that an abnormal sFLC ratio was related with poor outcomes of OS (studies all included newly diagnosed MM patients) and PFS (two studies included newly-diagnosed MM patients and one study included non-newly-diagnosed MM patients). Two studies stated that the sFLC ratio is not associated with OS (both studies included non-newly-diagnosed MM patients) and one study reported that the sFLC ratio is not associated with PFS (study included non-newly-diagnosed MM patients). Conclusion sFLC ratio could be used to predict adverse outcomes in newly-diagnosed MM patients, but is not suitable for non-newly-diagnosed MM patients.

P21 The fly in the ointment: a rare cause of cytopaenia

Introduction/BackgroundThose working in Rheumatology frequently encounter cytopaenia both as adverse effects of many conventional and biological disease modifying drugs and as manifestations of autoimmune diseases. In cases where cytopaenia progresses, other explanations should be sought. Here we present an unusual cause of cytopaenia, seen in a patient with psoriatic arthritis on immunosuppression.Description/MethodAn 81-year-old woman with a diagnosis of psoriatic arthritis had been maintained on Infliximab (3mg/kg 8 weekly) and Methotrexate (7.5mg PO weekly) for 12 years. She then developed mild neutropaenia (1.52 x 109/L). Methotrexate was stopped and she continued on Infliximab monotherapy for several months, until ongoing neutropenia resulted in Infliximab being withheld. Sustained mild neutropenia and the development of thrombocytopenia (platelets 95 x 109/L) prompted a referral to Haematology, who suspected autoimmune neutropenia and advised checking haematinics and monitoring the FBC monthly. The neutropenia improved and Infliximab was restarted.Over 6 months she developed gradually worsening and persistent pancytopenia (neutrophils 1.36 x 109/L, platelets 134 x 109/L, Hb 91 g/L). There was no history of fevers, night sweats, bone pain, headaches, recurrent infections, lumps or bumps, chest symptoms, change in bowel habit or urinary symptoms. Examination revealed no pallor, jaundice, clubbing, peripheral lymphadenopathy or sternal tenderness. The chest was clear and the abdomen soft with no organomegaly.Repeat bloods showed microcytic hypochromic anaemia (Hb 86 g/L, MCV 73 fL, MCH 24.6 pg) with target cells on blood film. Iron was low (iron 5.1 umol/L, transferrin 1.52 g/L). Serum protein electrophoresis showed paraprotein bands with abnormal serum free light chain assay possibly consistent with myeloma, lymphoma or MGUS. Haematology suspected myelodysplasia, potentially secondary to previous long term Methotrexate therapy. A bone marrow biopsy was performed; the unexpected result was Leishmaniasis. Our patient was admitted to hospital and, following microbiology review, was treated with liposomal amphotericin (IV) for Visceral Leishmaniasis as an inpatient. Her Infliximab was suspended and her joints have remained quiescent.Discussion/ResultsLeishmania is an intracellular parasitic protozoan that is transmitted to humans by sandfly bites. It is endemic in Africa, America, Central Asia and the Mediterranean. It usually has an incubation period of 3-8 months. Those who are immunosuppressed, for example due to immunotherapy or HIV, are at increased risk of infection. The patient had only ever travelled to Portugal and Gibraltar. One trip to Gibraltar occurred whilst on dual immunosuppression with Infliximab and Methotrexate and prior to the development of neutropenia. She has no recollection of any insect bites.Visceral leishmaniasis is variable in the severity of symptoms experienced ranging from asymptomatic to a febrile illness with lymphadenopathy, hepatospenomegaly, night sweats, cutaneous pigmentation, appetite loss and weight loss, which can be drastic. Our patient indeed described significant weight loss in the preceding year.This case demonstrated the laboratory abnormalities which are typical of Visceral Leishmaniasis: pancytopenia and hypergammaglobulinaemia.Treatment for visceral Leishmaniasis has classically been with pentavalent antimonials (based on the heavy metal antimony) such as Sodium stibogluconate. Their toxicity profile and growing resistance however has meant amphotericin B is often used first line. This is given intravenously, with the liposomal form preferred as it has lower toxicity. Our patient received 4 mg per kg daily on days 1–5, 10, 17, 24, 31, and 38, following the FDA-approved regimen for immunosuppressed patients.Without treatment Leishmaniasis is usually fatal, with haemmorrhage, infection, cachexia and multiorgan failure as causes of death.Key learning points/ConclusionWhilst patients with Rheumatological conditions may have cytopaenia due to their disease or its treatment, persistent, progressive or unexplained results should be investigated further. In patients with cytopaenia of unclear aetiology, thorough investigation with Haematology involvement is mandatory. You may find an unexpected diagnosis … we did!

Prevalence of Monoclonal Gammopathy of Undetermined Significance in Black South African Men.

Both multiple myeloma and its precursor, monoclonal gammopathy of undetermined significance (MGUS), occur twice as often within Black compared with White populations, suggesting that racial disparities lie within the development of MGUS. Nonetheless, MGUS has been studied mainly in White cohorts; the study that first described the natural history of MGUS was conducted in 97.3% White Olmsted County, Minnesota. We determined the prevalence of MGUS among 386 Black South African (SA) men >30 years at Chris Hani Baragwanath Hospital in Johannesburg. We conducted serum protein electrophoresis and free light chain quantification to define MGUS by the same criteria as the Olmsted County studies. We also investigated the association between MGUS and various clinical factors, including human immunodeficiency virus (HIV) infection and smoking. We found the prevalence of MGUS to be 8.03% [95% confidence interval (CI), 5.32-10.74], nearly 1.6-fold higher than in the White Olmsted County male population. In a univariable logistic regression model, MGUS was associated with HIV status (OR, 2.39; 95% CI, 0.95-5.49), but in an adjusted model that included body mass index and cigarette use, the association was not statistically significant. Those who were current (vs. never) cigarette smokers were more likely to have MGUS in both univariable (OR, 5.60; 95% CI, 2.16-17.42) and multivariable models (OR, 4.49; 95% CI, 1.63-14.56). The prevalence of MGUS in Black SA men is substantially higher than in White populations and may be associated with HIV status and cigarette use. Racial disparities in MGUS exist and may be associated with potentially modifiable risk factors.

An echo score raises the suspicion of cardiac amyloidosis in Chinese with heart failure with preserved ejection fraction.

Transthyretin cardiac amyloidosis (ATTR-CA) has been realized as an important cause of heart failure with preserved ejection fraction (HFpEF). We aim to provide insights into its prevalence in Chinese HFpEF patients, which is not known to date, using increased wall thickness (IWT) score by echocardiography. Consecutive patients with HFpEF (EF≥40%) and IWT (≥12mm) were prospectively screened. Echocardiography was performed, and the IWT score incorporated relative wall thickness, E/e' ratio, longitudinal strains, and tricuspid annular plane systolic excursion, and septal apical-to-base ratio was calculated. ATTR-CA was defined as score ≥8 in the absence of serum and urine free light chain. Six hundred twenty-four HFpEF patients from January 2019 to December 2021 were enrolled, of which 65.2% were males and the median (interquartile range [IQR]) age was 66 (IQR 57, 73) years. Thirty-three patients (5.3%, 95% CI 3.5-7.0%) were with score ≥8, and 33.3% were females. They were younger (58 vs. 69years, P<0.001), had higher NT-proBNP (6525.0 vs. 1741.5pg/mL, P<0.001) and troponin I (105.2 vs. 27.7pg/mL, P=0.001) level, and lower LVEF (47% vs. 57%, P<0.001) compared with the patients with score <5. In the internal cohort (82 patients) who had undergone scintigraphy, the IWT score ≥8 was shown to have a sensitivity of 85.7% (95% CI 56.2-97.5%) and a specificity of 92.6% (95% CI 83.0-97.3%) for diagnosing CA, and the IWT score <5 had great accuracy in excluding CA with the negative predictive value of 100%, supporting the clinical usefulness of the IWT score to guide further dedicated testing for ATTR-CA. The IWT score by echocardiography was an excellent tool for screening ATTR-CA in HFpEF. In Chinese HFpEF patients associated with a hypertrophic phenotype, the proportion of highly suspected ATTR-CA as detected by IWT score ≥8 was 5.3%, lower than the reported prevalence of ATTR-CA in non-Asian patients with the disease.

Defining new reference intervals for serum free light chains in individuals with chronic kidney disease: Results of the iStopMM study

Serum free light chain (FLC) concentration is greatly affected by kidney function. Using a large prospective population-based cohort, we aimed to establish a reference interval for FLCs in persons with chronic kidney disease (CKD). A total of 75422 participants of the iStopMM study were screened with serum FLC, serum protein electrophoresis and immunofixation. Estimated glomerular filtration rate (eGFR) was calculated from serum creatinine. Central 99% reference intervals were determined, and 95% confidence intervals calculated. Included were 6461 (12%) participants with measured FLCs, eGFR < 60 mL/min/1.73 m2, not receiving renal replacement therapy, and without evidence of monoclonality. Using current reference intervals, 60% and 21% had kappa and lambda FLC values outside the normal range. The FLC ratio was outside standard reference interval (0.26–1.65) in 9% of participants and outside current kidney reference interval (0.37–3.10) in 0.7%. New reference intervals for FLC and FLC ratio were established. New reference intervals for the FLC ratio were 0.46–2.62, 0.48–3.38, and 0.54–3.30 for eGFR 45–59, 30–44, and < 30 mL/min/1.73 m2 groups, respectively. The crude prevalence of LC-MGUS in CKD patients was 0.5%. We conclude that current reference intervals for FLC and FLC ratio are inaccurate in CKD patients and propose new eGFR based reference intervals to be implemented.

Laboratory characteristics of IgG4-related disease: A retrospective study from a single tertiary medical center.

Immunoglobulin G4-related disease (IgG4-RD) is an immune-mediated fibroinflammatory condition with unique histopathological features that can affect most organs, making diagnosis challenging. This study characterized detailed laboratory characteristics of IgG4-RD. Baseline clinical and laboratory features of 33 patients with IgG4-RD were reviewed, including serum IgG4 concentrations, serum free light chains (sFLCs), IgGĸ- and IgGλ-heavy/light chains (HLCs), capillary serum protein electrophoresis (SPE), and immunofixation electrophoresis (IFE) of IgG4 subclass. The cohort of 33 patients showed male predominance (94%), with 8 (24%) exhibiting multiple organ involvement. Most patients (88%) had an elevated IgG4 concentration, and 67% had elevated erythrocyte sedimentation rate and IgE levels. Median IgG4 concentration at baseline was significantly higher in patients with >2 organs involved than those with ≤2. Furthermore, erythrocyte sedimentation rate was significantly correlated with serum IgG4 concentrations at baseline. SPE results demonstrated polyclonal gammopathy in most patients. Half of the patients had an increased κ/λ sFLC ratio, 42% had an increased IgGκ/IgGλ HLC ratio. Most patients exhibited hypergammaglobulinemia in the anodal end of the ɤ region on SPE. This study describes detailed laboratory features of IgG4-RD. Although none of these tests are considered diagnostically sufficient by itself, the provided laboratory characteristics can increase awareness of this disorder and help distinguish it from other IgG4-RD mimics.

Primary amyloidosis of conjunctiva.

A 64-year old male presented in ophthalmology OPD for follow up for refractive errors. He had been wearing bifocal lenses from last 1 year. On local examination, medial canthus of left eye showed a pale yellowish plaque. The rest of the ocular examination and contralateral eye were normal. Slit lamp examination showed a confluent fusiform papule (Figure 1A,B). Computed tomography (CT) scan of the orbit showed no abnormality. Surgical excision was done and tissue sent for histopathological examination showed deposition of a pink homogeneous material in the fibrocollagenous tissue beneath the conjunctival epithelium (Figure 1,C). Congo red stain followed by polarizing microscopy confirmed the pink material to be amyloid with apple green birefringence (Figure 1D,E). Immunohistochemistry (IHC) showed lambda light chain restriction. A diagnosis of conjunctival amyloidosis was made. The patient was then evaluated to rule out systemic amyloidosis or plasma cell dyscrasia. The patient was asymptomatic with no history of trauma, systemic illness, and family history was non-contributory. General and systemic examinations were unremarkable. Complete hemogram, urea, creatinine, liver function test, erythrocyte sedimentation rate, blood glucose, and serum calcium were within normal limits. No abnormality was detected on skeletal survey, electrocardiography, 2D-echocardiography, and ultrasonography of abdomen. Serum protein electrophoresis, serum immunofixation electrophoresis, serum free light chain assay, 24-h urine protein, and urine protein electrophoresis were non-contributory. Bone marrow showed mildly increased plasma cells without light chain restriction on immunohistochemistry. Finally, a diagnosis of primary conjunctival amyloidosis was made and the patient was asked to follow up. Conjunctival amyloidosis occurs secondary to local inflammation and primary form of the disease is very rare. In a previous series published on ocular amyloidosis, only two of 26 ocular cases collected over 30 years had conjunctival involvement. Identification of the disease is a daunting task owing to protean manifestations. Differentiating it from conjunctival inflammation and malignancies is essential. Evaluation for systemic causes is important as the treatment modalities are completely different. While localized form might just need observation or surgical debulking, systemic AL amyloidosis or myeloma would necessitate administration of specific chemotherapy. A long-term follow up is also necessary as recurrence is a known phenomenon and evolution into systemic disease is also a possibility. Concepts design, definition of intellectual content, Literature search, data analysis, manuscript preparation, and manuscript editing: Karuna Jha. Concepts design, definition of intellectual content, manuscript editing: Maitreyee Bhattacharyya. Dr Bhaskar Mitra, Consultant Pathologist, Tribedi and Roy Lab, Kolkata. The authors declare they have no conflicts of interest. The authors received no specific funding for this work. No research on human was performed on this study. Informed consent was taken from the patient. The data that support the findings of this study are available from the corresponding author upon reasonable request.

Circulating Tumor and Immune Cells for Minimally Invasive Risk Stratification of Smoldering Multiple Myeloma.

Early intervention in smoldering multiple myeloma (SMM) requires optimal risk stratification to avoid under- and overtreatment. We hypothesized that replacing bone marrow (BM) plasma cells (PC) for circulating tumor cells (CTC), and adding immune biomarkers in peripheral blood (PB) for the identification of patients at risk of progression due to lost immune surveillance, could improve the International Myeloma Working Group 20/2/20 model. We report the outcomes of 150 patients with SMM enrolled in the iMMunocell study, in which serial assessment of tumor and immune cells in PB was performed every 6 months for a period of 3 years since enrollment. Patients with >0.015% versus ≤0.015% CTCs at baseline had a median time-to-progression of 17 months versus not reached (HR, 4.9; P<0.001). Presence of >20% BM PCs had no prognostic value in a multivariate analysis that included serum free light-chain ratio >20, >2 g/dL M-protein, and >0.015% CTCs. The 20/2/20 and 20/2/0.015 models yielded similar risk stratification (C-index of 0.76 and 0.78). The combination of the 20/2/0.015 model with an immune risk score based on the percentages of SLAN+ and SLAN- nonclassical monocytes, CD69+HLADR+ cytotoxic NK cells, and CD4+CXCR3+ stem central memory T cells, allowed patient' stratification into low, intermediate-low, intermediate-high, and high-risk disease with 0%, 20%, 39%, and 73% rates of progression at 2 years. This study showed that CTCs outperform BM PCs for assessing tumor burden. Additional analysis in larger series are needed to define a consensus cutoff of CTCs for minimally invasive stratification of SMM.