Free Light Chain Measurements Research Articles

Performance goals for immunoglobulins and serum free light chain measurements in plasma cell dyscrasias can be based on biological variation.

Measurements of immunoglobulins and serum free light chains (sFLC) are frequently used in patients with monoclonal plasma cell dyscrasia (PCD). For optimum patient care, well-defined performance standards or goals for the measured concentrations of immunoglobulins and sFLC are required. Generally, data based on biological variation is a good and reliable method for setting desirable performance standards; this also applies for the measurements of paraprotein and sFLC. The benefits of this approach are several. Among others, it is independent of the clinician, and it provides us with information about reference change value and index of individuality. Several studies on biological variation of both immunoglobulins and sFLC have been published, and mostly the studies are well performed. The studies normally show small within-subject biological variation resulting in strict analytical goals, which in most cases are difficult to meet. Nevertheless, we still need further information on biological variation of immunoglobulins and sFLC in patients with PCD and in the elderly, which are the main target populations for the two measurands. Furthermore, to improve data on biological variation of immunoglobulins and sFLC, studies accounting for number of individuals, samples, and replicates, as well as time length of the studies are needed.

Minimal tumour burden in haematological diseases: a step forward with quantitative assessment of Bence-Jones in nephelometry?

Minimal tumour burden in haematological diseases: a step forward with quantitative assessment of Bence-Jones in nephelometry?

Serum Versus Urine Free Light Chains for Assessment of Monoclonal Gammopathies

IntroductionMonoclonal gammopathies comprise a spectrum of disorders including Monoclonal Gammopathy of Undetermined Significant (MGUS), Smoldering Multiple Myeloma (SMM), and Active Multiple Myeloma (MM) characterized by production of monoclonal immunoglobulin heavy and/or light chains. Prior to availability of the FREELITE™ (Binding Site Ltd; Birmingham, UK) assay for measurement of immunoglobulin free light chains (FLC), laboratory monitoring of these disorders used predominantly SPEP, quantitation of immunoglobulin heavy chains (quantitative immunoglobulins), and 24 urine collection for total protein and UPEP to extrapolate production of immunoglobulin light chains. The FREELITE™ assay has up to 3-log increased sensitivity (1.5-3.0 mg/L) for detection of free light chains over standard electrophoresis (500-2,000 mg/L) and immunofixation (150-500 mg/L), and since its introduction, has been an integral tool in diagnosis and monitoring of monoclonal gammopathies. This assay detects more plasma cell disorders than SPEP, UPEP and IFE combined due to its higher sensitivity and ability to derive the ratio of affected to unaffected light chain. Measurement of urine FLC using FREELITE™ has not been integrated into standard practice due to presumed variability in FLC concentration due to changes in glomerular filtration, variability in tubular reabsorption of light chains, and lack of data regarding this use. In our practice, we routinely use random urine samples instead of 24 hour urine collections which are cumbersome and suffer from poor patient compliance.Methods:The study was approved by the Stratton VA Medical Center Institutional Review Board. As it has been our practice to obtain both random urine along with serum for FLC, we retrospectively reviewed patients diagnosed with monoclonal gammopathies and compared random urine free light chains measured by FREELITE™ to serum FLC and serum quantitative immunoglobulins. Data was analyzed for correlation using Pearson product moment correlation. P values of >0.05 were considered significant.Results:We identified 23 individuals, all male (consistent with VA population). Mean (±SD) age was 68±10 years at diagnosis, creatinine 1.3±0.5 mg/dl, and 9±6 pairs of data points per patient. Five (5) had MGUS, 5 SM, and 13 MM (2 light chain only). Results are illustrated in the Table. Normalization of urine results using concurrent serum and urine creatinine did not change the statistical significance of any of the results.TableCorrelation (p<0.05) between affected serum immunoglobulin, urine FLC, and serum FLCSerum ImmunoglobulinUrine FLCSerum FLC#(%) of PatientsYESYESno2(10)YESnoYES6(29)YESYESYES5(24)Nonono7(33)YESYES11(48)DiscussionWhile serum FLC is adequate in the majority of patients for monitoring monoclonal gammopathies, urine FLC correlates as well as serum FLC in about ½ of the patients. In addition, in a small number of individuals, urine FLC correlates with serum total serum immunoglobulin better than serum FLC. We feel that random urine FLC is useful for monitoring monoclonal gammopathies, and in a minority of instances, provides more accurate assessment of disease activity than serum. DisclosuresNo relevant conflicts of interest to declare.

Patients with AL Amyloidosis and Low Free Light Chain Burden Have Distinct Clinical Features and Outcome

Background: In AL amyloidosis, circulating free light chains (FLC) are not only a clonal marker, but they are the causal agent of the disease. The recently validated criteria of hematologic response are based solely on the measurement of FLC. Patients with a difference between involved and uninvolved FLC (dFLC) <50 mg/L do not have measurable disease and cannot be assessed for response. Hence, they are commonly excluded from clinical trials. Nevertheless, these subjects represent a significant proportion of the patients suffering from AL amyloidosis, and given the lower burden of the toxic amyloidogenic precursor we hypothesize that they have distinctive clinical features.Patients and methods: The study population is composed of 984 consecutive, newly diagnosed patients with AL amyloidosis evaluated between 2004 and 2013 at the Pavia Amyloidosis Center and prospectively followed for response and survival. We compared the 187 subjects (19%) who had a baseline dFLC <50 mg/L (low-dFLC group) with the remaining 797 patients (evaluable-dFLC group).Results: Patients' characteristics are reported in the table. Heart involvement was less common in the low-dFLC group, and cardiac and renal dysfunction was more advanced in the evaluable-dFLC group, whereas renal involvement was more frequent and severe in the low-dFLC group. The Mayo clinic staging system based on NT-proBNP and cardiac troponin was able to discriminate three groups with significantly different outcomes in both groups (P<0.001). Overall survival was significantly better in the low-dFLC group (median 90 vs. 48 months, P<0.001). Within each Mayo stage patients with low-dFLC had a longer survival (at 3 years, 88% vs. 78%, P=0.03 in stage I, 69% vs. 45%, P=0.001 in stage II, 36% vs. 22%, P=0.02 in stage III). Complete response was associated with a significant survival advantage in the low-dFLC group (median 122 months vs. 92 months, P=0.02).Conclusions: Nineteen percent of newly diagnosed patients with AL amyloidosis have very low dFLC burden. They represent a subgroup with a distinct and better outcome compared to other patients. Nevertheless, complete response can still significantly improve survival. Patients with low baseline dFLC can be included in clinical trials with appropriate stratification and are evaluable for complete response.Table 1Patients' characteristics.VariableLow dFLC group 187 patients N (%) - median (range)Evaluable dFLC group 797 patients N (%) - median (range)PAge, years66 (40-85)65 (30-87)0.124Male sex99 (52)444 (55)0.272Organ involvement Heart Kidney Liver >2 organs88 (47) 142 (76) 26 (13) 99 (53)662 (83) 511 (64) 112 (14) 566 (71)<0.001 0.001 0.532 <0.001Mayo Stage 1 2 369 (37) 91 (49) 27 (14)109 (12) 338 (42) 350 (44)<0.001 0.071 <0.001NYHA class 336 (19)331 (40)<0.001NT-proBNP >8500 ng/L12 (6)212 (27)0.0001Renal Stage 1 2 362 (33) 85 (45) 40 (21)384 (48) 320 (40) 93 (11)0.001 0.107 0.001Alkaline phosphatase, U/L144 (82-215)159 (99-239)0.077Involved ligh-chain type κλ37 (20) 150 (80)190 (24) 607 (76)0.02 0.13Bone marrow plasma cells, %8 (5-12)12 (8-20)0.001Melphalan-dexamethasone (MDex) Cyclophosphamide-bortezomib-Dex Bortezomib-MDexOthers67 (35) 41 (22) 19 (10) 60 (33)289 (36) 207 (26) 102 (12) 199 (26)0.491 0.145 0.194 0.030 DisclosuresMerlini:Millennium-Takeda: Honoraria; Janssen-Cilag: Honoraria; Pfizer: Honoraria; Prothena: Honoraria.

Effect of specimen type on free immunoglobulin light chains analysis on the Roche Diagnostics cobas 8000 analyzer.

The measurement of free immunoglobulin light chains is typically performed on serum; however, the use of alternative specimen types has potential benefits. Using the Freelite™ kappa and lambda free light chains assay on a Roche Diagnostics cobas 8000 c502 analyzer, we compared three specimen types (serum, EDTA-plasma and lithium heparin plasma separator gel-plasma) on 100 patients. Using Deming regression and eliminating outliers (limiting data to light chain concentrations below 400 mg/L), the three specimen types showed comparable results for kappa light chain concentration, lambda light chain concentration, and kappa/lambda ratio with slopes close to 1.0 and y-intercepts close to zero. EDTA-plasma showed slightly more positive bias relative to serum than lithium heparin. Analysis using EDTA-plasma and lithium heparin plasma showed comparable linearity, precision, and temperature stability. A single sample showing hook effect (not in the comparison set) gave comparable results using either plasma specimen type. For the Freelite™ kappa and lambda free light chains assay, both EDTA-plasma or lithium heparin-plasma can serve as acceptable substitutes for serum, at least for the Roche cobas 8000 analyzer.

A Prospective Phase II Trial of Lenalidomide and Dexamethasone (Len-Dex) in POEMS Syndrome

Abstract Background. POEMS syndrome is a rare form of B cell dyscrasia combining a proliferation usually of plasma cells, a polyneuropathy, osteocondensing bone lesions and multiple other clinical signs. The pathogenesis is not well understood but VEGF plays a major role. In patients with one or two sclerotic plasmacytoma and no bone marrow involvement, first line therapy should include radiation. For patients with diffuse sclerotic lesions, bone marrow involvement or absence of any bone lesion and for those who have not demonstrated stabilization of their disease 3 to 6 months after completing radiation systemic therapy is indicated, the most effective being high dose chemotherapy with autologous stem cell transplant (ASCT). Radiation of a single lesion is effective in about every other case and is accompanied by a fairly slow improvement of the neurological symptoms, often after initial worsening. ASCT seems to be accompanied by a number of important complications, in particular engraftment syndrome. Outside these 2 treatments there is no consensus therapy. Lenalidomide (LEN), a drug without serious neurological toxicity, has the advantage of being both anti-angiogenic and cytotoxic to malignant plasma cells (Richardson PG, Blood 2002;100(9):3063-7). We have recently reported a series of 20 French patients with POEMS syndrome treated by LEN with a good efficacy. We now report the first 27 patients of a prospective phase II trial using LEN + dexamethasone (LEN-DEX), 2 cycles preceding radiation or high dose treatment trying to obtain a rapid clinical response and to avoid engraftment syndrome or 9 cycles followed by 1 year LENalone in patients who cannot receive radiation or ASCT. Methods. Newly diagnosed or relapsing patients with POEMS syndrome who needed to be treated were eligible. Patients who can be treated by local radiation or intensive treatment with stem cell support receive two 28 day cycles of LEN 25 mg PO Days 1-21 and DEX 40 mg PO Days 1,8,15,22 before radiation or intensive treatment (Group 1), the other patients receive 9 cycles of the same LEN-DEX (Group 2) and then 12 cycles of continuous low dose LEN (10 mg). LEN dose was tapered to 10 mg for patients with a creatinine clearance between 30 and 50 ml/min and DEX to 20 mg for patients above 75 years of age and for those who were frail patients. Main eligibility criteria included a diagnosis of POEMS syndrome according to criteria by Dispenzieri et al (Am J Hematol 2012;87(8):804-14), an age of 18 or more, a creatinine clearance above 30 ml/min, no prior treatment with or contraindication to LEN and no uncontrolled thrombosis. Serum and plasma VEGF, serum electrophoresis, immunofixation and free light chain measurements were centrally monitored. Neurologic evaluations were performed using the Overall Neuropathy Limitations Scale (ONLS), the Neurological Impairment Scale (NIS) and the 10 meter walk test (10MWT). The primary endpoint was evaluation of the effectiveness of LEN-DEX combination using biological responses (decrease of monoclonal protein and serum VEGF level) and secondary endpoints were clinical and particularly neurological responses. Results. Twenty-seven patients have been included in 12 centres, median age was 61 (range 32-75), the median follow-up was 6.6 months (range 2-24). Eighteen patients were in group 1, with radiotherapy in 10 patients and ASCT in 8 patients; 9 patients were in group 2. Nineteen patients were in first line and 8 already treated. Only 2 patients experienced grade 3-4 adverse events due to LEN (cytopenia) and 2 patients had allergic rashes, no thrombotic event occurred. No engraftment syndrome was noted in the 5 patients already treated with ASCT. To date, no patient have died. Evolution of VEGF median values in serum and plasma, M-spike and dFLC levels and evolution of neurological measurements are reported in table 1. Neurological improvement was very rapid in some patients, using ONLS and 10MWT 11/18 evaluable patients had a neurological improvement after 2 cycles with an improvement of 1 or more of the ONLS score and/or change of 0.1 m/s or more in the 10MWT. Only one patient who progressed after nine cycles received another therapy. Conclusion. This is the first prospective trial of LEN-DEX in POEMS syndrome. This combination seems well tolerated in this disease with a good efficacy on VEGF measurements and rapid neurological improvement in the majority of patients. Updated data will be presented at the meeting. Download : Download high-res image (32KB) Download : Download full-size image Figure 1 . Disclosures Jaccard:Celgene: Drug supply to Trial Other. Tournilhac:mundipharma: Honoraria, Other, Research Funding; GSK: Honoraria, Other, Research Funding; Roche: Honoraria, Other, Research Funding. Moreau:celgene: Honoraria, Membership on an entity’s Board of Directors or advisory committees.

The treatment strategies in patients with partial response to high-dose chemotherapy and ASCT

FH Campus Vienna, University of Applied Sciences, Vienna, Austria; Department of Laboratory Medicine, Wilhelminenhospital, Vienna; Wilhelminen Cancer Research Institute, c/o Department of Medicine I, Center of Oncology, Hematology with Outpatient Department and Palliative Care, Wilheminenhospital, Vienna, Austria Background: Up to 10% of newly diagnosed multiple myeloma (MM) patients present with oligosecretory disease. In these instances serum free light chain (FLC) measurements are recommended for monitoring M-Ig changes; provided FLC ratio is abnormal and involved FLC (iFLC) >100 mg/L at presentation. Novel immunoassays have become available that measure immunoglobulin heavy/light chain (HLC) subsets and may offer a sensitive method of monitoring oligosecretory patients. Patients and Methods: FLC and and HLC Ig’ and Ig’ were quantified turbidimetrically in sequential sera from 16 oligosecretory MM patients (6 IgG and 10 IgA) assessed at Wilhelminenspital, Vienna, Austria. Median age was 72 (53-90) years and median follow-up 992 (2533743) days. 9/16 patients were ISS stage 1, 3/16 were ISS stage 2 and 4/16 were ISS stage 3. HLC measurements were compared to serum protein electrophoresis (SPE) and nephelometric total Ig’ concentrations. Results: In 6/16 patients the M-Ig bands were nonquantifiable by SPEP and in the remaining 10/16 patients the M-Ig concentrations were 100 mg/L; and could be monitored by serum FLC measurements. For the remaining 8/16 patients none of the standard techniques provided information for monitoring M-Ig changes. All 8/8 patients had an abnormal HLC ratio and 5/8 had elevated involved HLC (iHLC) concentrations (IgA n1⁄43, HLC ratio: 6.7 (2.9-85.5), iHLC: 7.0 (3.1-8.6) g/L; IgA n1⁄42, HLC ratio: 0.08 and 0.05; iHLC: 5.9 and 5.0 g/L). In these 5 patients there was overall concordance between changes in HLC ratio /dHLC (involved-uninvolved HLC) and clinical assessment. Importantly in 2/3 patients an increasingly abnormal HLC ratio and dHLC preceded relapse; including 1 IgA patient where immunofixation remained negative and total IgA concentrations within the normal range for 12 months before progression. Conclusion: HLC immunoassays may offer a sensitive method of quantifying M-Ig concentrations in oligosecretory MM patients. Further studies on larger cohorts and comparison with clinical assessment are necessary to validate the assays for monitoring these patients.

Utility of Serum Free Light Chain Measurements in Multiple Myeloma Patients Not Achieving Complete Response

![Graphic][1] Background: Normalization of serum free light chain ratio (FLCr) following achievement of a complete response (CR) to therapy denotes a stringent CR in multiple myeloma, and is associated with improved overall survival (OS). However, its value in patients achieving less than a CR is not clear. We hypothesized that patients in whom FLCr normalizes with the initial anti-myeloma therapy will have an improved outcome, even in the absence of a CR. Methods: We analyzed the medical records of patients with MM who were evaluated at Mayo Clinic, Rochester, MN between January 2004 and December 2011, seen within 90 days of diagnosis. We selected patients with measurable disease on serum protein electrophoresis, defined as a serum M-spike ≥1 g/dl at the time of diagnosis. Then, we excluded all patients with negative immunofixation for monoclonal M-protein in serum and urine at their first response as well as those who did not have serum FLC results available from the time of their first best response. M protein measurements were collected serially from the time of diagnosis until the time of the first relapse or last follow up date if no relapse was recorded. Progression-free survival (PFS) and OS were analyzed using the Kaplan-Meier method. Results: Out of the 1354 MM patients seen at our institution during this period, a total of 453 patients were included in our study. The median age of the cohort at the time of diagnosis was 65 (range, 29-91); 49% were older than 65 years and 284 (63%) were male. The estimated median follow up for the whole cohort was 66 months (95% CI; 61, 71); 241 patients were alive with a median follow up of 4.7 years (range, 0.4 to 10). Three hundred and seventy three (82%) patients had relapsed following the initial therapy with a median time to progression (TTP) of 19.5 months (95% CI; 18, 22) and PFS of 19 months (95% CI; 17, 20). The median OS for the entire cohort is 67 months (95% CI; 59, 73). The median time to the best first response was 6.9 months (range: 0.3-83 months). At the time of the best first response 154 patients (34%) had normal FLCr. Patients with normal FLCr had a longer PFS (29 vs. 16 months, P< .0001) and OS (91 vs. 58 months, P= .0002). We then examined the impact of FLCr normalization on survival outcomes according to their IMWG response categories. We found that FLCr normalization had a favorable impact on PFS in each of the three groups [very good partial response (VGPR), Partial response (PR), and Stable disease (SD)], However, when looking at the OS, only patients with PR had a better OS associated with a normal sFLC κ/λ ratio ( Table 1) . Conclusions: Obtaining a normal sFLC ratio appears to confer a more favorable prognosis irrespective of the depth of the response, suggesting an important role for serial-sFLC measurements in disease monitoring even in patients who achieve only a PR to therapy as their best response. This supports the inclusion of sFLC at all levels of response included in the current International Myeloma Working Group (IMWG) criteria. | | SD | PR | VGPR | | --------------------------- | ------------- | ----------- | ----------- | ------------- | | Normal FLCr | Abnormal FLCr | p-value | Normal FLCr | Abnormal FLCr | p-value | Normal FLCr | Abnormal FLCr | p-value | | Number of patients | 7 | 70 | | 62 | 157 | | 85 | 60 | | | PFS, month, median (95% CI) | 27 (14, 36) | 7 (5, 12) | < .02 | 26 (20, 36) | 17 (13, 20) | .0005 | 29 (23, 32) | 21 (18, 26) | .03 | | OS, months, median (95% CI) | 68 (49, 68) | 41 (31, 61) | > .1 | NR (65, NR) | 61 (50, NR) | .004 | 74 (67, 107) | 73 (59, NR) | > .9 | Abstract 3427. Table 1 Effect of FLCr normalization according to IMWG response categories • Abbreviations: NR; statistically not reached •• Progressive disease (PD) was not included because none of the 12 patients achieved a normal FLCr. Disclosures No relevant conflicts of interest to declare. [1]: /embed/inline-graphic-2.gif

Utility of serum free light chain measurements in multiple myeloma patients not achieving complete response to therapy.

Normalization of the serum free light chain ratio (FLCr) with the absence of bone marrow monoclonal plasma cells following achievement of a complete response (CR) to therapy denotes a stringent CR in multiple myeloma (MM), and is associated with improved overall survival (OS). However, its value in patients achieving <CR is not clear. We hypothesized that patients achieving a normalization of FLCr with initial therapy of MM will have an improved outcome, even in the absence of a CR. We retrospectively evaluated 449 patients with newly-diagnosed MM with measurable disease at baseline, who did not achieve a CR with initial therapy. One hundred fifty three patients (34%) had a normal FLCr while 296 (66%) had an abnormal ratio. Patients with a normal FLCr had a longer progression-free survival (PFS) (29 vs. 16 months, P <.001) and OS (91 vs. 58 months, P <.001). Normalization of FLCr retained its prognostic value in a multivariable model. Our results suggest an important role for sFLC measurement in disease monitoring even in patients who achieve only a partial response to therapy. Obtaining a normal FLCr confers a favorable prognosis independent from other factors, supporting the inclusion of sFLC in all levels of response criteria.

Detecting monoclonal immunoglobulins in human serum using mass spectrometry

Established guidelines from the International Myeloma Working Group recommend diagnostic screening for patients suspected of plasma cell proliferative disease using protein electrophoresis (PEL), free light chain measurements and immunofixation electrophoresis (IFE) of serum and urine in certain cases. Plasma cell proliferative disorders are generally classified as monoclonal gammopathies given most are associated with the excess secretion of a monoclonal immunoglobulin or M-protein. In clinical practice, the M-protein is detected in a patients’ serum by the appearance of a distinct protein band migrating within regions typically occupied by immunoglobulins. Given each M-protein is comprised by a sequence of amino acids pre-defined by somatic recombination unique to each clonal plasma cell, the molecular mass of the M-protein can act as a surrogate marker. We established a mass spectrometry based method to assign molecular mass to the immunoglobulin light chain of the M-protein and used this to detect the presence of M-proteins. Our method first enriches serum for immunoglobulins, followed by reduction to separate light chains from heavy chains, followed by microflow LC-ESI-Q-TOF MS. The multiply charged light chain ions are converted to their molecular mass and reconstructed peak area calculations are used for quantification. Using this method, we term “monoclonal immunoglobulin Rapid Accurate Molecular Mass” or miRAMM, the presence of M-proteins can be reliably detected with superior sensitivity compared to current gel-based PEL and IFE techniques.

Assessment of free light chains in HCV-positive patients with mixed cryoglobulinaemia vasculitis undergoing rituximab treatment.

Mixed cryoglobulinaemia (MC) is an HCV-related lymphoproliferative disorder characterized by the presence of circulating immune complexes called cryoglobulins. Treatment with anti-CD20 monoclonal antibody rituximab is proved to be very useful, especially in patients ineligible to interferon-based antiviral therapy. Recently, free light chain (FLC) κ/λ ratio and FLC patterns were associated with MC. The aim of this study was to evaluate changes in FLC-κ, FCL-λ, FLC ratio following rituximab treatment in patients with HCV-related MC and to correlate FLC-κ, FCL-λ and FLC ratio values with therapy response. We retrospectively enrolled 46 patients with HCV infection (26 females, 20 males), including 10 patients without signs/symptoms of MC-related vasculitis, 36 with MC vasculitis. Clinical and biological data were recorded at baseline and 6months after RTX treatment. Nephelometric measurement of serum FLCs was taken. The mean serum FLC-κ level and FLC ratio were significantly higher in patients with MC, compared to HCV patients without MC and to blood donors. An abnormal FLC ratio at baseline correlated with the presence of cryoglobulins, C4 consumption, higher RF level and higher vasculitis rate. To evaluate the predictive value of FLCs, patients with MC were divided into two groups according to RTX therapy outcome (responders and no/partial responders). Abnormal baseline FLC ratio was significantly associated with no/partial response. RTX treatment in HCV-related MC induces a reduction in FLC-κ and RF levels. Moreover, pretreatment FLC ratio, which can be easily assessed by a routine test, may be useful to predict response to this expensive treatment for patients with HCV-related MC ineligible to IFN-based therapy.

Kinetics of organ response and survival following normalization of the serum free light chain ratio in AL amyloidosis

Despite successful treatment of the clonal plasma cell implicated in its pathogenesis, patients with AL amyloidosis (AL) experience significant morbidity related to underlying amyloid mediated organ dysfunction. While normalization of the serum free light chain measurements [normal ratio of involved and uninvolved free light chains (nFLCr)] is the goal of therapy and centerpiece of hematologic response criteria, achieving (or not achieving) meaningful organ response (OR) is clinically significant for its implications on long-term symptomatology as well as overall survival (OS), and remains the ultimate goal of treatment. Expectations for organ recovery following successful therapy leading to nFLCr in AL remain poorly described. We evaluated the timeframe and predictive factors for OR, and long-term outcome, in 313 AL patients who achieved nFLCr following therapy initiation. OR was seen in 80% of surviving AL patients within 1-year of nFLCr. Patients achieving early OR within 1 year of nFLCr had superior OS compared with those who despite obtaining nFLCr did not achieve early OR. We further evaluated factors predicting OR and OS among patients achieving nFLCr. Higher values of dFLC (involved-uninvolved) at diagnosis predict OR, and early OR predicts improved OS following successful hematologic therapy in AL.

Elevation of serum immunoglobulin free light chains during the preclinical period of rheumatoid arthritis.

Immunoglobulin free light chains (FLC) represent biomarkers of B cell activity in rheumatoid arthritis (RA) and are associated with all-cause mortality in the general population. Our objective was to evaluate the relationships of serum FLC to preclinical disease, RA characteristics, and mortality in RA compared to non-RA subjects. A population-based study in Olmsted County, Minnesota, USA, was performed by crosslinking a large cohort in the general population having available serum FLC measurements with established RA incidence and prevalence cohorts. Serum κ, λ, and total FLC and their trends relative to RA incidence were compared between RA and non-RA subjects. Regression models were used to determine the associations between FLC, disease characteristics, and mortality, testing for differential effects of FLC on mortality in RA. Among 16,609 subjects, 270 fulfilled the criteria for RA at the time of FLC measurement. Mean total FLC were significantly higher in RA compared to non-RA subjects (4.2 vs 3.3 mg/dl, p < 0.001). FLC became elevated 3-5 years before the clinical onset of RA and remained elevated during followup. Polyclonal FLC were found to predict higher mortality in persons with RA, though elevation to the highest decile had a relatively lower effect on mortality in RA compared to non-RA subjects. Elevation of serum FLC precedes the development of RA and may be useful in monitoring B cell activity and disease progression. FLC are associated with mortality among patients with RA as well as the general population.

Clinical recommendations for the measurement of serum free light chains and the emerging role of heavy/light chain pair analysis in the management of monoclonal gammopathies: When and how to use it?

Protein and immunofixation electrophoresis are the laboratory backbone assays for testing multiple myeloma (MM) and related plasma cell proliferative diseases. The secreted monoclonal proteins act as serological tumor markers, and the electrophoretic assays identify monoclonal proteins for diagnosis as well as quantitate these markers for monitoring clinical course and response to therapy. However these techniques present limitations, especially in patients with nonsecretory MM, light chain MM, and amyloid light chain amyloidosis. Freelite, a nephelometric/turbidimetric serum assay for the measurement of serum free light chains, has improved the management of patients with monoclonal gammopathies and has been included in international myeloma working group guidelines for the screening, prognostic, monitoring and as response criteria. Recently, a new assay, Hevylite; , was also approved by Food and Drug Administration for the monitoring of IgA and IgG MM. It allows the quantification of each immunoglobulin subtype (i.e., IgGκ, IgGλ, IgAκ, IgAλ), enables analysis of specific involved/monoclonal and uninvolved/polyclonal immunoglobulins heavy/light chain pairs, which produce immunoglobulin heavy/light chain ratios and may improve sensitivity for identifying residual disease and relapse in some patients. Together, these assays are important tools which will help us to achieve the correct diagnostic and follow-up of B cell dyscrasias.

A Prospective Phase II Trial of Lenalidomide and Dexamethasone ( LEN-DEX) in POEMS Syndrome

Background. POEMS syndrome is a rare form of B cell dyscrasia combining a proliferation usually of plasma cells, a polyneuropathy, osteocondensing bone lesions and multiple other clinical signs. The pathogenesis is not well understood but VEGF plays a major role. In patients with one or two sclerotic plasmacytoma and no bone marrow involvement, first line therapy should include radiation. For patients with diffuse sclerotic lesions, bone marrow involvement or absence of any bone lesion and for those who have not demonstrated stabilization of their disease 3 to 6 months after completing radiation systemic therapy is indicated, the most effective being high dose chemotherapy with autologous stem cell transplant (ASCT). Radiation of a single lesion is effective in about every other case and is accompanied by a fairly slow improvement of the neurological symptoms, often after initial worsening. ASCT seems to be accompanied by a number of important complications, in particular engraftment syndrome. Outside these 2 treatments there is no consensus therapy. Lenalidomide (LEN), a drug without serious neurological toxicity, has the advantage of being both anti-angiogenic and cytotoxic to malignant plasma cells (Richardson PG, Blood 2002;100(9):3063-7). We have recently reported a series of 20 French patients with POEMS syndrome treated by LEN with a good efficacy. We now report the first 27 patients of a prospective phase II trial using LEN + dexamethasone (LEN-DEX), 2 cycles preceding radiation or high dose treatment trying to obtain a rapid clinical response and to avoid engraftment syndrome or 9 cycles followed by 1 year LENalone in patients who cannot receive radiation or ASCT.Methods. Newly diagnosed or relapsing patients with POEMS syndrome who needed to be treated were eligible. Patients who can be treated by local radiation or intensive treatment with stem cell support receive two 28 day cycles of LEN 25 mg PO Days 1-21 and DEX 40 mg PO Days 1,8,15,22 before radiation or intensive treatment (Group 1), the other patients receive 9 cycles of the same LEN-DEX (Group 2) and then 12 cycles of continuous low dose LEN (10 mg). LEN dose was tapered to 10 mg for patients with a creatinine clearance between 30 and 50 ml/min and DEX to 20 mg for patients above 75 years of age and for those who were frail patients. Main eligibility criteria included a diagnosis of POEMS syndrome according to criteria by Dispenzieri et al (Am J Hematol 2012;87(8):804-14), an age of 18 or more, a creatinine clearance above 30 ml/min, no prior treatment with or contraindication to LEN and no uncontrolled thrombosis. Serum and plasma VEGF, serum electrophoresis, immunofixation and free light chain measurements were centrally monitored. Neurologic evaluations were performed using the Overall Neuropathy Limitations Scale (ONLS), the Neurological Impairment Scale (NIS) and the 10 meter walk test (10MWT). The primary endpoint was evaluation of the effectiveness of LEN-DEX combination using biological responses (decrease of monoclonal protein and serum VEGF level) and secondary endpoints were clinical and particularly neurological responses.Results. Twenty-seven patients have been included in 12 centres, median age was 61 (range 32-75), the median follow-up was 6.6 months (range 2-24). Eighteen patients were in group 1, with radiotherapy in 10 patients and ASCT in 8 patients; 9 patients were in group 2. Nineteen patients were in first line and 8 already treated. Only 2 patients experienced grade 3-4 adverse events due to LEN (cytopenia) and 2 patients had allergic rashes, no thrombotic event occurred. No engraftment syndrome was noted in the 5 patients already treated with ASCT. To date, no patient have died. Evolution of VEGF median values in serum and plasma, M-spike and dFLC levels and evolution of neurological measurements are reported in table 1. Neurological improvement was very rapid in some patients, using ONLS and 10MWT 11/18 evaluable patients had a neurological improvement after 2 cycles with an improvement of 1 or more of the ONLS score and/or change of 0.1 m/s or more in the 10MWT. Only one patient who progressed after nine cycles received another therapy.Conclusion. This is the first prospective trial of LEN-DEX in POEMS syndrome. This combination seems well tolerated in this disease with a good efficacy on VEGF measurements and rapid neurological improvement in the majority of patients. Updated data will be presented at the meeting. [Display omitted] DisclosuresJaccard:Celgene: Drug supply to Trial Other. Tournilhac:mundipharma: Honoraria, Other, Research Funding; GSK: Honoraria, Other, Research Funding; Roche: Honoraria, Other, Research Funding. Moreau:celgene: Honoraria, Membership on an entity’s Board of Directors or advisory committees.

Prognostic Implications of Oligoclonal Bands in Patients with Multiple Myeloma:Its Development per Se did Not Confer the Prologation of Survival

Background: Multiple myeloma (MM) is characterised by the production of monoclonal immunoglobulin (Ig) and clonal proliferation of neoplastic plasma cells in bone marrow. The emergence of oligoclonal bands (OB) in serum and/or urine immunofixation electrophoresis (IFE) that is different to that observed at diagnosis has been reported with varying frequency in patients with stem cell transplantation (SCT) or favourable response to chemotherapy, although its prognostic relevance remains unclear. Here, we retrospectively analysed clinical records and results serial serum and/or urine IFE to determine the frequency, clinical characteristics and prognostic impact of OB that developed after treatment. We also analysed the effects of OB on the results of free light chain (FLC) assay that may affect the stringent CR (sCR) criteria proposed by the IMWG.Patients: We retrospectively reviewed 180 patients with MM admitted to the Department of Hematology/Oncology at Kameda Medical Center, Kamogawa, Japan, from January 2006 to May 2014.Seventeen patients were excluded from the analysis due to lack of appropriate follow-up data.Method: An OB was defined as the presence of a serum and/or urine IFE monoclonal spike that was different from the original myeloma protein in heavy and/or light chains, as well as a different IFE migration pattern. IFE was performed at least every three months after obtaining very good partial response (VGPR) and then until the disease progression or relapse was confirmed. Overall survival (OS) and progression free survival (PFS) were analyzed in 173 MM patients by the Kaplan–Meier method, and differences between the curves were calculated by two-sided log-rank test. Free light chain (FLC) and minimal residual disease measurement by multicolour flowcytometory (MFC) were performed to evaluate the response to treatment.Results: Myeloma response more than VGPR and CR were achieved in 87 (53.3%) and 54 (33.1%) patients, respectively. None of the patients with less than PR developed OB, and OB developed in 36.3% (12/33) and 51.9% (28/54) of the patients with VGPR and CR, respectively. Among the 30 patients who received SCT, 18 patients (60.0%) developed OB, whereas only 12 of 133 patients who did not receive SCT developed OB. The emergence of OB was significantly higher in patients receiving SCT (P < 0.001). The median PFS of the patients with and without OB were 79.4 months and 19.2 months (P = 0.028), respectively, and those of OS did not reached and 41.1 months (P < 0.001), respectively (Figure 1). However, if patient response was limited to CR, median PFS and OS of patients with and without OB were not significantly different between the two groups (79.4 months vs. 80.1months, respectively, P = 0.646; and not reached and not reached, respectively, P = 0.776; Figure 2). OB was observed in 60% of patients after SCT, and in 36.6% of patients more than VGPR without SCT (P<0.001). In the patients who received SCT, the presence of OB was not associated with either PFS or OS (20.0 months vs 44.7 months, P=0.543 and not reached vs not reached, P= 0.739, respectively; Figure3). We also analyzed the association between residual myeloma cells assessed by the MFC in the patients who achieved CR according to the development of OB. Although the patients with OB showed slightly lower levels of residual myeloma cells than those without OB (2.49 x10-4vs 4.15 x10-4), there was no significant difference between the two groups (P= 0.054). As the presence of OB may result in an abnormal serum FLC kappa/lambda ratio, we examined the association of FLC kappa/lambda ratio and development of OB in 28 patients who achieved CR. Abnormal FLC kappa/lambda ratio was observed in 10 (35.7%) and 4 (15.3%) patients with and without OB, respectively, among those with CR (P = 0.118). Conversely, OB was detected in 10 of the 14 patients with an abnormal FLC ratio (71.4%) versus 18 of 40 (45.0%) with a normal serum FLC ratio (P = 0.164) among those with CR.Conclusions: In conclusion, we showed that the emergence of OB occurred exclusively in patients with favourable myeloma responses (more than VGPR) and was associated with prolonged survival. Patients with OB appeared to have fewer residual myeloma cells compared to those without OB. However, its development did not confer an additional survival benefit among the patients who achieved CR. In addition, it is possible that the emergence of OB affects the sCR response criteria proposed by the IMWG. [Display omitted] [Display omitted] [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Utility of Serum Free Light Chain Measurements in Multiple Myeloma Patients Not Achieving Complete Response

▪Background: Normalization of serum free light chain ratio (FLCr) following achievement of a complete response (CR) to therapy denotes a stringent CR in multiple myeloma, and is associated with improved overall survival (OS). However, its value in patients achieving less than a CR is not clear. We hypothesized that patients in whom FLCr normalizes with the initial anti-myeloma therapy will have an improved outcome, even in the absence of a CR.Methods: We analyzed the medical records of patients with MM who were evaluated at Mayo Clinic, Rochester, MN between January 2004 and December 2011, seen within 90 days of diagnosis. We selected patients with measurable disease on serum protein electrophoresis, defined as a serum M-spike ≥1 g/dl at the time of diagnosis. Then, we excluded all patients with negative immunofixation for monoclonal M-protein in serum and urine at their first response as well as those who did not have serum FLC results available from the time of their first best response. M protein measurements were collected serially from the time of diagnosis until the time of the first relapse or last follow up date if no relapse was recorded. Progression-free survival (PFS) and OS were analyzed using the Kaplan-Meier method.Results: Out of the 1354 MM patients seen at our institution during this period, a total of 453 patients were included in our study. The median age of the cohort at the time of diagnosis was 65 (range, 29-91); 49% were older than 65 years and 284 (63%) were male. The estimated median follow up for the whole cohort was 66 months (95% CI; 61, 71); 241 patients were alive with a median follow up of 4.7 years (range, 0.4 to 10). Three hundred and seventy three (82%) patients had relapsed following the initial therapy with a median time to progression (TTP) of 19.5 months (95% CI; 18, 22) and PFS of 19 months (95% CI; 17, 20). The median OS for the entire cohort is 67 months (95% CI; 59, 73). The median time to the best first response was 6.9 months (range: 0.3-83 months). At the time of the best first response 154 patients (34%) had normal FLCr. Patients with normal FLCr had a longer PFS (29 vs. 16 months, P< .0001) and OS (91 vs. 58 months, P= .0002).We then examined the impact of FLCr normalization on survival outcomes according to their IMWG response categories. We found that FLCr normalization had a favorable impact on PFS in each of the three groups [very good partial response (VGPR), Partial response (PR), and Stable disease (SD)], However, when looking at the OS, only patients with PR had a better OS associated with a normal sFLC κ/λ ratio (Table 1).Conclusions: Obtaining a normal sFLC ratio appears to confer a more favorable prognosis irrespective of the depth of the response, suggesting an important role for serial-sFLC measurements in disease monitoring even in patients who achieve only a PR to therapy as their best response. This supports the inclusion of sFLC at all levels of response included in the current International Myeloma Working Group (IMWG) criteria.Abstract 3427. Table 1Effect of FLCr normalization according to IMWG response categoriesSDPRVGPRNormal FLCrAbnormal FLCrp-valueNormal FLCrAbnormal FLCrp-valueNormal FLCrAbnormal FLCrp-valueNumber of patients770621578560PFS, month, median (95% CI)27 (14, 36)7 (5, 12)< .0226 (20, 36)17 (13, 20).000529 (23, 32)21 (18, 26).03OS, months, median (95% CI)68 (49, 68)41 (31, 61)> .1NR (65, NR)61 (50, NR).00474 (67, 107)73 (59, NR)> .9- Abbreviations: NR; statistically not reached-- Progressive disease (PD) was not included because none of the 12 patients achieved a normal FLCr. DisclosuresNo relevant conflicts of interest to declare.

Comparing the Performance of Serum Free Light Chain Measurements with Urine Electrophoresis and Immunofixation for Monitoring and Assessing Response to Therapy in Patients with Multiple Myeloma

The introduction of the serum free light chain (sFLC) changed the diagnostic paradigm for patients with B cell disorders. IMWG guideline recommends the assay as a replacement for 24h urine at diagnosis, however with the exception of oligosecretory disease the assay is not recommended as a tool to monitor patients. One rationale for this recommendation is that to date, studies have compared the concentrations of FLC as measured by the two tests rather than determine which test provides the more reliable clinical assessment. Here we compare the sensitivities of FLC and 24h urine and comment on the reliability of each to monitor patients.Sequential sera from 25 LCMM (14 FLCκ, 11 FLCλ; stage I: 10, II: 10, III: 5) and 157 IIMM patients (79 IgGκ, 34 IgGλ, 26 IgAκ, 18 IgAλ; Stage I: 46, II: 75, III: 35, 1 missing) enrolled onto the IFM 2007-02 MM trial were analysed. Serum FLCκ and FLCλ levels were measured by Freelite® in samples collected at presentation, after cycles 2 and 4 of therapy and post ASCT. Results were compared to previously published sFLC reference ranges (sFLCκ 3.3-19.4 mg/L, sFLCλ 5.7-26.3 mg/L, sFLCκ/λ ratio 0.26-1.65), SPEP, UPEP sIFE and uIFE. IMWG guidelines were used to define measurable disease and to assess response the therapy. Quadratic Weighted Kappa (WK) analysis was performed to assess agreement in responses assigned by sFLC and urine tests.All 25 LCMM patients had abnormal sFLC ratios (14 FLCκ, 11 FLCλ) and measurable disease at presentation (iFLC 3620 (689-22000) mg/L). Similarly, all patients were positive by uIFE and had measurable disease by UPE (1940 (490-42000) mg/24h). However, in keeping with previous reports quantitative correlation between the two assays was poor (r=0.27).Responses assigned by sFLC and UPEP were concordant in 11/25 (44%) patients, although in 4/11 (40%) timing of the response was different (UPEP 89 (58-118) days; sFLC 226 (216-227) days). In the remaining 14/25 patients the responses assigned using the two tests differed. In 7/14 patients UPEP and uIFE became negative whilst the FLC ratio remained abnormal; in 1/7 patient sIFE confirmed the presence of the M protein. In 3 patients FLC identified relapse, while UPEP was negative or indicated response. In a further 2 patients FLC identified no response whilst UPEP initially identified a response and subsequent relapse. Overall, a moderate concordance was identified between the responses assigned by sFLC and urine tests (WK (95% CI): 0.59 (0.36-0.82)).At presentation, 154/157 (98%) IIMM patients had abnormal FLC ratios (FLCκ ratio 57 (2-33191); FLCλ ratio 0.009 (0.00003-0.25)), whereas only 85/157 (54%) patients were positive by uIFE and 67/157 (43%) by UPEP. 98/157 (62%) had measurable disease using sFLC (κFLC 491 (101-15600) mg/L; λFLC 441 (101-14100) mg/L), and 55/157 (35%) had measurable disease by UPEP (1000 (210-9200) mg/24h). 53/157 (34%) patients had measurable disease by both methods. The correlation between sFLC and UPEP measurements was poor (r=0.36) as was the correlation between intact immunoglobulin measurements by SPEP and sFLC (r=-0.06) or UPEP (r=-0.26).In 53 IIMM patients with measurable disease by both FLC and UPEP, sFLC ratios normalised in 14/53 patients (in 8/14 sIFE remained positive) while uIFE became negative in 33/53 patients (20/33 remained sIFE positive). WK showed better agreement for response assignment between intact immunoglobulin and sFLC measurements (WK (95% CI): 0.63 (0.48-0.79); substantial agreement) than with urine tests (0.49 (0.27-0.72); moderate agreement). Additionally, there was an association between depth of response by sFLC pre- and post-transplant: patients achieving >VGPR before transplant were more likely to achieve >VGPR post-transplant compared with patients who achieved <VGPR prior to transplant (96.2% vs. 63.2%, respectively; p=0.001).Finally 5/157 IIMM patients were oligosecretory and had measurable levels of disease by both UPEP and sFLC, but not by SPEP. In all 5 patients UPEP became negative by cycle 2; however, an abnormal sFLC ratio and positive sIFE indicated persistent disease.sFLC was a more sensitive tool and showed a greater degree of concordance with IFE and SPEP than UPEP in LCMM and IIMM patients respectively during patient monitoring. Furthermore, >90% reduction in sFLC prior to transplant was associated with post-transplant response in IIMM patients. Larger studies with patient outcome are required to validate our findings. DisclosuresNo relevant conflicts of interest to declare.

Serum free light chains in patients with HIV infection: their association with markers of disease severity and antiretroviral use

AimSerum free light chain measurements are used to follow-up and manage patients with monoclonal gammopathies, and abnormal ratios are associated with risk of progression in certain diseases. B cell dysfunction...

Oligoclonal bands in patients with multiple myeloma: its emergence per se could not be translated to improved survival.

The emergence of oligoclonal bands (OB) has been reported in patients with multiple myeloma (MM) after stem cell transplantation (SCT) or successful chemotherapy. However, their clinical relevance remains unclear. We reviewed the clinical records of MM patients from January 2006 to May 2014. Treatment response was evaluated by International Working Group (IMWG) criteria. Serum immunofixation tests were performed at least every 3 months if the patient achieved more than very good partial response (VGPR). Free light chain (FLC) and minimal residual disease measurement by multicolor flow cytometry (MFC) were performed to evaluate the response to treatment. Among the 163 patients included in the study, 40 developed OB. Detection rates of OB in patients with complete response (CR), VGPR and partial response (PR) or less were 51.8, 36.3 and 0%, respectively. Patients with OB showed better progression-free survival (PFS) and overall survival (OS) rates than those without OB (P = 0.028 and P < 0.001, respectively). However, if the patients were limited to ≥VGPR or CR, development of OB did not affect PFS (P = 0.621 and P = 0.646, respectively) or OS (P = 0.189 and P = 0.766, respectively). OB was observed in 60% of patients after SCT, and in 36.6% of patients with more than VGPR without SCT (P < 0.001). Patients with OB tended to have less minimal residual disease than those without OB (P = 0.054) and its presence may affect the stringent CR criteria. In conclusion, the emergence of OB was seen exclusively in patients with favorable responses, but its emergence per se could not be translated to improved survival.