Murray, David

e20515 Background: IgA monoclonal proteins (MCPs), unlike IgG MCPs, often migrate in the beta region on serum protein electrophoresis (SPEP) which can lead to underestimation of their size due to the co-migration with physiologic proteins. In IgA multiple myeloma (MM), the utility of quantitative IgA levels in assessing disease response in comparison to SPEP is not well studied. Methods: We retrospectively analyzed 304 IgA MM patients, diagnosed between 2004 and 2018, with available serial MCP and quantitative IgA levels. Kaplan Meier analysis was used to estimate the median progression free survival (mPFS) using the IMWG criteria and our study definition of IgA progression (2 consecutive IgA values that are > 25% above the nadir IgA value and above upper limit of normal (ULN) of 356 mg/dL, and a detectable IgA MCP on serum immunofixation). The mPFS was defined as the time from treatment initiation until disease progression or death. Results: IgA MCP migrated in the beta region in 134 (44%) patients, and in the gamma region in 150 (56%) patients. At diagnosis the median MCP was 3 (IQR 1.9-4) g/dL and the median IgA was 3240 (IQR 2008-4420) mg/dL. The median time from treatment initiation to MCP nadir was 80 (IQR 42-144) days and median time to IgA nadir was 154 (IQR 90-238) days. At MCP nadir 40% of patients had an IgA above the ULN. All complete responders (n = 104) had normal IgA levels, with a median IgA of 54 (IQR 27-88) g/dL. A ≥90% decrease in IgA between treatment initiation and IgA nadir, compared to a < 90% decrease, was associated with a longer mPFS (34 vs. 20 months, p = 0.006) and overall survival (97 vs. 33 months, p = 0.003). Patients with serial MCP and IgA levels available prior to progression (n = 195) were used to compare the mPFS using the IMWG and IgA progression criteria. The mPFS using the IgA criteria was 32 (95% CI 29-39) months, versus 39 (95% CI 33-45) months using IMWG criteria. Overall, 92 (47%) patients progressed by both IMWG and IgA criteria. At the time of progression using the IgA criteria compared to at IMWG progression, the median hemoglobin was higher (13.3 vs. 11.6 g/dL, respectively, p < 0.001) and fewer patients had new symptomatic bone lesions (2% vs. 33%, respectively, p < 0.001). Conclusions: In IgA MM patients, monitoring quantitative IgA levels predicts disease response and allows for earlier detection of disease progression, prior to the development of end organ damage.

Key PointsPatients with an abnormal sFLC ratio because of LC suppression have outcomes similar to those with a normal ratio. sFLC ratio has prognostic value in patients achieving CR who have an absence of clonal BMPCs assessed by MFC.

Abstract Detection of an M-protein (monoclonal immunoglobulin) has been used to diagnose and monitor multiple myeloma (MM). As therapies for MM have improved, more sensitive methods have been used to define response: immunofixation electrophoresis (IFX) of serum and urine, normalization of the serum immunoglobulin free light chain (FLC) ratio, and high sensitivity flow cytometry to detect clonal plasma bone marrow cells. It is hoped that these more sensitive approaches will differentiate those patients with minimal residual disease (MRD) versus no residual disease (NRD), that later which could mean a cure. Flow cytometry of plasma cells requires bone marrow aspiration, which is inconvenient and expensive and is potentially limited by sampling bias. More sensitive methods to differentiate MRD from NRD using serum would be advantageous. We have developed a sensitive test for the presence of the monoclonal antibody produced by the plasma cells which may serve as a substitute for invasive bone marrow biopsy. Briefly, patient serum is enriched for immunoglobulins (Ig) and the Ig light chains are decoupled from the heavy chain by reduction with DTT. The mass distribution of the light chains is resolved using a micro LC-ESI-Q-TOF mass spectrometry and the presence of the M-protein is detected as a spike in the mass distribution. In addition to the detection, the accurate mass measurement of the light chain serves as a unique individualized marker which can aid in detection in subsequent patient monitoring. We have termed this method: monoclonal-immunoglobulin-Rapid-Accurate-Mass-Measurement (miRAMM). By spiking monoclonal immunoglobulins into human serum, we have demonstrated that miRAMM is approximately 1000x more sensitive than SPEP at detecting M-proteins. Initial results on 21 patients in stringent complete response (sCR) demonstrated that 67% (n=14) had detectable malignant specific clones by miRAMM. Given these promising results we have extended the method to a larger series of myeloma patients for whom we had long term follow-up and serum samples prior to treatment and at 6-12 months status post stem cell treatment. The results of this data will be correlated to the clinical outcomes for these patients. Early results demonstrate the potential of miRAMM to be a more sensitive, cost effective approach to detect MRD compared with current methods. Disclosures Murray: Mayo Clinic: Patent Application filed Patents & Royalties. Barnidge:Mayo Clinic: Patent filed Patents & Royalties.

Introduction: The clinical presentation and frequency of organ involvement are quite distinct in patients with IgM amyloidosis compared to patients with non-IgM amyloidosis, with less heart involvement and more nerve, soft tissue and lung involvement in IgM patients. It has been observed that immunoglobulin light chain variable region (IGVL) gene and gene family of the plasma cell clone impact phenotypic manifestations in AL amyloidosis, particularly organ tropism. (Kourelis et al. Blood. 2017;129). Given the variability in clinical features amongst patients with IgM vs. non IgM AL amyloidosis, we hypothesized that IGVL gene usage would be different between these two groups. Methods: Patients with newly diagnosed IgM and non-IgM amyloidosis seen at our institution from 01/2006 to 12/2015 were identified. IGVL gene usage was assessed by liquid chromatography tandem mass spectrometry (LC/MS/MS) as previously described. (Vrana et al Blood 2009;Dasari et al J of Proteome Res; Kourelis et al. Blood. 2017). Mass spectrometry data for IGVL analysis was obtained from clinically available data. LC/MS/MS was performed on available archival specimens when such data was not available clinically. Results: Patients with newly diagnosed amyloidosis who had sufficient mass spectrometry data on fat aspirate or tissue biopsy for IGVL gene usage identification were included (IgM AL, N=44 and Non-IgM AL, N=391). Clinical Features: As expected, differences were noted in baseline characteristics of patients with IgM vs. non-IgM AL amyloidosis. Patients with IgM amyloidosis were older (68 vs. 64 years, p=0.04) and more likely to have kappa as the involved light chain (34% vs. 25%, p=0.2), though the difference did not reach statistical significance. IgM amyloidosis patients had lower difference in involved and uninvolved free light chains (dFLC) (15.5 vs. 25.8 mg/dL, p=0.01) and lower NTProBNP levels (median: 1987 vs. 2655 pg/mL, p=0.02) compared with non-IgM amyloid patients. Presence of t(11;14) was less common by FISH (30% vs. 48%, p=0.1), though the difference did not reach statistical significance. Patterns of organ involvement were different, with heart involvement being less frequent in IgM patients (64% vs. 76%), while peripheral nerve (30% vs. 18%, p=0.05), soft tissue (41% vs. 21%, p=0.005) and lung involvement (7% vs. 3%, p=0.1) were more common in patients with IgM vs. non-IgM AL amyloidosis. There was no difference in liver (9% vs. 15%), kidney (50% vs. 51%) GI (23% vs. 25%) or autonomic nervous system involvement (18% vs. 14%). IGVL Gene Usage: As shown in Figure 1, IGVL usage differed across the two groups. In the lambda group, LV2-08 (14% vs. 2%, p<0.001) and LV2-14 (36% vs. 10%, p<0.001) were more common in IgM amyloid patients, while LV1-44 (0% vs. 10%, p=0.02) and LV6-57 (2% vs. 18%, p=0.004) were less common in the IgM group. There was also a trend towards less frequent involvement of LV3-01 (2% vs. 12%, p=0.07) in IgM amyloid patients. In the kappa group, KV4-01 (11% vs. 4%, p=0.04) was more common in patients with IgM amyloidosis. Correlation Between IGVL Gene usage and Clinical Features: IGVL gene usage correlated with disease characteristics and differences in the clinical presentation of patients across the two groups. LV2-14 was more commonly seen in IgM amyloid patients (above). It has been previously associated with a higher frequency of peripheral nerve involvement (Kourelis et al. Blood 2017), which was more frequent in IgM group. LV2-14 is also associated with low dFLC levels, which were noted to be lower in the IgM AL cohort. LV1-44 has been associated with higher likelihood of cardiac involvement. LV1-44 was seen less frequently in the IgM amyloid patients, who also had lower rates of cardiac involvement. LV6-57 has been historically associated with higher rates of t(11;14). It was less common in IgM AL patients, who also had lower rates of t(11;14). Conclusions: IGVL gene usage differed significantly in patients with IgM vs. non-IgM amyloidosis, correlating with differences in disease characteristics and organ involvement. This is the first study to our knowledge to report IGVL gene usage differences in IgM vs. non-IgM AL amyloidosis. Differences in IGVL gene usage may explain the distinct clinical presentation seen in IgM amyloidosis, the biology of which has so far remained poorly understood. *SS and SD contributed equally Disclosures Dasari: The Binding Site: Patents & Royalties: US Patent Rights on Mass Spectroscopy Licensing agreement with The Binding Site, Research Funding. Dispenzieri:Alnylam: Research Funding; Celgene: Research Funding; Takeda: Research Funding; Pfizer: Research Funding; Janssen: Consultancy; Akcea: Consultancy; Intellia: Consultancy. Murray:The Binding Site: Patents & Royalties: US Patent Rights on Mass Spectroscopy Licensing agreement with The Binding Site, Research Funding. Kumar:Janssen: Consultancy, Research Funding; Celgene: Consultancy, Research Funding; Takeda: Research Funding. Gertz:Prothena: Honoraria; Alnylam: Honoraria; Ionis: Honoraria; Janssen: Honoraria; Spectrum: Honoraria, Research Funding; Celgene: Honoraria.

Introduction: Primary cold agglutinin disease (CAD) is a rare disorder and accounts for 15% of all autoimmune hemolytic anemia. It is associated with an underlying monoclonal protein, which is IgM kappa in the majority of patients. The IgM protein triggers complement mediated extravascular hemolysis. The underlying pathophysiology by which autoimmune hemolysis affects only a small subset of patients with IgM monoclonal protein is poorly understood. Changes in the IgM structure via post-translational modifications of light chains (LCs), such as glycosylation are enriched in patients with AL amyloidosis, another rare disorder where disease manifestations are driven by the protein structure. (Kumar S et al Leukemia 2017). We hypothesized that presence of post-translational glycosylation is a potential mechanism allowing the IgM protein to trigger complement mediated hemolysis in CAD. Methods: Serum samples of patients being evaluated for monoclonal protein disorders were analyzed by immuno-enrichment-based matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS), termed MASS-FIX. Patients who had an underlying IgM monoclonal protein identified by MASS-FIX from 07/23/2018 to 5/14/2019 were included this study. Glycosylated LCs were identified by the shift in their molecular mass due to the added molecular mass of the carbohydrates. Glycosylation was compared in patients with IgM associated CAD vs. other IgM disorders. Results: 438 unique patients with an IgM monoclonal protein identified on MASS-FIX were included in the study. CAD was present in 3% (n=14) of patients. The primary underlying diagnosis in the remaining patients was monoclonal gammopathy of undetermined significance (MGUS): 47% (n=206), lymphoplasmacytic lymphoma (LPL)/Waldenstrom Macroglobulinemia (WM): 30% (n=132), non-LPL lymphoma: 6% (n=28), AL amyloidosis: 4% (n=19), CLL: 4% (n=18), cryoglobulinemia: 2% (n=10), multiple myeloma: 2% (n=9) and other: 0.5% (n=2) Glycosylation was present in 7% (31/438) of patients in this cohort. Glycosylation was significantly more prevalent in patients with CAD than other IgM gammopathies (64% vs. 5%, p &lt;0.001). CAD was present in 29% of patients with glycosylation and in only 1% of patients without glycosylation, p&lt;0.001. Disease characteristics of patients with CAD, including patients with and without glycosylated LCs are listed in Table 1. No statistically significant differences were observed in the features of CAD patients with and without glycosylation, though sample size was small. All patients (100%) had IgM kappa monoclonal protein. Median M-spike was 0.5 g/dL. The associated disorder was IgM MGUS in 50% of patients, LPL/WM in 29%, marginal zone lymphoma in 14% and a non-specific B-cell lymphoproliferative disorder in 7% of patients. Median hemoglobin was 11.2 g/dL. All patients had characteristic RBC agglutination on peripheral smear. Elevated LDH and undetectable haptoglobin were seen in 100% of patients. Cold agglutinin titer was positive (&gt;1:64) in 12 of 13 patients (85%) in whom it was tested. The remaining patient had a titer of 1:8. Polyspecific Coombs test/direct antiglobin test (DAT) was positive in 100% of patients (n=12 tested) and all patients who were tested with monospecific DAT had an antiC3 antibody (n=10). One patient also had monospecific anti IgG antibody. Transfusions were needed in 21% of patients. Other clinical manifestations included acrocyanosis in 43% (n=6) and Raynaud's phenomenon in 50% (n=7) of patients, respectively. Treatment for CAD included rituximab with or without steroids in 50% (n=7) of patients, rituximab-based chemotherapy in 29% (n=4) of patients, with observation alone in 21% (n=3) of patients. Conclusions: Modification of the IgM immunoglobulin structure by post-translational glycosylation of LCs was significantly more common in patients with CAD compared with other IgM monoclonal gammopathies (64% vs. 5%, p &lt;0.001). CAD was seen in only 1% of patients without glycosylation, while one-third of patients with glycosylation had CAD. This is a novel finding. As MASS FIX is now routinely done for detection of monoclonal proteins at our institution, the presence of LC glycosylation can provide diagnostic clues and can be a potential therapeutic target in the future. Future studies in larger, independent cohorts of patients are needed to confirm these findings. *SS & DLM contributed equally Disclosures Murray: The Binding Site: Patents & Royalties: US Patent Rights on Mass Spectroscopy Licensing agreement with The Binding Site, Research Funding. Dasari:The Binding Site: Patents & Royalties: US Patent Rights on Mass Spectroscopy Licensing agreement with The Binding Site, Research Funding. Kumar:Celgene: Consultancy, Research Funding; Janssen: Consultancy, Research Funding; Takeda: Research Funding. Dispenzieri:Janssen: Consultancy; Pfizer: Research Funding; Takeda: Research Funding; Celgene: Research Funding; Alnylam: Research Funding; Intellia: Consultancy; Akcea: Consultancy. Gertz:Spectrum: Honoraria, Research Funding; Janssen: Honoraria; Celgene: Honoraria; Prothena: Honoraria; Ionis: Honoraria; Alnylam: Honoraria.