BackgroundThe monoclonal antibody daratumumab, approved for treating myeloma, targets CD38, a protein on myeloma and also on CD34+ hematopoietic progenitor cells. Because mobilized CD34+ cells are critical for stem cell transplant, we investigated the in vitro activity of daratumumab on mobilized CD34+ cells from myeloma patients with no prior exposure to daratumumab.MethodsWe determined the number of CD38 molecules per CD34+ cell, and whether daratumumab bound to CD34+ cells, whether C1q bound to daratumumab-coated CD34+ cells and whether daratumumab-related complement-dependent cytotoxicity (CDC) occurred. We also examined CD34+ cell progenitor cell colony capacity in assays with pre-plating incubation of CD34+ cells with daratumumab alone or with daratumumab and the CD59 inhibitory antibody BRIC229, and also assessed CD34+ cell responses to increasing doses of daratumumab in caspase 3/7 activity assays.ResultsAlthough 75% of mobilized CD34+ cells co-express CD38, CD38 was minimally present on CD34+ cells compared to Daudi and KG-1 controls, C1q did not bind to daratumumab-coated CD34+ cells, and CDC did not occur. CD34+ cells incubated in complement-rich human serum with daratumumab alone or with daratumumab and BRIC229, and then plated in progenitor cell assays, produced similar numbers of colonies as controls. In progenitor cell assays with cryopreserved or fresh unselected or CD34-selected cells, daratumumab did not affect progenitor cell capacity, and in caspase 3/7 activity assays CD34+ cells were not affected by increasing doses of daratumumab.ConclusionIn vitro, daratumumab is not toxic to mobilized CD34+ progenitor cells from myeloma patients.
Introduction: Response criteria in light-chain (AL) amyloidosis, revised in 2012, did not specify a depth of change in the pathologic involved FLC (iFLC) for either an amyloid complete response (aCR) or a very good partial response (VGPR) (JCO 2012;30:4541). The FLC ratio upon which an aCR is based is impacted if renal function deteriorates and if either the iFLC or uninvolved FLC (uFLC) become undetectable; moreover, the VGPR was based on the difference between iFLC and uFLC and not on the response in the iFLC. Emerging data have controverted the role of the FLC ratio, suggesting the need for a revised criteria, particularly since a new CD38-based treatment regimen may achieve iFLC levels <10mg/L in over 70% of patients and may often make the ratio incalculable (Abs #S875 24th European Hematology Association Congress;6/15/19). In a revisionist mode, we examined overall survival (OS) from diagnosis in patients achieving > VGPR as a function of baseline, treatment-related and iFLC response variables. Patients and Methods: Patients with AL diagnosed between 2005-2017 with > VGPR after treatment were included in this IRB-approved retrospective study. Cox proportional-hazards regression analyses were used to assess the impact of baseline variables (gender, age at diagnosis, pathologic light-chain isotype, marrow plasma cells, iFLC, dFLC, and cardiac and renal stage), and also to assess the impact of treatment and iFLC-response related variables (types of initial therapy, number of courses of therapy [induction if needed+MEL SCT+consolidation = 1 course], exposure to daratumumab, achievement of aCR and VGPR, and iFLC response). iFLC responses were defined as levels <10, 10-20 or > 20mg/L. MedCalc 19.0.3 statistical software was used for all statistical analyses. Results: One hundred and thirty-three patients met the criteria and were 78M/55W a median of 60.5 yo (range, 35-81) with AL-lambda type in 114 (86%) and marrows with a median 10% plasma cells (1-50). Median iFLC and dFLC were 135mg/L (29.4-9780) and 123mg/L (4-9770) respectively, and 89 patients (66%) had cardiac (stage II=49, III=33) and 87 (65%) renal involvement (stage II=55, III=16). Of baseline variables, only age and cardiac stage were significant predictors of OS. Ninety one (68%) had bortezomib-based initial therapy and 62 (47%) went to SCT+consolidation. Eighty-three (63%) received 2 courses of therapy and 14 (11%) received 3 courses; 7 received daratumumab in second-line therapy and 14 in the third-line. Eighty four (63%) achieved an aCR. With a median follow up > 5 years, of treatment and response related variables, only the iFLC response predicted OS (p<0.01). Log-rank (Kaplan-Meier) analysis showed that patients achieving iFLC <10mg/L had over 95% survival at 120 months, compared to those achieving iFLC 10-20mg/L or >20mg/L whose median OS were 96 and 121 months respectively (p<0.01) (Figure 1). We then compared patients in the iFLC groups. They did not differ by Mann-Whitney in age at diagnosis or by χ2 with respect to cardiac stage, types of initial therapy, number of courses of therapy, or exposure to daratumumab. However, the groups differed significantly by Mann-Whitney in baseline iFLC/dFLC [medians, 75/62 (<10), 212/192 (10-20), 227/190 (>20) (all comparisons, p<0.01)], and by χ2 in aCR rates [81% (<10), 67% (10-20) and 47% (>20) (p<0.01)]. Conclusions: Multiple variables impact OS in patients achieving > VGPR to therapy, including age, cardiac stage at diagnosis, and iFLC response. In this series, the optimal hematologic response is an iFLC <10mg/L and the major challenge to achieving that goal is the scale of FLC disease at diagnosis. Further studies are needed to validate a revision of the hematologic response criteria in AL in the modern era of monoclonal antibody therapy. Disclosures Comenzo: Unum: Membership on an entity's Board of Directors or advisory committees, Research Funding; Caelum: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Myself: Patents & Royalties: Patent 9593332, Pending 20170008966; Karyopharm: Research Funding; Prothena Biosciences: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Sanofi-Aventis: Membership on an entity's Board of Directors or advisory committees; Takeda: Research Funding.
Introduction: Systemic light-chain (AL) amyloidosis results from clonal plasma cells that secrete toxic fibril-forming free light chains. Therapies directed at the plasma cell clone form the backbone of its management. Identification of cell-surface receptors on the clonal cells can provide targets for therapy. BCMA is one such cell-surface glycoprotein; it is principally expressed on plasma cells and supports their long-term survival (J Exp Med. 2004;199:91-98). Anti-BCMA immunotherapies are currently being studied in multiple myeloma (N Engl J Med. 2019;380:1726-1737). Membrane-bound BCMA (mBCMA) is also shed as a soluble form, sBCMA, due to γ-secretase activity that can be inhibited by a small molecule (GSI, LY-411575) (Nat Commun. 2015;6:7333; J Immunol. 2017;198(8):3081-3088). We report on mBCMA on the clonal plasma cells of AL patients and its modulation by GSI in vitro, and on sBCMA in the blood of AL patients and of mice xenografted with an AL cell line, demonstrating its correlations in vivo with free light chain (FLC) levels and plasma cell tumor burden. Methods: We analyzed mBCMA and sBCMA levels in marrow aspirate and peripheral blood samples from AL patients under an IRB approved protocol. We isolated mononuclear cells (MNC) from patient marrow aspirates with anti-CD138 microbeads (Miltenyi Biotec, Auburn, CA), and used the CD138-selected cells in culture with LY-411575 (Sigma Aldrich, St Louis, MO). We analyzed mBCMA expression by flow cytometry using APC conjugated anti-CD269 (BCMA) antibody (Biolegend, San Diego, CA, USA) and CD138 expression by PE-conjugated anti-CD138 antibody (Biolegend, San Diego, CA, USA), along with appropriate isotype controls. We injected 107 ALMC-1 reporter cells in the flanks of NOD scid gamma (NSG) mice to create a xenograft model of AL clonal plasma cell disease (Jackson Laboratories, Bar Harbor, ME). sBCMA in patients and mice and FLC in mice were measured by ELISA (R&D Systems, Minneapolis, MN; Bethyl lab Montgomery, TX respectively). Pearson and Spearman correlation analysis was used to examine associations of sBCMA and clinical disease parameters. Paired t-test was applied to compare BCMA expression before and after treatment with GSI. Results: Marrow and blood were obtained from 20 AL patients, 8 newly diagnosed, 4 with progression of disease, and 8 after treatment with >VGPR. Their median age was 65 years (range, 48-77) and 50% were female. Median plasma cells in the marrow aspirates and involved FLC levels were 5% (1-20%) and 33 mg/L (6.6-2220mg/L) respectively. Median mBCMA expression on CD138+ marrow MNC and sBCMA levels in plasma were 39% (4-83) and 28.5 ng/ml (6.6-100.3) respectively (Figure 1A-B). sBCMA levels correlated with bone marrow plasma cell percentage and iFLC (both p<0.001, Figure 1C-D). In culture with LY-411575, the percentages of CD138 cells positive for mBCMA increased from 85% to 100% with ALMC-1 cells and from 36% to 68% (p < 0.01) with patient CD138-selected cells while the sBCMA levels in culture supernatant decreased by over 50%. In NSG mice with ALMC-1 reporter cell xenografts, medians of luciferin-based bioluminescence FLUX (photons/s), λ FLC and sBCMA were 3.9x1010 (2.02x109-1.2x1011), 949.1 mg/L (868.8-23629.2), and 3.8 ng/ml (0.9-23.6) respectively. sBCMA levels correlated with FLC (Pearson r= 0.99, p<0.0001) and with FLUX (Pearson r=0.61, p=0.07). Conclusions: BCMA is expressed on AL plasma cells and sBCMA is detected in the blood of all AL patients. In this light chain disease, sBCMA may be useful as a marker of disease activity even in patients with low FLC. Furthermore, expression of mBCMA can be manipulated by treatment with a GSI, an approach which may be useful therapeutically in AL. These results provide the basis for applying anti-BCMA immunotherapies in clinical trials in relapsed refractory AL patients. Disclosures Comenzo: Sanofi-Aventis: Membership on an entity's Board of Directors or advisory committees; Unum: Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Research Funding; Caelum: Consultancy, Membership on an entity's Board of Directors or advisory committees; Janssen: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Karyopharm: Research Funding; Prothena Biosciences: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Myself: Patents & Royalties: Patent 9593332, Pending 20170008966.
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