In localized light chain amyloidosis (locAL), amyloidogenic light chains (aLC) are produced and deposited locally by a B‐cell clone. We present 293 patients with immunohistochemically confirmed ...locAL. Lung (nodular pulmonary) with 63 patients was the most involved organ. The aLC was λ in 217 cases (κ:λ ratio 1:3). A local B‐cell clone was identified in 30% of cases. Sixty‐one (21%) had a concomitant autoimmune disorder (cAD). A monoclonal component (MC) were present in 101 (34%) patients and were more frequent in subjects with cAD (51% vs 34%; P = .03). Cigarette smoking was more prevalent in lung locAL (54% vs 37%; P = .018). After a median follow‐up of 44 months, 16 patients died and 5‐ and 10‐years locAL progression‐free survival (PFS) were 62% and 44%. Interestingly, locAL‐PFS was shorter among patients with an identified clonal infiltrate at amyloid deposition site (40 vs 109 months; P = .02) and multinuclear giant cells and/or an inflammatory infiltrate resulted in longer locAL‐PFS in lung involvement (65 vs 42 months; P = .01). However, no differences in locAL PFS were observed in patients with cAD, a MC and involved organ site. Treatment was administered in 163 (54%) patients and was surgical in 135 (46%). Median locAL‐PFS after first treatment was 56 months. Responders had longer locAL‐PFS (78 vs 17 months; P < .001). Three patients with lung locAL and a MC were diagnosed as systemic AL amyloidosis at follow‐up. In summary, locAL pathogenesis seems to be heterogeneous and the clonal infiltrate leads local progression.
Systemic light chain amyloidosis is a rare and life-threatening disorder, for which accurate risk stratification is crucial. Current cardiac staging systems (MAYO2004, MAYO3b, and MAYO2012) are ...mainly based on biomarkers, which have uncertain reliability in the context of atrial fibrillation, arrhythmia or pacemaker stimulation as well as renal insufficiency. We compared the performance of the established staging systems with particular regard to these comorbidities in 1,224 patients with systemic light chain amyloidosis diagnosed at our center from July 2002 until March 2017. We first characterized the subsets with an estimated glomerular filtration rate <50 mL/min/1.73 m
(415 patients) and any kind of atrial arrhythmia (183 patients) as unique high-risk subgroups with similarly increased cardiac biomarkers (χ
-test, all
<0.001). This resulted in a shift towards higher risk stages and reduced median overall survival compared to those of patients with better kidney function or without atrial arrhythmia in univariate analyses (13
46 months and 17
53 months, respectively; both
<0.001). Performance analysis revealed that predictions in the entire cohort were least precise with the MAYO2004 staging system and most precise with the MAYO3b system. This performance pattern was almost preserved for patients with an estimated glomerular filtration rate <50 mL/min/1.73 m
, but less so for those with atrial arrhythmias. The MAYO3b staging system was most robust. Importantly, atrial arrhythmia retained its prognostic value in multivariable analysis including age, difference between involved and uninvolved free light chains, and any staging system, while estimated glomerular filtration rate <50 mL/min/1.73 m
was not statistically significant in multivariable analysis with the MAYO3b staging system. In conclusion, our results favor the MAYO3b staging system due to its consistently best performance and retained applicability in the subgroups with atrial arrhythmia and estimated glomerular filtration rate <50 mL/min/1.73 m
.
Monoclonal antibodies against the cell surface antigen CD38, e.g., isatuximab, daratumumab, or Mor202, have entered the therapeutic armamentarium in multiple myeloma due to single agent overall ...response rates of 29 vs. 36 vs. 31%, effectivity in combination regimen, e.g., with lenalidomide or bortezomib plus dexamethasone, and tolerable side effects. Despite clinical use, many questions remain. In this manuscript, we address three of these: first, upfront CD38 target-expression in AL-amyloidosis, monoclonal gammopathy of unknown significance (MGUS), asymptomatic, symptomatic, and relapsed multiple myeloma. Second, relation of CD38-expression to survival, disease stages, molecular entities, and high-risk definitions. Third, alternative splicing or lack of CD38-expression as potential mechanisms of upfront resistance. We assessed CD138-purified plasma cell samples from 196 AL-amyloidosis, 62 MGUS, 259 asymptomatic, 764 symptomatic, and 90 relapsed myeloma patients, including longitudinal pairs of asymptomatic/symptomatic (
= 34) and symptomatic/relapsed myeloma (
= 57) regarding interphase fluorescence
hybridization (
= 1,380), CD38-expression by gene expression profiling (
= 1,371), RNA-sequencing (
= 593), and flow cytometry (
= 800). Samples of normal bone marrow plasma cells (
= 10), memory B-cells (
= 9), polyclonal plasmablastic cells (
= 9), and human myeloma cell lines (
= 54) were used as comparators. CD38 was expressed in all malignant plasma cell samples, but significantly lower compared to normal plasma cells with small but significant downregulation in longitudinal sample pairs. Higher CD38 expression was associated with the presence of t(4;14) and high-risk according to the UAMS70-gene score, lower expression was associated with del17p13 and hyperdiploidy in symptomatic myeloma as well as t(11;14) in asymptomatic myeloma. Higher CD38-expression was associated with slower progression to symptomatic and relapsed myeloma and better overall survival in the latter two entities. CD38 expression, t(4;14), del17p13, and gain of 1q21 are independently prognostic in multivariate analysis. By contrast, high CD38-expression is associated with adverse survival in AL-amyloidosis. Regarding mechanisms of upfront anti-CD38-treatment resistance, lack of CD38-expression and alternative splicing of receptor binding-sites could be excluded. Here, of the two protein coding CD38-transcripts CD38-001 (eight-exon, full length) and CD38-005 (truncated), CD38-001 conveyed >97% of reads spanning the respective CD38 splice junction.
Electron microscopy is the gold standard to characterize cellular ultrastructure. However, production of significant morphometrical data is highly limited by acquisition time. Here, we describe a ...semi-automated high-throughput strategy using single-axis serial section electron tomography to investigate and analyze centriole ultrastructure in bone-marrow-derived, primary human CD138pos plasma cells. The protocol comprises steps for electron microscopy sample preparation, semi-automated transmission electron microscopy screening, and screening evaluation for cells of interest. Thereafter, we detail tomography acquisition, data reconstruction, and joining.
For complete details on the use and execution of this protocol, please refer to Dittrich et al.1
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•High-throughput production of volumetric EM data•From sample acquisition to tomography data in one week•Protocol can be used in both cell lines and primary patient material•Protocol is adaptable to various organelles and subcellular features
Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.
Electron microscopy is the gold standard to characterize cellular ultrastructure. However, production of significant morphometrical data is highly limited by acquisition time. Here, we describe a semi-automated high-throughput strategy using single-axis serial section electron tomography to investigate and analyze centriole ultrastructure in bone-marrow-derived, primary human CD138pos plasma cells. The protocol comprises steps for EM sample preparation, semi-automated transmission electron microscopy screening, and screening evaluation for cells of interest. Thereafter, we detail tomography acquisition, data reconstruction, and joining.
Structured patient data play a key role in all types of clinical research. They are often collected in study databases for research purposes. In order to describe characteristics of a next-generation ...study database and assess the feasibility of its implementation a proof-of-concept study in a German university hospital was performed. Key characteristics identified include FAIR access to electronic case report forms (eCRF), regulatory compliant Electronic Data Capture (EDC), an EDC with electronic health record (EHR) integration, scalable EDC for medical documentation, patient generated data, and clinical decision support. In a local case study, we then successfully implemented a next-generation study database for 19 EDC systems (n = 2217 patients) that linked to i.s.h.med (Oracle Cerner) with the local EDC system called OpenEDC. Desiderata of next-generation study databases for patient data were identified from ongoing local clinical study projects in 11 clinical departments at Heidelberg University Hospital, Germany, a major tertiary referral hospital. We compiled and analyzed feature and functionality requests submitted to the OpenEDC team between May 2021 and July 2023. Next-generation study databases are technically and clinically feasible. Further research is needed to evaluate if our approach is feasible in a multi-center setting as well.
Daratumumab has shown promising first results in systemic amyloid light-chain (AL) amyloidosis. We analyzed a consecutive series of 168 patients with advanced AL receiving either ...daratumumab/dexamethasone (DD, n = 106) or daratumumab/bortezomib/dexamethasone (DVD, n = 62). DD achieved a remission rate (RR) of 64% and a very good hematologic remission (VGHR) rate of 48% after 3 months. Median hematologic event-free survival (hemEFS) was 11.8 months and median overall survival (OS) was 25.6 months. DVD achieved a 66% RR and a 55% VGHR rate. Median hemEFS was 19.1 months and median OS had not been reached. Cardiac organ responses were noted in 22% with DD and 26% with DVD after 6 months. Infectious complications were common (Common Terminology Criteria CTC grade 3/4: DD 16%, DVD 18%) and likely related to a high rate of lymphocytopenia (CTC grade 3/4: DD 20%, DVD 17%). On univariable analysis, hyperdiploidy and gain 1q21 conferred an adverse factor for OS and hemEFS with DD, whereas translocation t(11;14) was associated with a better hemEFS. N-terminal prohormone of brain natriuretic peptide >8500 ng/L could not be overcome for survival with each regimen. Multivariable Cox regression analysis revealed plasma cell dyscrasia (difference between serum free light chains dFLC) >180 mg/L as an overall strong negative prognostic factor. Additionally, nephrotic-range albuminuria with an albumin-to-creatinine-ratio (ACR) >220 mg/mmol was a significantly adverse factor for hemEFS (hazard ratio, 2.1 and 3.1) with DD and DVD. Daratumumab salvage therapy produced good results and remission rates challenging any therapy in advanced AL. Outcome is adversely influenced by the activity of the underlying plasma cell dyscrasia (dFLC) and nephrotic-range albuminuria (ACR).
•Daratumumab can achieve good results in advanced AL amyloidosis, but is less effective in nephrotic patients with high free light chains.•Translocation t(11;14) is beneficial, and myeloma-like hyperdiploidy is adverse for outcome with daratumumab therapy in AL.
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The difference between involved minus uninvolved serum free light chains (dFLC) has been established as an invaluable hematologic parameter in systemic amyloid light chain (AL) amyloidosis. However, ...patients with an initial dFLC level <50 mg/L are currently deemed not evaluable for response to therapy. Therefore, we aimed to characterize this subgroup of patients and to define novel hematologic response parameters. We retrospectively analyzed 783 AL patients newly diagnosed at our center between 2002 and 2016. Patients with a dFLC level <50 mg/L showed smaller bone marrow plasmacytosis compared to patients with a dFLC level ≥50 mg/L (7% vs 10%, P < .001), but no significant differences in all analyzed chromosomal aberrations. Cardiac involvement was less frequent (45% vs 80%, P < .001) and less severe (Mayo 2004 stage III: 18% vs 51%, P < .001), whereas kidney involvement was more prevalent (83% vs 53%, P < .001) and proteinuria was higher (7.3 g/L vs 5.0 g/L, P < .001). In multivariate analyses, a dFLC level <50 mg/L appeared to be an independent prognostic factor with respect to overall survival (hazard ratio HR = 0.50, P = .003) and renal survival (HR = 0.56, P = .020). Patients with a dFLC level <50 mg/L showed a higher proportion of complete hematologic response after first-line therapy compared to patients with a dFLC level ≥50 mg/L (39% vs 9%, P < .001). Receiver-operating characteristics analysis identified a low-dFLC partial response (dFLC <10 mg/L for patients with a dFLC between 20 and 50 mg/L), which predicted overall and renal survival already at 3 months after the start of therapy. Importantly, a parallel Italian study validated this new hematologic remission parameter. The outcome of prospective clinical trials might be adversely influenced by exclusion of the favorable clinical subgroup with an initial dFLC <50 mg/L. We propose the appreciation of dFLC in hematologic response assessment for all patients with a baseline dFLC >20 mg/L.
•AL patients with an initial dFLC <50 mg/L represent a distinct clinical subgroup with mostly renal disease and a favorable prognosis.•These patients are evaluable for hematologic response including a novel low-dFLC partial response, which predict renal and overall survival.
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