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• In those with cardiac amyloidosis and heart failure-related cardiogenic shock, there was significant augmentation of cardiac index after intra-aortic balloon pump (IABP) and significant decreases in left ventricular and right ventricular filling pressures. • Predictors of lack of response to IABP by cardiac index were a smaller baseline left ventricular end-diastolic diameter size per centimeter and higher pre-IABP systemic vascular resistance. • Most patients with Society for Cardiovascular Angiography and Interventions stage C heart failure-related cardiogenic shock were stabilized successfully and bridged to advanced therapies with IABP alone. In those with heart failure-related cardiogenic shock (HF-CS), an intra-aortic balloon pump (IABP) may improve hemodynamics and be useful as a bridge to advanced therapies. We explore whether those with cardiac amyloidosis and HF-CS might experience hemodynamic improvement and describe the hemodynamic response after IABP. We retrospectively identified consecutive patients with a diagnosis of cardiac amyloid, either light chain or transthyretin, who were admitted to our intensive care unit with HF-CS. Patients were excluded if an IABP was placed during heart transplant or for shock related to acute myocardial infarction. Invasive hemodynamics before and after IABP placement were assessed. We identified 23 patients with cardiac amyloid who had an IABP placed for HF-CS. The 1-year survival rate was 74% and most (65%) were bridged to heart transplant, although 1 patient was bridged to destination left ventricular assist device. After IABP, the mean arterial pressure, cardiac index, and cardiac power index were significantly increased, whereas mean right atrial pressure, mean pulmonary artery pressure, and pulmonary capillary wedge pressure were all significantly decreased. A smaller left ventricular end-diastolic diameter (per cm) was associated with a higher likelihood of a cardiac index of <2.2 L/min/m2 after IABP (odds ratio 0.16, 95% confidence interval 0.01–0.93, P =.04). IABP significantly improved cardiac index while decreasing right atrial pressure, mean pulmonary artery pressure, and pulmonary capillary wedge pressure in cardiac amyloidosis patients with HF-CS. [Display omitted] [ABSTRACT FROM AUTHOR]

Summary Patients with AL amyloidosis can have persistent organ dysfunction despite achieving a haematological complete response (hemCR). We aimed to identify factors for organ non‐response among 143 patients who achieved hemCR for ≥6 months. Kidney, heart and liver non‐response were observed in 40/117 (34%), 19/68 (28%) and 3/17 (18%) patients respectively. Predisposing factors varied by organ system. Kidney non‐responders had more advanced organ dysfunction at diagnosis, whereas heart non‐responders had disproportionately more lambda‐typic amyloidogenic light chains. Most patients without an apparent reason for organ non‐response had detectable residual clonal disease. The interplay of factors impeding organ recovery in AL amyloidosis is complex. [ABSTRACT FROM AUTHOR]

Purpose of Review: To discuss the recent advances in the diagnosis and treatment of light chain cardiac amyloidosis (AL-CM). Recent Findings: The improved recognition of AL-CM is primarily driven by improved diagnostic techniques. Cardiac MRI has emerged as a sensitive modality for the diagnosis of AL-CM. Positron emission tomography (PET) with a whole host of amyloid fibril-binding tracers has tremendous promise. On the therapeutics front, better anti-plasma cells therapies are in clinical use or under investigation, and a new front of fibril depleters are opening the doors for targeting organ recovery by promoting the resorption of amyloid fibrils. Summary: AL-CM results in significant morbidity and mortality mainly due to delayed diagnosis and lack of therapies that acutely address the underlying organ dysfunction. The combination of improved disease detection with improved therapy promises to continue transforming AL-CM into either chronic treatable or curable disease. [ABSTRACT FROM AUTHOR]

Purpose of Review: Cardiac amyloidosis (CA) is a condition characterized by misfolding and extracellular deposition of proteins, leading to organ dysfunction. While numerous forms of CA exist, two subtypes dominate clinical prevalence: Transthyretin amyloid (ATTR) and immunoglobulin light chain amyloid. Recent Findings: The current scientific landscape reflects the urgency to advance therapeutic interventions with over 100 ongoing clinical trials. Heart failure treatment is affected by CA phenotype with poor tolerance of otherwise frequently used medications. Treating comorbidities including atrial fibrillation and valvular disease remains a challenge in CA, driven by technical difficulties and uncertain outcomes. Tafamidis is the first ATTR-stabilizer approved with a rapidly growing rate of clinical use. In parallel, various new therapeutic classes are in late-stage clinical trials including silencers, antibodies and genetic therapy. Summary: Managing CA is a critical challenge for future heart failure care. This review delineates the current standard-of-care and scientific landscape of CA therapy. [ABSTRACT FROM AUTHOR]

Amyloid is an amorphous, fibrillar material formed from various abnormally folded proteins that deposits locally or systemically. Over 95% of cases have been attributed to light chain deposition (AL) or transthyretin deposition (ATTR) amyloidosis. The basic investigations in the evaluation of cardiac amyloidosis include the electrocardiogram, echocardiography and cardiac biomarkers. Echocardiography in a patient with cardiac amyloidosis shows biatrial enlargement, biventricular hypertrophy, diastolic dysfunction, interatrial septal thickening, valvular thickening, a glistening appearance of the interventricular septum, and pericardial effusion. Magnetic resonance imaging can help distinguish amyloidosis from other causes of infiltrative/restrictive cardiomyopathy, from example, sarcoidosis, hemochromatosis, and Fabry disease based on characteristic enhancement patterns in these diseases. The latest Expert Consensus recommends that serum/urine immunofixation electrophoresis along with a serum free light chain assay must be done in all the cases of suspected cardiac amyloidosis. If the light chain assays are positive, we proceed with tissue diagnosis for confirmation of AL amyloidosis. If the screening assays are negative for monoclonal gammopathy, the next step is to obtain cardiac scintigraphy. If the nuclear scan is negative, but the index of suspicion remains high, an endomyocardial biopsy can be done. Once amyloid is demonstrated in histopathologic specimens, it must be typed to distinguish between AL and ATTR. The ideal method for this is tandem mass spectrometry, although this may not be widely available. It has a sensitivity of 88% and specificity of 96% higher than other techniques 23. In resource-poor settings, immunohistochemistry or immunoelectron microscopy can allow this distinction, although with lesser sensitivity.

In contrast to natural antibodies that rely mainly on the heavy chain to establish contacts with their cognate antigen, we have developed a bispecific antibody format in which the light chain (LC) drives antigen binding and specificity. To better understand epitope-paratope interactions in this context, we determined the X-ray crystallographic structures of an antigen binding fragment (Fab) in complex with human CD47 and another Fab in complex with human PD-L1. These Fabs contain a κ-LC and a λ-LC, respectively, which are paired with an identical heavy chain (HC). The structural analysis of these complexes revealed the dominant contribution of the LCs to antigen binding, but also that the common HC provides some contacts in both CD47 and PD-L1 Fab complexes. The anti-CD47 Fab was affinity optimized by diversifying complementary-determining regions of the LC followed by phage display selections. Using homology modeling, the contributions of the amino acid modification to the affinity increase were analyzed. Our results demonstrate that, despite a less prominent role in natural antibodies, the LC can mediate high affinity binding to different antigens and neutralize their biological function. Importantly, Fabs containing a common variable heavy (VH) domain enable the generation of bispecific antibodies retaining a truly native structure, maximizing their therapeutic potential.

Simple Summary: Light-chain (AL) amyloidosis is a rare disease similar to the more common disease, multiple myeloma (MM). Both are caused by proliferation of malignant plasma cells. In AL amyloidosis, disease is a result of the deposition of aggregates of proteins, namely immunoglobulin light chains, secreted by the malignant plasma cells, in target organs such as the heart or kidneys. Historically, treatment of AL amyloidosis has followed that of MM. A wide range of novel immunotherapies, i.e., therapies which utilize or activate immune mechanisms to eliminate the disease, are already established in MM and are gradually being adopted in AL amyloidosis as well. Although promising, the increased frailty of typical AL amyloidosis compared to MM patients is a concern in the administration of these therapies, which may be associated with severe side effects. We review both the promise and the challenges with the expansion of MM immunotherapies to AL amyloidosis. Light-chain (AL) amyloidosis is a rare plasma cell disorder characterized by the deposition of misfolded immunoglobulin light chains in target organs, leading to multi-organ dysfunction. Treatment approaches have historically mirrored but lagged behind those of multiple myeloma (MM). Recent advancements in MM immunotherapy are gradually being evaluated and adopted in AL amyloidosis. This review explores the current state of immunotherapeutic strategies in AL amyloidosis, including monoclonal antibodies, antibody–drug conjugates, bispecific antibodies, and chimeric antigen receptor T-cell therapy. We discuss the unique challenges and prospects of these therapies in AL amyloidosis, including the exposure of frail AL amyloidosis patients to immune-mediated toxicities such as cytokine release syndrome (CRS) and immune effector-cell-associated neurotoxicity syndrome (ICANS), as well as their efficacy in promoting rapid and deep hematologic responses. Furthermore, we highlight the need for international initiatives and compassionate programs to provide access to these promising therapies and address critical unmet needs in AL amyloidosis management. Finally, we discuss future directions, including optimizing treatment sequencing and mitigating toxicities, to improve outcomes for AL amyloidosis patients. [ABSTRACT FROM AUTHOR]

Amyloidosis can lead to cardiac, renal, and other multiorgan failure. New treatments have become available that can prolong survival but rely on early diagnosis. Manifestations of amyloidosis in hand surgery include carpal tunnel syndrome, trigger finger, peripheral neuropathy, and spontaneous distal biceps rupture. Often, these can predate systemic amyloidosis, offering hand surgeons an opportunity to diagnose patients with amyloidosis before systemic disease, refer them for treatment, and potentially alter disease course and prolong survival. In this review, we describe the pathophysiology and two most common subtypes of amyloidosis seen by hand surgeons. We provide guidance on biopsy practices and referral for patients with amyloidosis. Lastly, we provide a brief overview of the treatments for amyloidosis and their effect on disease course. [ABSTRACT FROM AUTHOR]

Introduction: Analysis of an individual's immunoglobulin (IG) gene repertoire requires the use of high-quality germline gene reference sets. When sets only contain alleles supported by strong evidence, AIRR sequencing (AIRR-seq) data analysis is more accurate and studies of the evolution of IG genes, their allelic variants and the expressed immune repertoire is therefore facilitated. Methods: The Adaptive Immune Receptor Repertoire Community (AIRR-C) IG Reference Sets have been developed by including only human IG heavy and light chain alleles that have been confirmed by evidence from multiple high-quality sources. To further improve AIRR-seq analysis, some alleles have been extended to deal with short 3' or 5' truncations that can lead them to be overlooked by alignment utilities. To avoid other challenges for analysis programs, exact paralogs (e.g. IGHV1-69*01 and IGHV1-69D*01) are only represented once in each set, though alternative sequence names are noted in accompanyingmetadata. Results and discussion: The Reference Sets include less than half the previously recognised IG alleles (e.g. just 198 IGHV sequences), and also include a number of novel alleles: 8 IGHV alleles, 2 IGKV alleles and 5 IGLV alleles. Despite their smaller sizes, erroneous calls were eliminated, and excellent coverage was achieved when a set of repertoires comprising over 4 million V(D)J rearrangements from 99 individuals were analyzed using the Sets. The versiontracked AIRR-C IG Reference Sets are freely available at the OGRDB website (https://ogrdb.airr-community.org/germline_sets/Human) and will be regularly updated to include newly observed and previously reported sequences that can be confirmed by new high-quality data. [ABSTRACT FROM AUTHOR]

Dynein, an ancient microtubule-based motor protein, performs diverse cellular functions in nearly all eukaryotic cells, with the exception of land plants. It has evolved into three subfamilies—cytoplasmic dynein-1, cytoplasmic dynein-2, and axonemal dyneins—each differentiated by their cellular functions. These megadalton complexes consist of multiple subunits, with the heavy chain being the largest subunit that generates motion and force along microtubules by converting the chemical energy of ATP hydrolysis into mechanical work. Beyond this catalytic core, the functionality of dynein is significantly enhanced by numerous non-catalytic subunits. These subunits are integral to the complex, contributing to its stability, regulating its enzymatic activities, targeting it to specific cellular locations, and mediating its interactions with other cofactors. The diversity of non-catalytic subunits expands dynein's cellular roles, enabling it to perform critical tasks despite the conservation of its heavy chains. In this review, we discuss recent findings and insights regarding these non-catalytic subunits. [ABSTRACT FROM AUTHOR]

Amyloidosis involving the lacrimal sac is extremely rare. In this study, we demonstrated a rare case of localized light chain amyloidosis in the lacrimal sac region. The lacrimal sac lesion presented as infiltrative with bony erosion. Given the slow growth of the lesion and the absence of a blood flow signal inside, we concluded that the lesion was less likely to be malignant. Complete removal of the lacrimal sac lesion combined with simultaneous lacrimal passage reconstruction was performed. The diagnosis of light chain amyloidosis was confirmed by histology. The surgical results were favorable, and no recurrence was observed over one-year follow-up. Our case report enriches the understanding of amyloid deposition in the ocular adnexa.

Amyloid is an amorphous, fibrillar material formed from various abnormally folded proteins that deposits locally or systemically. Over 95% of cases have been attributed to light chain deposition (AL) or transthyretin deposition (ATTR) amyloidosis. The basic investigations in the evaluation of cardiac amyloidosis include the electrocardiogram, echocardiography and cardiac biomarkers. Echocardiography in a patient with cardiac amyloidosis shows biatrial enlargement, biventricular hypertrophy, diastolic dysfunction, interatrial septal thickening, valvular thickening, a glistening appearance of the interventricular septum, and pericardial effusion. Magnetic resonance imaging can help distinguish amyloidosis from other causes of infiltrative/restrictive cardiomyopathy, from example, sarcoidosis, hemochromatosis, and Fabry disease based on characteristic enhancement patterns in these diseases. The latest Expert Consensus recommends that serum/urine immunofixation electrophoresis along with a serum free light chain assay must be done in all the cases of suspected cardiac amyloidosis. If the light chain assays are positive, we proceed with tissue diagnosis for confirmation of AL amyloidosis. If the screening assays are negative for monoclonal gammopathy, the next step is to obtain cardiac scintigraphy. If the nuclear scan is negative, but the index of suspicion remains high, an endomyocardial biopsy can be done. Once amyloid is demonstrated in histopathologic specimens, it must be typed to distinguish between AL and ATTR. The ideal method for this is tandem mass spectrometry, although this may not be widely available. It has a sensitivity of 88% and specificity of 96% higher than other techniques 23. In resource-poor settings, immunohistochemistry or immunoelectron microscopy can allow this distinction, although with lesser sensitivity. [ABSTRACT FROM AUTHOR]

Although most patients with cardiac amyloidosis are diagnosed with either light chain (AL) or transthyretin (ATTR) disease, coexisting amyloid subtypes can occur. We present three cases of coexisting AL and ATTR cardiac amyloidosis and demonstrate the importance of clinical history and endomyocardial biopsy in diagnosis of this rare entity.

IntroductionAnalysis of an individual’s immunoglobulin (IG) gene repertoire requires the use of high-quality germline gene reference sets. When sets only contain alleles supported by strong evidence, AIRR sequencing (AIRR-seq) data analysis is more accurate and studies of the evolution of IG genes, their allelic variants and the expressed immune repertoire is therefore facilitated.MethodsThe Adaptive Immune Receptor Repertoire Community (AIRR-C) IG Reference Sets have been developed by including only human IG heavy and light chain alleles that have been confirmed by evidence from multiple high-quality sources. To further improve AIRR-seq analysis, some alleles have been extended to deal with short 3’ or 5’ truncations that can lead them to be overlooked by alignment utilities. To avoid other challenges for analysis programs, exact paralogs (e.g. IGHV1-69*01 and IGHV1-69D*01) are only represented once in each set, though alternative sequence names are noted in accompanying metadata.Results and discussionThe Reference Sets include less than half the previously recognised IG alleles (e.g. just 198 IGHV sequences), and also include a number of novel alleles: 8 IGHV alleles, 2 IGKV alleles and 5 IGLV alleles. Despite their smaller sizes, erroneous calls were eliminated, and excellent coverage was achieved when a set of repertoires comprising over 4 million V(D)J rearrangements from 99 individuals were analyzed using the Sets. The version-tracked AIRR-C IG Reference Sets are freely available at the OGRDB website (https://ogrdb.airr-community.org/germline_sets/Human) and will be regularly updated to include newly observed and previously reported sequences that can be confirmed by new high-quality data.

Abstract Background Proliferative glomerulonephritis with monoclonal immunoglobulin G (IgG) deposits (PGNMID) is a disease entity with nonorganized granular glomerular deposition with monoclonal proteins of both heavy and light chains. Dysproteinemia was observed in only 30% of the patients with PGNMID. We herein report a case of PGNMID with discrepancy between serum and glomerular deposits. Case presentation The patient was a 50-year-old man who had been followed at a local clinic due to hypertension, type 2 diabetes, hyperlipidemia, hyperuricemia, fatty liver, and obesity. Proteinuria had been noted five years previously, and he had been referred to a hematology department due to hyperproteinemia, high gamma globulin, and κ Bence-Jones protein (BJP) positivity one year previously. Bone marrow aspiration showed 5% plasma cells, and he was referred to the nephrology department to evaluate persistent proteinuria. He was hypertensive, and his estimated glomerular filtration rate was 54.2 ml/min/1.73 m2. His urinary protein level was 0.84 g/g⋅Cr. Urine and serum immunofixation showed BJP-κ type and IgG-κ type, respectively. Kidney biopsy showed an increase in mesangial cells and matrix without nodular lesions under a light microscope. Immunofluorescence microscopy showed granular deposits of IgG and C3 on the capillary wall and weak positivity for C1q. IgG3 was predominant among the IgG subclasses, and intraglomerular κ and λ staining was negative for κ and positive for λ. Direct fast scarlet staining was negative. Electron microscopy showed lumpy deposits without a fibrillar structure in the subepithelial area. Based on the above findings, a diagnosis of membranous nephropathy-type PGNMID was made. Since proteinuria increased gradually after three years of treatment with valsartan (40 mg, daily), oral prednisolone (30 mg, daily) was initiated, which led to decreased proteinuria. The dose of oral prednisolone was gradually tapered to 10 mg per day. At that time, proteinuria was 0.88 g/g⋅Cr. We found 204 cases in 81 articles in the PubMed database, among which 8 showed discrepancy in the heavy and/or light chains between serum and kidney. Conclusions We experienced a case of membranous nephropathy-type PGNMID with discrepancy in light chains between serum and kidney that was successfully treated with oral prednisolone.

Objective·To screen the human-mouse chimeric antibodies that neutralize the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and analyse the key amino acid site changes in their light chain.Methods·Antibody heavy chain gene-humanized mice were immunized with the spike protein of SARS-CoV-2. Enzyme-linked immunosorbent assay (ELISA) and pseudovirus neutralization assay were performed to analyse the antibody titer in serum level. Flow cytometry was used to sort plasma cells and germinal center (GC) B cells from the mouse spleen and lymph nodes, and the proportions of different B cells were analysed. RNA was extracted to construct a mouse-origin antibody light chain gene library, which was paired with the knocked-in human-origin antibody heavy chain gene for antibody expression. ELISA was employed to screen for high-affinity chimeric antibodies. Amino acid changes in the light chain of high-affinity chimeric antibodies were analysed by comparing them with mouse-origin germline antibody light chain genes. Chimeric antibodies with single amino acid changes in the light chain were constructed and expressed, and their affinity and neutralization abilities were tested through ELISA and pseudovirus neutralization experiments. By comparing the affinity and neutralization abilities of the chimeric antibodies with or without single amino acid changes in the light chain, the key single amino acid change was analysed.Results·Compared to unimmunized mice, the immunized mice showed an increased titer of specific antibodies against SARS-CoV-2 on the serum level, along with an elevated proportion of plasma cells and GC B cells in the spleen and lymph nodes. ELISA showed that a high-affinity chimeric antibody was screened from GC B cells in the mouse spleen. The light chain of the antibody had a 76th serine-to-isoleucine substitution and a 98th phenylalanine deletion. Another ELISA showed that the chimeric antibody exhibited high affinity, the antibody without amino acid change on the light chain and the antibody with only the 98th phenylalanine deletion showed low affinity, and the antibody with only the 76th serine-to-isoleucine substitution demonstrated intermediate affinity between the chimeric antibody and the antibody without amino acid change on the light chain. Pseudovirus neutralization experiments revealed that the chimeric antibody had a half-maximal inhibitory concentration (IC50) of 0.995 ng/μL; the antibody with only the 76th serine-to-isoleucine substitution had an IC50 of 1.724 ng/μL, indicating a slight decrease in neutralization ability; the antibody with only the 98th phenylalanine deletion had an IC50 of 71.05 ng/μL; the antibody without amino acid change on the light chain had an IC50 of 42.06 ng/μL, suggesting weak neutralization ability.Conclusion·For the screened human-mouse chimeric antibodies that neutralize SARS-CoV-2 in this study, the key amino acid change determining the neutralization ability of the chimeric antibody is the 76th serine-to-isoleucine substitution.

Cardiac involvement is the foremost determinant of the clinical progression of amyloidosis. The diagnostic role of cardiac magnetic resonance (CMR) imaging in cardiac amyloidosis has been established, but the prognostic role of various right and left CMR tissue characterization and functional parameters, including global longitudinal strain (GLS), late gadolinium enhancement (LGE), and parametric mapping, is yet to be delineated. We searched EMBASE, PubMed, and MEDLINE for studies analysing the prognostic use of CMR imaging in patients with light chain amyloidosis or transthyretin amyloidosis cardiac amyloidosis. The primary endpoint was all-cause mortality. A random effects model was used to calculate a pooled odds ratio using inverse-variance weighting. Nineteen studies with 2199 patients [66% males, median age 59.7 years, interquartile range (IQR) 58–67] were included. Median follow-up was 24 months (IQR 20–32), during which 40.8% of patients died. Both tissue characterization left heart parameters such as elevated extracellular volume [hazard ratio (HR) 3.95, 95% confidence interval (CI) 3.01–5.17], extension of left ventricular (LV) LGE (HR 2.69, 95% CI 2.07–3.49) elevated native T1 (HR 2.19, 95% CI 1.12–4.28), and functional parameters such as reduced LV GLS (HR 1.91, 95% CI 1.52–2.41) and reduced LV ejection fraction (EF; HR 1.20, 95% CI 1.17–1.23) were associated with increased all-cause mortality. Unlike the presence of right ventricular (RV) LGE (HR 3.40, 95% CI 0.51–22.54), parameters such as RV GLS (HR 2.08, 95% CI 1.6–2.69), RVEF (HR 1.13, 95% CI 1.05–1.22), and tricuspid annular systolic excursion (TAPSE) (HR 1.11, 95% CI 1.02–1.21) were also associated with mortality. In this large meta-analysis of patients with cardiac amyloidosis, CMR parameters assessing RV and LV function and tissue characterization were associated with an increased risk of mortality. Graphical Abstract [ABSTRACT FROM AUTHOR]

Clostridium botulinum neurotoxins (BoNTs) are the most potent toxins known, causing the deadly disease botulism. They function through Zn2+-dependent endopeptidase cleavage of SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, preventing vesicular fusion and subsequent neurotransmitter release from motor neurons. Several serotypes of BoNTs produced by Clostridium botulinum (BoNT/A-/G and/X) have been well-characterised over the years. However, a BoNT-like gene (homologue of BoNT) was recently identified in the non-clostridial species, Enterococcus faecium, which is the leading cause of hospital-acquired multi-drug resistant infections. Here, we report the crystal structure of the catalytic domain of a BoNT homologue from Enterococcus faecium (LC/En) at 2.0 Å resolution. Detailed structural analysis in comparison with the full-length BoNT/En AlphaFold2-predicted structure, LC/A (from BoNT/A), and LC/F (from BoNT/F) revealed putative subsites and exosites (including loops 1–5) involved in recognition of LC/En substrates. LC/En also appears to possess a conserved autoproteolytic cleavage site whose function is yet to be established. [ABSTRACT FROM AUTHOR]

Objectives Amyloid light chain (AL)-κ and AL-λ share common histopathologic changes; however, the potential difference in clinical manifestations, histologic findings, and clinical significance between the 2 subtypes remain unclear. Methods In a retrospective study, 94 kidney biopsies for AL amyloidosis were evaluated using the composite scarring injury score (CSIS) and amyloid score (AS). Results were then compared between AL-κ and AL-λ. Results Comparing AS and CSIS between AL-κ and AL-λ, the AS was significantly higher in AL-κ than in AL-λ, with 2 components of AS (capillary wall and vascular amyloid) scoring higher in AL-κ than in AL-λ, while mesangial and interstitial ASs were similar in the 2 cohorts. In addition, the proportion of periodic acid–Schiff strong-staining amyloid in AL-κ was markedly higher than in AL-λ. There was no significant difference in CSIS and its components between the 2 subtypes of AL amyloidosis. Conclusions Overall, AL-κ presents with higher serum creatinine and a higher AS score than AL-λ at biopsy, which may indicate a worse prognosis and be an important reference for clinical management. [ABSTRACT FROM AUTHOR]

Abstract Background Systemic amyloidosis is group of disorders characterized by the accumulation of insoluble proteins in tissues. The most common form of systemic amyloidosis is light chain amyloidosis, which results from the accumulation of misfolded immunoglobulins. The disease is progressive, with treatment targeted at the underlying plasma cell dyscrasia. Since essentially any organ system can be affected, the presentation is variable and delays in diagnosis are common. Given this diagnostic difficulty, we discuss four different manifestations of light chain amyloidosis. Case presentations In this case series, we discuss four cases of light chain amyloidosis. These include cardiac, hepatic, and gastrointestinal as well as autonomic and peripheral nerve involvement with amyloidosis. The patients in our series are of Caucasian background and include a 69-year-old female, a 29-year-old female, a 68-year-old male, and a 70-year-old male, respectively. The case discussions highlight variability in presentation and diagnostic challenges. Conclusions Amyloidosis is a rare but serious disease that is often complicated by long delays in diagnosis. Morbidity and mortality can sometimes be limited if diagnosed earlier. We hope our real life cases will contribute to understanding and to early suspicion that can lead to early diagnosis and management.

Abstract Background In a range of human disorders such as multiple myeloma (MM), immunoglobulin light chains (IgLCs) can be produced at very high concentrations. This can lead to pathological aggregation and deposition of IgLCs in different tissues, which in turn leads to severe and potentially fatal organ damage. However, IgLCs can also be highly soluble and non-toxic. It is generally thought that the cause for this differential solubility behaviour is solely found within the IgLC amino acid sequences, and a variety of individual sequence-related biophysical properties (e.g. thermal stability, dimerisation) have been proposed in different studies as major determinants of the aggregation in vivo. Here, we investigate biophysical properties underlying IgLC amyloidogenicity. Results We introduce a novel and systematic workflow, Thermodynamic and Aggregation Fingerprinting (ThAgg-Fip), for detailed biophysical characterisation, and apply it to nine different MM patient-derived IgLCs. Our set of pathogenic IgLCs spans the entire range of values in those parameters previously proposed to define in vivo amyloidogenicity; however, none actually forms amyloid in patients. Even more surprisingly, we were able to show that all our IgLCs are able to form amyloid fibrils readily in vitro under the influence of proteolytic cleavage by co-purified cathepsins. Conclusions We show that (I) in vivo aggregation behaviour is unlikely to be mechanistically linked to any single biophysical or biochemical parameter and (II) amyloidogenic potential is widespread in IgLC sequences and is not confined to those sequences that form amyloid fibrils in patients. Our findings suggest that protein sequence, environmental conditions and presence and action of proteases all determine the ability of light chains to form amyloid fibrils in patients.

Light-chain (AL) amyloidosis is a rare plasma cell disorder characterized by the deposition of misfolded immunoglobulin light chains in target organs, leading to multi-organ dysfunction. Treatment approaches have historically mirrored but lagged behind those of multiple myeloma (MM). Recent advancements in MM immunotherapy are gradually being evaluated and adopted in AL amyloidosis. This review explores the current state of immunotherapeutic strategies in AL amyloidosis, including monoclonal antibodies, antibody–drug conjugates, bispecific antibodies, and chimeric antigen receptor T-cell therapy. We discuss the unique challenges and prospects of these therapies in AL amyloidosis, including the exposure of frail AL amyloidosis patients to immune-mediated toxicities such as cytokine release syndrome (CRS) and immune effector-cell-associated neurotoxicity syndrome (ICANS), as well as their efficacy in promoting rapid and deep hematologic responses. Furthermore, we highlight the need for international initiatives and compassionate programs to provide access to these promising therapies and address critical unmet needs in AL amyloidosis management. Finally, we discuss future directions, including optimizing treatment sequencing and mitigating toxicities, to improve outcomes for AL amyloidosis patients.

Dynein, an ancient microtubule-based motor protein, performs diverse cellular functions in nearly all eukaryotic cells, with the exception of land plants. It has evolved into three subfamilies—cytoplasmic dynein-1, cytoplasmic dynein-2, and axonemal dyneins—each differentiated by their cellular functions. These megadalton complexes consist of multiple subunits, with the heavy chain being the largest subunit that generates motion and force along microtubules by converting the chemical energy of ATP hydrolysis into mechanical work. Beyond this catalytic core, the functionality of dynein is significantly enhanced by numerous non-catalytic subunits. These subunits are integral to the complex, contributing to its stability, regulating its enzymatic activities, targeting it to specific cellular locations, and mediating its interactions with other cofactors. The diversity of non-catalytic subunits expands dynein’s cellular roles, enabling it to perform critical tasks despite the conservation of its heavy chains. In this review, we discuss recent findings and insights regarding these non-catalytic subunits.

Background: Proliferative glomerulonephritis with monoclonal immunoglobulin G (IgG) deposits (PGNMID) is a disease entity with nonorganized granular glomerular deposition with monoclonal proteins of both heavy and light chains. Dysproteinemia was observed in only 30% of the patients with PGNMID. We herein report a case of PGNMID with discrepancy between serum and glomerular deposits. Case presentation: The patient was a 50-year-old man who had been followed at a local clinic due to hypertension, type 2 diabetes, hyperlipidemia, hyperuricemia, fatty liver, and obesity. Proteinuria had been noted five years previously, and he had been referred to a hematology department due to hyperproteinemia, high gamma globulin, and κ Bence-Jones protein (BJP) positivity one year previously. Bone marrow aspiration showed 5% plasma cells, and he was referred to the nephrology department to evaluate persistent proteinuria. He was hypertensive, and his estimated glomerular filtration rate was 54.2 ml/min/1.73 m2. His urinary protein level was 0.84 g/g⋅Cr. Urine and serum immunofixation showed BJP-κ type and IgG-κ type, respectively. Kidney biopsy showed an increase in mesangial cells and matrix without nodular lesions under a light microscope. Immunofluorescence microscopy showed granular deposits of IgG and C3 on the capillary wall and weak positivity for C1q. IgG3 was predominant among the IgG subclasses, and intraglomerular κ and λ staining was negative for κ and positive for λ. Direct fast scarlet staining was negative. Electron microscopy showed lumpy deposits without a fibrillar structure in the subepithelial area. Based on the above findings, a diagnosis of membranous nephropathy-type PGNMID was made. Since proteinuria increased gradually after three years of treatment with valsartan (40 mg, daily), oral prednisolone (30 mg, daily) was initiated, which led to decreased proteinuria. The dose of oral prednisolone was gradually tapered to 10 mg per day. At that time, proteinuria was 0.88 g/g⋅Cr. We found 204 cases in 81 articles in the PubMed database, among which 8 showed discrepancy in the heavy and/or light chains between serum and kidney. Conclusions: We experienced a case of membranous nephropathy-type PGNMID with discrepancy in light chains between serum and kidney that was successfully treated with oral prednisolone. [ABSTRACT FROM AUTHOR]

The article reports a rare case of mantle cell lymphoma (MCL) with dual expression of kappa and lambda light chains, which is a phenomenon rarely observed in bursa (B) -cell non-Hodgkin lymphomas. It highlights the importance of careful analysis and avoiding diagnostic pitfalls when interpreting flow cytometry results, particularly in cases of dual expression, to ensure accurate diagnosis and appropriate treatment.

Objective ·To screen the human-mouse chimeric antibodies that neutralize the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and analyse the key amino acid site changes in their light chain. Methods ·Antibody heavy chain gene-humanized mice were immunized with the spike protein of SARS-CoV-2. Enzyme-linked immunosorbent assay (ELISA) and pseudovirus neutralization assay were performed to analyse the antibody titer in serum level. Flow cytometry was used to sort plasma cells and germinal center (GC) B cells from the mouse spleen and lymph nodes, and the proportions of different B cells were analysed. RNA was extracted to construct a mouse-origin antibody light chain gene library, which was paired with the knocked-in human-origin antibody heavy chain gene for antibody expression. ELISA was employed to screen for high-affinity chimeric antibodies. Amino acid changes in the light chain of high-affinity chimeric antibodies were analysed by comparing them with mouse-origin germline antibody light chain genes. Chimeric antibodies with single amino acid changes in the light chain were constructed and expressed, and their affinity and neutralization abilities were tested through ELISA and pseudovirus neutralization experiments. By comparing the affinity and neutralization abilities of the chimeric antibodies with or without single amino acid changes in the light chain, the key single amino acid change was analysed. Results ·Compared to unimmunized mice, the immunized mice showed an increased titer of specific antibodies against SARS-CoV-2 on the serum level, along with an elevated proportion of plasma cells and GC B cells in the spleen and lymph nodes. ELISA showed that a high-affinity chimeric antibody was screened from GC B cells in the mouse spleen. The light chain of the antibody had a 76th serine-to-isoleucine substitution and a 98th phenylalanine deletion. Another ELISA showed that the chimeric antibody exhibited high affinity, the antibody without amino acid change on the light chain and the antibody with only the 98th phenylalanine deletion showed low affinity, and the antibody with only the 76th serine-to-isoleucine substitution demonstrated intermediate affinity between the chimeric antibody and the antibody without amino acid change on the light chain. Pseudovirus neutralization experiments revealed that the chimeric antibody had a half-maximal inhibitory concentration (IC50) of 0.995 ng/μL; the antibody with only the 76th serine-to-isoleucine substitution had an IC50 of 1.724 ng/μL, indicating a slight decrease in neutralization ability; the antibody with only the 98th phenylalanine deletion had an IC50 of 71.05 ng/μL; the antibody without amino acid change on the light chain had an IC50 of 42.06 ng/μL, suggesting weak neutralization ability. Conclusion ·For the screened human-mouse chimeric antibodies that neutralize SARS-CoV-2 in this study, the key amino acid change determining the neutralization ability of the chimeric antibody is the 76th serine-to-isoleucine substitution. [ABSTRACT FROM AUTHOR]

Background Increased intracardiac thrombus formation in light-chain cardiac amyloidosis (AL-CA) has been associated with thrombotic events such as stroke and high rates of mortality and morbidity. Case summary A 51-year-old man was admitted to the emergency department with a sudden change in consciousness. His emergency brain magnetic resonance imaging showed two foci of cerebral infarction on the bilateral temporal lobes. His electrocardiogram displayed a normal sinus rhythm with low QRS voltage. Transthoracic echocardiography showed concentric thickened ventricles, dilatation of both atria, a left ventricular ejection fraction of 53%, and a Grade 3 diastolic dysfunction. The bull-eye plot on speckle tracking echocardiography displayed a distinctive apical sparing pattern. A serum-free immunoglobulin analysis showed increased lambda-free light chains (295.59 mg/L) with a reduced kappa/lambda ratio (0.08). Light-chain amyloidosis was subsequently confirmed by examining the histology of the abdominal fat-pad tissue. On transoesophageal echocardiography (TEE), an elongated static thrombus and a mobile bouncing oval thrombus were found on the left and right atrial appendages, respectively. Atrial thrombi were managed with a full dose of dabigatran of 150 mg twice daily, which resulted in a complete resolution after 2 months of TEE follow-up. Discussion Complicating intracardiac thrombosis has been considered one of the major contributions of death in cardiac amyloidosis. Transoesophageal echocardiography should be established to aid in the detection and management of atrial thrombus in AL-CA. [ABSTRACT FROM AUTHOR]

Waldenström macroglobulinemia (WM) is a rare lymphoplasmacytic disorder characterized by an IgM paraprotein. The clinical presentation of WM varies and can include common manifestations such as anemia and hyperviscosity, in addition to less common features such as cryoglobulinemia, IgM-related neuropathy, and immunoglobulin light chain (AL) amyloidosis. Amyloidosis is a protein-folding disorder in which vital organ damage occurs due to the accumulation of misfolded protein aggregates. The most common type of amyloidosis in patients with an IgM paraprotein is AL amyloidosis, although other types of amyloidosis may occur. IgM-related amyloidosis has distinct clinical features when compared with other subtypes of AL amyloidosis. This review highlights the diagnostic criteria of IgM-related AL amyloidosis, as well as the clinical characteristics and treatment options for this disorder.

Background: Systemic amyloidosis is group of disorders characterized by the accumulation of insoluble proteins in tissues. The most common form of systemic amyloidosis is light chain amyloidosis, which results from the accumulation of misfolded immunoglobulins. The disease is progressive, with treatment targeted at the underlying plasma cell dyscrasia. Since essentially any organ system can be affected, the presentation is variable and delays in diagnosis are common. Given this diagnostic difficulty, we discuss four different manifestations of light chain amyloidosis. Case presentations: In this case series, we discuss four cases of light chain amyloidosis. These include cardiac, hepatic, and gastrointestinal as well as autonomic and peripheral nerve involvement with amyloidosis. The patients in our series are of Caucasian background and include a 69-year-old female, a 29-year-old female, a 68-year-old male, and a 70-year-old male, respectively. The case discussions highlight variability in presentation and diagnostic challenges. Conclusions: Amyloidosis is a rare but serious disease that is often complicated by long delays in diagnosis. Morbidity and mortality can sometimes be limited if diagnosed earlier. We hope our real life cases will contribute to understanding and to early suspicion that can lead to early diagnosis and management. [ABSTRACT FROM AUTHOR]

The XpressCF+® cell-free protein synthesis system is a robust platform for the production of non-natural amino acids containing antibodies, which enable the site-specific conjugation of homogeneous antibody drug conjugates (ADCs) via click chemistry. Here, we present a robust and scalable means of achieving a 50–100% increase in IgG titers by combining the high productivity of cell-based protein synthesis with the unique ability of XpressCF+® reactions to produce correctly folded and assembled IgGs containing multiple non-natural amino acids at defined positions. This hybrid technology involves the pre-expression of an IgG light-chain (LC) protein in a conventional recombinant E. coli expression system, engineered to have an oxidizing cytoplasm. The prefabricated LC subunit is then added as a reagent to the cell-free protein synthesis reaction. Prefabricated LC increases IgG titers primarily by reducing the protein synthesis burden per IgG since the cell free translation machinery is only responsible for synthesizing the HC protein. Titer increases were demonstrated in four IgG products in scales ranging from 100-µL microplate reactions to 0.25-L stirred tank bioreactors. Similar titer increases with prefabricated LC were also demonstrated for a bispecific antibody in the scFvFc-FabFc format, demonstrating the generality of this approach. Prefabricated LC also increases robustness in cell-free reactions since it eliminates the need to fine-tune the HC-to-LC plasmid ratio, a critical parameter influencing IgG assembly and quality when the two IgG subunits are co-expressed in a single reaction. ADCs produced using prefabricated LC were shown to be identical to IgGs produced in cell-free alone by comparing product quality, in vitro cell killing, and FcRn receptor binding assays. This approach represents a significant step towards improving IgG titers and the robustness of cell-free protein synthesis reactions by integrating in vivo and in vitro protein production platforms. [ABSTRACT FROM AUTHOR]

Background: In a range of human disorders such as multiple myeloma (MM), immunoglobulin light chains (IgLCs) can be produced at very high concentrations. This can lead to pathological aggregation and deposition of IgLCs in different tissues, which in turn leads to severe and potentially fatal organ damage. However, IgLCs can also be highly soluble and non-toxic. It is generally thought that the cause for this differential solubility behaviour is solely found within the IgLC amino acid sequences, and a variety of individual sequence-related biophysical properties (e.g. thermal stability, dimerisation) have been proposed in different studies as major determinants of the aggregation in vivo. Here, we investigate biophysical properties underlying IgLC amyloidogenicity. Results: We introduce a novel and systematic workflow, Thermodynamic and Aggregation Fingerprinting (ThAgg-Fip), for detailed biophysical characterisation, and apply it to nine different MM patient-derived IgLCs. Our set of pathogenic IgLCs spans the entire range of values in those parameters previously proposed to define in vivo amyloidogenicity; however, none actually forms amyloid in patients. Even more surprisingly, we were able to show that all our IgLCs are able to form amyloid fibrils readily in vitro under the influence of proteolytic cleavage by co-purified cathepsins. Conclusions: We show that (I) in vivo aggregation behaviour is unlikely to be mechanistically linked to any single biophysical or biochemical parameter and (II) amyloidogenic potential is widespread in IgLC sequences and is not confined to those sequences that form amyloid fibrils in patients. Our findings suggest that protein sequence, environmental conditions and presence and action of proteases all determine the ability of light chains to form amyloid fibrils in patients. [ABSTRACT FROM AUTHOR]

Systemic light-chain amyloidosis (AL) is a human disease caused by overexpression of monoclonal immunoglobulin light chains that form pathogenic amyloid fibrils. These amyloid fibrils deposit in tissues and cause organ failure. Proteins form amyloid fibrils when they partly or fully unfold and expose segments capable of stacking into β-sheets that pair and thereby form a tight, dehydrated interface. These structures, termed steric zippers, constitute the spines of amyloid fibrils. Here, using a combination of computational (with ZipperDB and Boston University ALBase), mutational, biochemical, and protein structural analyses, we identified segments within the variable domains of Ig light chains that drive the assembly of amyloid fibrils in AL. We demonstrate that there are at least two such segments and that each one can drive amyloid fibril assembly independently of the other. Our analysis revealed that peptides derived from these segments form steric zippers featuring a typical dry interface with high-surface complementarity and occupy the same spatial location of the Greek-key immunoglobulin fold in both λ and κ variable domains. Of note, some predicted steric-zipper segments did not form amyloid fibrils or assembled into fibrils only when removed from the whole protein. We conclude that steric-zipper propensity must be experimentally validated and that the two segments identified here may represent therapeutic targets. In addition to elucidating the molecular pathogenesis of AL, these findings also provide an experimental approach for identifying segments that drive fibril formation in other amyloid diseases.

In light chain amyloidosis, a reduction in dFLC to below 40 mg/l is a prerequisite for organ recovery as nearly half of the patients who achieve very good partial haematological responses have improvement in the function of the involved organ. We describe a patient who developed new-onset cardiac amyloidosis despite a post-treatment reduction in dFLC to

The aim of this work was to compare the classification of cardiac MR-images of AL versus ATTR amyloidosis by neural networks and by experienced human readers. Cine-MR images and late gadolinium enhancement (LGE) images of 120 patients were studied (70 AL and 50 TTR). A VGG16 convolutional neural network (CNN) was trained with a 5-fold cross validation process, taking care to strictly distribute images of a given patient in either the training group or the test group. The analysis was performed at the patient level by averaging the predictions obtained for each image. The classification accuracy obtained between AL and ATTR amyloidosis was 0.750 for cine-CNN, 0.611 for Gado-CNN and between 0.617 and 0.675 for human readers. The corresponding AUC of the ROC curve was 0.839 for cine-CNN, 0.679 for gado-CNN (p < 0.004 vs. cine) and 0.714 for the best human reader (p < 0.007 vs. cine). Logistic regression with cine-CNN and gado-CNN, as well as analysis focused on the specific orientation plane, did not change the overall results. We conclude that cine-CNN leads to significantly better discrimination between AL and ATTR amyloidosis as compared to gado-CNN or human readers, but with lower performance than reported in studies where visual diagnosis is easy, and is currently suboptimal for clinical practice. [ABSTRACT FROM AUTHOR]

Waldenström macroglobulinemia (WM) is a rare lymphoplasmacytic disorder characterized by an IgM paraprotein. The clinical presentation of WM varies and can include common manifestations such as anemia and hyperviscosity, in addition to less common features such as cryoglobulinemia, IgM-related neuropathy, and immunoglobulin light chain (AL) amyloidosis. Amyloidosis is a protein-folding disorder in which vital organ damage occurs due to the accumulation of misfolded protein aggregates. The most common type of amyloidosis in patients with an IgM paraprotein is AL amyloidosis, although other types of amyloidosis may occur. IgM-related amyloidosis has distinct clinical features when compared with other subtypes of AL amyloidosis. This review highlights the diagnostic criteria of IgM-related AL amyloidosis, as well as the clinical characteristics and treatment options for this disorder. [ABSTRACT FROM AUTHOR]

Studies carried out in the last three decades have significantly advanced our knowledge about the structural factors that drive the amyloid aggregation of the immunoglobulin light chains. Solid-state nuclear magnetic resonance and cryo-electron microscopy studies have resulted in huge progress in our knowledge about the AL fibril structure. Now, it is known that the assembly of the light chain into AL fibrils implies an extensive conformational rearrangement that converts the beta-sandwich fold of the protein into a near flat structure. On the other hand, there has also been significant progress made in understanding the role that some cell types play as facilitators of AL formation. Such a role has been studied in glomerular amyloidosis, where mesangial cells play an important role in the mechanism of AL deposition, as well as for the pathogenic mechanisms that result in glomerular/renal damage. This review addresses what we currently know about why and how certain light chains are prone to forming amyloid. It also summarizes the most recent publications on the structure of AL fibrils and analyzes the structural bases of this type of aggregate, including the origin of its structural diversity. Finally, the most relevant findings on the role of mesangial cells in the amyloid deposition of light chains in the glomerular space are summarized.

Clostridium botulinum neurotoxins (BoNTs) are the most potent toxins known, causing the deadly disease botulism. They function through Zn2+-dependent endopeptidase cleavage of SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptor) proteins, preventing vesicular fusion and subsequent neurotransmitter release from motor neurons. Several serotypes of BoNTs produced by Clostridium botulinum (BoNT/A-/G and/X) have been well-characterised over the years. However, a BoNT-like gene (homologue of BoNT) was recently identified in the non-clostridial species, Enterococcus faecium, which is the leading cause of hospital-acquired multi-drug resistant infections. Here, we report the crystal structure of the catalytic domain of a BoNT homologue from Enterococcus faecium (LC/En) at 2.0 Å resolution. Detailed structural analysis in comparison with the full-length BoNT/En AlphaFold2-predicted structure, LC/A (from BoNT/A), and LC/F (from BoNT/F) revealed putative subsites and exosites (including loops 1–5) involved in recognition of LC/En substrates. LC/En also appears to possess a conserved autoproteolytic cleavage site whose function is yet to be established.

The extreme toxicity of botulinum neurotoxins (BoNTs) relies on their specific cleavage of SNARE proteins, which eventually leads to muscle paralysis. One newly identified mosaic toxin, BoNT/HA (aka H or FA), cleaves VAMP-2 at a unique position between residues L54 and E55, but the molecular basis underlying VAMP-2 recognition of BoNT/HA remains poorly characterized. Here, we report a ∼2.09 Å resolution crystal structure of the light chain protease domain of BoNT/HA (LC/HA). Structural comparison between LC/HA and LC of BoNT/F1 (LC/F1) reveals distinctive hydrophobic and electrostatic features near the active sites, which may explain their different VAMP-2 cleavage sites. When compared to BoNT/F5 that cleaves VAMP-2 at the same site as BoNT/HA, LC/HA displays higher affinity for VAMP-2, which could be caused by their different surface charge properties surrounding a VAMP-2 exosite-binding cleft. Furthermore, systematic mutagenesis studies on VAMP-2 and structural modeling demonstrate that residues R47 to K59 spanning the cleavage site in VAMP-2 may adopt a novel extended conformation when interacting with LC/HA and LC/F5. Taken together, our structure provides new insights into substrate recognition of BoNT/HA and paves the way for rational design of small molecule or peptide inhibitors against LC/HA.

The engineering of antibodies and antibody fragments for affinity maturation, stability, and other biophysical characteristics is a common aspect of therapeutic development. Maturation of antibodies in B cells during the adaptive immune response is the result of a process called somatic hypermutation (SHM), in which the activation-induced cytidine deaminase (AID) acts to introduce mutations into immunoglobulin (Ig) genes. Iterative selection and clonal expansion of B cells containing affinity-enhancing mutations drive an increase in the overall affinity of antibodies. Here we describe the use of SHM coupled with mammalian cell surface display for the maturation of antibodies in vitro and the complementarity of these methods with the mining of immune lineages using next-generation sequencing (NGS).