Immune Complex Formation Research Articles

Amyloid-β (Aβ) immunotherapy induced microhemorrhages are associated with activated perivascular macrophages and peripheral monocyte recruitment in Alzheimer’s disease mice

BackgroundAmyloid-related imaging abnormalities (ARIA) have been identified as the most common and serious adverse events resulting from pathological changes in the cerebral vasculature during several recent anti-amyloid-β (Aβ) immunotherapy trials. However, the precise cellular and molecular mechanisms underlying how amyloid immunotherapy enhances cerebral amyloid angiopathy (CAA)-mediated alterations in vascular permeability and microhemorrhages are not currently understood. Interestingly, brain perivascular macrophages have been implicated in regulating CAA deposition and cerebrovascular function however, further investigations are required to understand how perivascular macrophages play a role in enhancing CAA-related vascular permeability and microhemorrhages associated with amyloid immunotherapy.MethodsIn this study, we examined immune responses induced by amyloid-targeting antibodies and CAA-induced microhemorrhages using histology and gene expression analyses in Alzheimer’s disease (AD) mouse models and primary culture systems.ResultsIn the present study, we demonstrate that anti-Aβ (3D6) immunotherapy leads to the formation of an antibody immune complex with vascular amyloid deposits and induces the activation of CD169+ perivascular macrophages. We show that macrophages activated by antibody mediated Fc receptor signaling have increased expression of inflammatory signaling and extracellular matrix remodeling genes such as Timp1 and MMP9 in vitro and confirm these key findings in vivo. Finally, we demonstrate enhanced vascular permeability of plasma proteins and recruitment of inflammatory monocytes around vascular amyloid deposits, which are associated with hemosiderin deposits from cerebral microhemorrhages, suggesting the multidimensional roles of activated perivascular macrophages in response to Aβ immunotherapy.ConclusionsIn summary, our study establishes a connection between Aβ antibodies engaged at CAA deposits, the activation of perivascular macrophages, and the upregulation of genes involved in vascular permeability. However, the implications of this phenomenon on the susceptibility to microhemorrhages remain to be fully elucidated. Further investigations are warranted to determine the precise role of CD169 + perivascular macrophages in enhancing CAA-mediated vascular permeability, extravasation of plasma proteins, and infiltration of immune cells associated with microhemorrhages.

Revealing the SARS-CoV-2 Spike Protein and Specific Antibody Immune Complex Formation Mechanism for Precise Evaluation of Antibody Affinity.

The profound understanding and detailed evaluation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (SCoV2-S) protein and specific antibody interaction mechanism is of high importance in the development of immunosensors for COVID-19. In the present work, we studied a model system of immobilized SCoV2-S protein and specific monoclonal antibodies by molecular dynamics of immune complex formation in real time. We simultaneously applied spectroscopic ellipsometry and quartz crystal microbalance with dissipation to reveal the features and steps of the immune complex formation. We showed direct experimental evidence based on acoustic and optical measurements that the immune complex between covalently immobilized SCoV2-S and specific monoclonal antibodies is formed in two stages. Based on these findings it was demonstrated that applying a two-step binding mathematical model for kinetics analysis leads to a more precise determination of interaction rate constants than that determined by the 1:1 Langmuir binding model. Our investigation showed that the equilibrium dissociation constants (KD) determined by a two-step binding model and the 1:1 Langmuir model could differ significantly. The reported findings can facilitate a deeper understanding of antigen-antibody immune complex formation steps and can open a new way for the evaluation of antibody affinity towards corresponding antigens.

Review: Advances in the Pathogenesis and Treatment of Immune Thrombocytopenia Associated with Viral Hepatitis.

Hepatitis B virus and hepatitis C virus are the hepatitis subtypes that most commonly induce immune thrombocytopenia (ITP). Although the pathogenesis of viral hepatitis-associated ITP remains unclear, it may involve antibody cross-reactivity due to molecular mimicry, the formation of virus-platelet immune complexes, and T cell-mediated suppression of bone marrow hematopoiesis. Moreover, there is significant correlation between platelet count and the severity of viral hepatitis, the risk of progression to liver cirrhosis, and clinical prognosis. However, treatment of viral hepatitis-associated ITP is hindered by some antiviral drugs. In this review, we summarize research progress to date on the pathogenesis and treatment of viral hepatitis-related ITP, hoping to provide a reference for clinical diagnosis and treatment.

Positive and negative regulation of the Fcγ receptor-stimulating activity of RNA-containing immune complexes by RNase.

The U1RNP complex, Ro/SSA, and La/SSB are major RNA-containing autoantigens. Immune complexes (ICs) composed of RNA-containing autoantigens and autoantibodies are suspected to be involved in the pathogenesis of some systemic autoimmune diseases. Therefore, RNase treatment, which degrades RNA in ICs, has been tested in clinical trials as a potential therapeutic agent. However, no studies to our knowledge have specifically evaluated the effect of RNase treatment on the Fcγ receptor-stimulating (FcγR-stimulating) activity of RNA-containing ICs. In this study, using a reporter system that specifically detects FcγR-stimulating capacity, we investigated the effect of RNase treatment on the FcγR-stimulating activity of RNA-containing ICs composed of autoantigens and autoantibodies from patients with systemic autoimmune diseases such as systemic lupus erythematosus. We found that RNase enhanced the FcγR-stimulating activity of Ro/SSA- and La/SSB-containing ICs, but attenuated that of the U1RNP complex-containing ICs. RNase decreased autoantibody binding to the U1RNP complex, but increased autoantibody binding to Ro/SSA and La/SSB. Our results suggest that RNase enhances FcγR activation by promoting the formation of ICs containing Ro/SSA or La/SSB. Our study provides insights into the pathophysiology of autoimmune diseases involving anti-Ro/SSA and anti-La/SSB autoantibodies, and into the therapeutic application of RNase treatment for systemic autoimmune diseases.

Dynamics of acute-phase and endothelial reactions and immune complex formation during bone replacement with germanium-doped calcium-phosphate ceramics of bone fragment fractures in dogs

Bone defects that occur with fragment fractures lead to an increase in the number of postoperative complications. Therefore, to restore the structure and function of the bone, there is a need for bone replacement, in particular doped calcium-phosphate ceramics. The purpose of the work is the biochemical assessment of acute-phase and endothelial reactions and immune complex formation during bone replacement with germanium-doped calcium-phosphate ceramics of bone fragment fractures in dogs. Dogs with fragment fractures of long tubular bones and plate osteosynthesis were included in the study. In the experimental group (n = 10), bone defects were replaced with germanium-doped calcium-phosphate ceramics (HА/β-TCP/l-Gе-700), in the control group (n = 10) – unalloyed (HА/β-TCP-700). Blood samples were taken after the injury and on the 3rd, 7th, 14th, 30th, and 60th days after osteosynthesis. Partial recovery of limb function in the experimental group was faster by 1.3 times (P < 0.001) and full by 1.2 times (P < 0.01) compared to the control. On the 60th day, in experimental animals, the defect was filled with a regenerate of high X-ray density without a periosteal reaction, while in the control animals, the bone regenerate did not have sufficient density with a significant periosteal reaction. In both groups, total protein and albumin content varied within the normal range. The activity of protein C on the 3rd day in the experimental group was 1.3 times higher (Р ˂ 0.001), and on the 7th day, it was twice as high (P ˂ 0.001) compared to the control group with normalization by the 14th day. The concentration of ceruloplasmin from the 7th day in the control animals was 1.1 times higher (Р ˂ 0.001) than in the experimental animals, with normalization in the latter by the 60th day. The level of small molecular circulating immune complexes (CIC) after the injury increased by 1.1 times (Р ˂ 0.001) and reached a peak in the control and experimental groups on the 14th day, with an increase in indicators by 2.1 and 1.4 times (Р ˂ 0.001), respectively, with normalization in the experimental group on the 60th day. The level of nitric oxide (NO) in the control group increased from the 7th to the 60th day, with a peak on the 30th day, and in the experimental group – from the 3rd to the 30th, with a peak on the seventh day. Osteoreplacement of fragment fractures of long tubular bones in dogs with calcium-phosphate ceramics doped with germanium is accompanied by a moderate level of the acute phase reaction and immune complex formation, an increase in the endothelial reaction and the anticoagulant potential of the blood, which contributes to a decrease in the intensity of the inflammatory-resorptive stage of reparative osteogenesis and an increase in its proliferative phase, which, respectively, accelerates the consolidation of fractures.

Study of SARS-CoV-2 Spike Protein Wild-Type and the Variants of Concern Real-Time Interactions with Monoclonal Antibodies and Convalescent Human Serum.

The spike (S) protein and its receptor-binding domain (RBD) of the coronavirus SARS-CoV-2 have been continually evolving, yielding the majority of significant missense mutations and new variants of concern. In this study, we examined how monoclonal antibodies against RBD (mAbs-SCoV2-RBD) and polyclonal antibodies present in convalescent human serum specifically interact with the S protein of wild-type and SARS-CoV-2 variants of concern (VOCs) in real time and how this can be reflected through surface mass density. Moreover, we combined two distinct, label-free measurement techniques: one based on changes in surface electromagnetic waves after reflection from the surface, and the other on changes in acoustic waves. The results demonstrated that dry surface mass density (ΓSE) of mAbs-SCoV2-RBD attached to the RBD of the S protein decreases three-fold, from 148 ng/cm2 to 46 ng/cm2, due to the B.1.351 or so-called beta mutation of coronavirus and its S protein (SCoV2-β). Consequently, the obtained wet mass ΓQCM-D resulted in values two times lower, from 319 ng/cm2 to 158 ng/cm2, and the hydration of mAbs-SCoV2-RBD/SCoV2-β immune complex was 70.88%. Conversely, when polyclonal antibodies present in convalescent human serum form immune complexes with the S protein of SARS-CoV-2 variants of concern, the ΓSE decreased from 279 ng/cm2 to 249 ng/cm2, and ΓQCM-D from 1545 ng/cm2 to 1366 ng/cm2. These results can give insights into the differences between the interaction of monoclonal and polyclonal antibodies with SARS-CoV-2 VOCs.

Extracellular vesicles of Trypanosoma cruzi and immune complexes they form with sialylated and non-sialylated IgGs increase small peritoneal macrophage subpopulation and elicit different cytokines profiles.

American trypanosomiasis, or Chagas disease, is caused by the protozoan parasite Trypanosoma cruzi and is characterized by the presence of cardiac or gastrointestinal symptoms in a large number of patients during the chronic phase of the disease. Although the origin of the symptoms is not clear, several mechanisms have been described involving factors related to T. cruzi and the host immune response. In this sense, the extracellular vesicles (EVs) secreted by the parasite and the immune complexes (ICs) formed after their recognition by host IgGs (EVs-IgGs) may play an important role in the immune response during infection. The aim of the present work is to elucidate the modulation of the immune response exerted by EVs and the ICs they form by analyzing the variation in the subpopulations of small and large peritoneal macrophages after intraperitoneal inoculation in mice and to evaluate the role of the sialylation of the host IgGs in this immunomodulation. Both macrophage subpopulations were purified and subjected to cytokine expression analysis by RT-qPCR. The results showed an increase in the small peritoneal macrophage subpopulation after intraperitoneal injection of parasite EVs, but a greater increase in this subpopulation was observed when sialylated and non-sialylated ICs were injected, which was similar to inoculation with the trypomastigote stage of the parasite. The cytokine expression results showed the ability of both subpopulations to express inflammatory and non-inflammatory cytokines. These results suggest the role of free EVs in the acute phase of the disease and the possible role of immune complexes in the immune response in the chronic phase of the disease, when the levels of antibodies against the parasite allow the formation of immune complexes. The differential expression of interleukins showed after the inoculation of immune complexes formed with sialylated and non-sialylated IgGs and the interleukins expression induced by EVs, demonstrates that the IgG glycosilation is involved in the type of immune response that dominates in each of the phases of the Chagas disease.

Клинико-лабораторные исследования в комплексной диагностике и лечении увеитов у детей

This work is devoted to the analysis of the effectiveness of new clinical and laboratory studies and their possible application in treatment of uveitis in children. The paper assesses the study of the glutathione-reducing system as a factor in the vulnerability of eye tissues to the effects of free radicals and peroxidation products during inflammation. The dependence of the clinical picture of uveitis and the frequency of detection of eye tissue antigens in the blood was analyzed to assess the localization and stage of the pathological process, which will undoubtedly determine the tactics of the therapy. Evaluation of the main indicators of the activity of peroxidation processes in uveitis is proposed to be carried out according to the original method for determining the functional activity of polymorphonuclear leukocytes (PNLs). It was proved that there is a close relationship between the level of PNLs activity, the level of intensity of peroxide processes and the ophthalmic picture of uveitis. Based on the results obtained in the following work, the use of pharmaceutical antioxidants, both traditional and those known drugs in which antioxidant properties were discovered for the first time, was proposed. This paper offers a concept of the development of toxic-allergic uveitis, including successive stages of the formation of immune complexes, inflammation, an increase in the functional activity of PMNs, their chemotaxis into the inflammation zone, infiltration through the capillary walls in the eye tissue, which leads to an avalanche-like increase in the level of oxidants, depletion of endogenous reserves of antioxidants, damage to cell membranes. Understanding these processes serves as an explication for developing optimal treatment regimens for uveitis and other inflammatory ophthalmpathology. Keywords: uveitis, retinal antigens, glutathione system, antioxidants, polymorphonuclear leukocytes, peroxidation, reactive oxygen species

Immunoglobulin G4 in eosinophilic esophagitis: Immune complex formation and correlation with disease activity.

Recent studies have shown deposition of immunoglobulin G4 (IgG4) and food proteins in the esophageal mucosa of eosinophilic esophagitis (EoE) patients. Our aims were to assess whether co-localization of IgG4 and major cow's milk proteins (CMPs) was associated with EoE disease activity and to investigate the proteins enriched in proximity to IgG4 deposits. This study included adult subjects with EoE (n = 13) and non-EoE controls (n = 5). Esophageal biopsies were immunofluorescence stained for IgG4 and CMPs. Co-localization in paired samples from active disease and remission was assessed and compared to controls. The proteome surrounding IgG4 deposits was evaluated by the novel technique, AutoSTOMP. IgG4-food protein interactions were confirmed with co-immunoprecipitation and mass spectrometry. IgG4-CMP co-localization was higher in the active EoE group compared to paired remission samples (Bos d 4, p = .02; Bos d 5, p = .002; Bos d 8, p = .002). Co-localization was also significantly higher in the active EoE group compared to non-EoE controls (Bos d 4, p = .0013; Bos d 5, p = .0007; Bos d 8, p = .0013). AutoSTOMP identified eosinophil-derived proteins (PRG 2 and 3, EPX, RNASE3) and calpain-14 in IgG4-enriched areas. Co-immunoprecipitation and mass spectrometry confirmed IgG4 binding to multiple food allergens. These findings further contribute to the understanding of the interaction of IgG4 with food antigens as it relates to EoE disease activity. These data strongly suggest the immune complex formation of IgG4 and major cow's milk proteins. These immune complexes may have a potential role in the pathophysiology of EoE by contributing to eosinophil activation and disease progression.

Whole blood hydroxychloroquine: Does genetic polymorphism of cytochrome P450 enzymes have a role?

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with a wide range of clinical manifestations and multifactorial etiologies ranging from environmental to genetic. SLE is associated with dysregulated immunological reactions, with increased immune complex formation leading to end-organ damages such as lupus nephritis, cutaneous lupus, and musculoskeletal disorders. Lupus treatment aims to reduce disease activity, prevent organ damage, and improve long-term patient survival and quality of life. Antimalarial, hydroxychloroquine (HCQ) is used as a first-line systemic treatment for lupus. It has shown profound efficacy in lupus and its associated conditions. However, wide variation in terms of clinical response to this drug has been observed among this group of patients. This variability has limited the potential of HCQ to achieve absolute clinical benefits. Several factors, including genetic polymorphisms of cytochrome P450 enzymes, have been stipulated as key entities leading to this inter-individual variation. Thus, there is a need for more studies to understand the role of genetic polymorphisms in CYP450 enzymes in the clinical response to HCQ. Focusing on the role of genetic polymorphism on whole blood HCQ in lupus disorder, this review aims to highlight up-to-date pathophysiology of SLE, the mechanism of action of HCQ, and finally the role of genetic polymorphism of CYP450 enzymes on whole blood HCQ level as well as clinical response in lupus.

BAK1 protects the receptor-like kinase BIR2 from SNIPER2a/b-mediated degradation to promote pattern-triggered immunity in Nicotiana benthamiana.

The detection of microbial infections by plants induces the rapid formation of immune receptor complexes at the plasma membrane. However, how this process is controlled to ensure proper immune signaling remains largely unknown. Here, we found that the Nicotiana benthamiana membrane-localized leucine-rich repeat receptor-like kinase BAK1-INTERACTING RLK 2 (NbBIR2) constitutively associates with BRI1-ASSOCIATED RECEPTOR KINASE 1 (NbBAK1) in vivo and in vitro and promotes complex formation with pattern recognition receptors. In addition, NbBIR2 is targeted by 2 RING-type ubiquitin E3 ligases, SNC1-INFLUENCING PLANT E3 LIGASE REVERSE 2a (NbSNIPER2a) and NbSNIPER2b, for ubiquitination and subsequent degradation in planta. NbSNIPER2a and NbSNIPER2b interact with NbBIR2 in vivo and in vitro and are released from NbBIR2 upon treatment with different microbial patterns. Furthermore, accumulation of NbBIR2 in response to microbial patterns is tightly associated with NbBAK1 abundance in N. benthamiana. NbBAK1 acts as a modular protein that stabilizes NbBIR2 by competing with NbSNIPER2a or NbSNIPER2b for association with NbBIR2. Similar to NbBAK1, NbBIR2 positively regulates pattern-triggered immunity and resistance to bacterial and oomycete pathogens in N. benthamiana, whereas NbSNIPER2a and NbSNIPER2b have the opposite effect. Together, these results reveal a feedback regulatory mechanism employed by plants to tailor pattern-triggered immune signaling.

Novel Strategy to Combat the Procoagulant Phenotype in Heparin-Induced Thrombocytopenia Using 12-LOX Inhibition.

Heparin-induced thrombocytopenia (HIT) is a major concern for all individuals that undergo cardiac bypass surgeries or require prolonged heparin exposure. HIT is a life- and limb-threatening adverse drug reaction with an immune response following the formation of ultra-large immune complexes that drive platelet activation through the receptor FcγRIIA. Thrombotic events remain high following the standard of care treatment with anticoagulants, while increasing risk of bleeding complications. This study sought to investigate a novel approach to treatment of HIT. Recent reports demonstrate increased procoagulant activity in HIT; however, these reports required analysis ex vivo, and relevance in vivo remains unclear. Using human and mouse model systems, we investigated the cooperativity of PARs (protease-activated receptors) and FcγRIIA in HIT. We challenged humanized FcγRIIA transgenic mice with or without endogenous mouse Par4 (denoted as IIA-Par4+/+ or IIA-Par4-/-, respectively) with a well-established model IgG immune complex (anti [α]-CD9). Furthermore, we assessed the procoagulant phenotype and efficacy to treat HIT utilizing inhibitor of 12-LOX (12[S]-lipoxygenase), VLX-1005, previously reported to decrease platelet activation downstream of FcγRIIA and PAR4, using the triple allele HIT mouse model. IIA-Par4+/+ mice given αCD9 were severely thrombocytopenic, with extensive platelet-fibrin deposition in the lung. In contrast, IIA-Par4-/- mice had negligible thrombocytopenia or pulmonary platelet-fibrin thrombi. We observed that pharmacological inhibition of 12-LOX resulted in a significant reduction in both platelet procoagulant phenotype ex vivo, and thrombocytopenia and thrombosis in our humanized mouse model of HIT in vivo. These data demonstrate for the first time the need for dual platelet receptor (PAR and FcγRIIA) stimulation for fibrin formation in HIT in vivo. These results extend our understanding of HIT pathophysiology and provide a scientific rationale for targeting the procoagulant phenotype as a possible therapeutic strategy in HIT.

Enhanced binding at fever temperatures of HER2 in complex with trastuzumab and pertuzumab.

Aim: Fever follows the administration of trastuzumab and pertuzumab used in HER2-relevant immunotherapy, but is often eliminated in clinical practice. This work explores the role of temperature (37-39°C) in the formation of immune complexes between HER2 with either trastuzumab or pertuzumab or with both antibodies. Materials & methods: Using molecular dynamics simulations and free energy calculations, the binding between HER2 and these immunotherapeutic monoclonal antibodies was investigated at different temperatures. Results: Trastuzumab and pertuzumab present the highest bindingfreeenergy to HER2 at febrile temperatures (39°C), or when HER2 is in complex with both antibodies. Conclusion: Performing molecular dynamics simulations under fever temperatures may be important for delineating their role in enhancing the binding affinity of mature antibodies used in immunotherapy.

#4458 SINGLE CELL RNA SEQUENCING OF CIRCULATING LEUKOCYTES FROM IGA NEPHROPATHY PATIENTS IDENTIFIES AN IMPORTANT ROLE FOR INNATE IMMUNE CELLS

Abstract Background and Aims IgA-nephropathy (IgAN) is a slowly progressive disease characterized by mesangial deposition of IgA1 that causes kidney injury and can eventually lead to end stage renal disease. A key factor underlying the development of IgAN is the presence of galactose-deficient IgA1 (Gd-IgA1) in the circulation. The formation of IgA and IgG antibodies against Gd-IgA1 results in the formation of immune complexes which deposit in the kidney mesangium. Production of Gd-IgA1 hints to defects in B cell regulation. However, the exact immunological dysregulation leading to the formation of Gd-IgA1, and the subsequent antibody formation against it, remains elusive. Here we use single cell RNA sequencing (scRNA-seq) of circulating leukocytes for in depth analysis of immune cell populations in patients with active IgAN to gain a better understanding of the pathogenesis of this disease. Method Peripheral blood mononuclear cells were isolated for scRNA-seq from the blood of four patients with active IgAN, and four controls, matched for age and estimated glomerular filtration rate (eGFR). Cell hashing (Biolegend) of individual samples was performed before pooling samples for scRNA-seq. scRNA expression libraries were performed using the chromium X machine (10xgenomics) according to 10XGenomics single cell 5’ V2 user guide. Libraries were sequenced on an Illumina Nextseq500 and reads were mapped to GrCh38 using CellRanger. Differential gene expression analysis was performed for each immune cell type (FDR < 0.05). Reactome pathway enrichment analysis was performed on the differentially expressed genes. Results Identification of immune cell subtypes in IgAN patients and matched controls did not show a significant difference in cell numbers or ratios. B cells showed only minor changes in gene expression between IgAN patients and controls, and T cell populations showed changes in a limited set of genes, whereas interestingly, most differentially expressed genes were found in monocytes and NK cells (Figure 1A). Analysis of the differentially expressed genes showed enrichment for genes in interferon signaling pathways, mostly in monocytes and NK cells (Figure 1B). Gene enrichment analysis showed differential expression of type I IFN genes and antigen presentation genes, also mainly in monocytes and NK cells. Conclusion Our data shows the specific endotype of circulating immune cells from patients with active IgAN. While gene expression changes in T and B cells are discrete, changes in monocytes and NK cells appear especially prominent. In these cell types, we observed an important role for interferon signaling. Further analysis of scRNA-seq data in a larger patient cohort will follow, as well as analysis of T and B cell receptor clones in IgAN patients, with which we aim to gain new insights into immune dysregulation in IgAN.

#4333 A PHASE 3, RANDOMIZED, DOUBLE-BLIND, PLACEBO-CONTROLLED STUDY OF BION-1301 IN ADULTS WITH IGA NEPHROPATHY

Abstract Background and Aims IgA nephropathy (IgAN) is the leading cause of primary glomerulonephritis and has limited treatment options, especially for high-risk patients. Approximately 30-45% of IgAN patients progress to end-stage kidney disease over a period of 20-25 years and proteinuria is the strongest predictor of disease progression [1]. BION-1301 is a novel, humanized monoclonal antibody that blocks a proliferation-inducing ligand (APRIL), a cytokine that is elevated in patients with IgAN. APRIL promotes the production of pathogenic galactose-deficient IgA1 (Gd-IgA1), resulting in immune complex formation and subsequent glomerular deposition, leading to inflammation and kidney injury. Blocking APRIL with BION-1301 is a potential disease-modifying approach to treating IgAN. Interim results from a Phase 1/2 trial of BION-1301 in patients with IgAN (NCT03945318) demonstrate rapid and durable reductions in Gd-IgA1 and sustained, clinically meaningful reductions in proteinuria with an acceptable safety profile [2]. Methods CHK02-02 is a Phase 3, randomized, double-blind, placebo-controlled study to evaluate the effect of BION-1301 in adults with primary IgAN at risk of progressive kidney function loss. Approximately 272 patients will be enrolled across North America, South America, Europe and Asia-Pacific. Key eligibility criteria include biopsy-proven IgAN within the past 10 years (not due to secondary causes), eGFR ≥ 30 ml/min/1.73 m2 (CKD-EPI) and total urine protein ≥ 1.0 g/day at screening. Patients must be stable on a maximally tolerated dose of angiotensin-converting enzyme inhibitor (ACEi)/angiotensin II receptor blocker (ARB) for at least 12 weeks prior to screening or intolerant to ACEi/ARB. Patients may also be on a stable dose of SGLT2i, mineralocorticoid receptor antagonists, and/or endothelin receptor antagonists for at least 12 weeks prior to screening. The study will be comprised of a screening period (6 weeks), a double-blind treatment period (104 weeks), and a safety follow-up period (24 weeks). Patients will be randomized 1:1 to receive subcutaneous 600 mg BION-1301 Q2W or placebo for 104 weeks. Randomization will be stratified by region (Asia vs. Rest of World), baseline proteinuria (≥ 2 g/day vs. < 2 g/day) and eGFR (≤ 45 ml/min/1.73 m2 vs. > 45 ml/min/1.73 m2). An additional ∼20 patients with eGFR 20 to < 30 mL/min/1.73 m2 will be enrolled into an exploratory cohort not included in the primary or secondary analyses. Results The primary endpoint is change in proteinuria (UPCR from a 24-hour urine collection) from baseline to week 36. The key secondary endpoint is change in eGFR from baseline to week 104. Additional secondary endpoints will evaluate the effect of BION-1301 vs. placebo on composite clinical outcomes including patients experiencing at least one of the following: 30% or 40% reduction in eGFR, eGFR < 15 mL/min/1.73 m2, dialysis, kidney transplantation or all-cause mortality. Safety endpoints include type, incidence, severity, and relatedness of adverse events (AEs) and serious AEs. Exploratory endpoints include impact of BION-1301 on disease biomarkers and health-related quality of life as well as analysis of BION-1301 pharmacokinetics and immunogenicity. Conclusion BION-1301 provides a potentially disease-modifying approach for the treatment of IgAN by directly targeting the disease pathogenesis. Interim results of the Phase 1/2 open-label trial demonstrated proof-of-concept for BION-1301 to reduce pathogenic Gd-IgA1 and provide sustained, clinically meaningful reductions in proteinuria while supporting SC dosing at 600 mg Q2W [2]. The Phase 3 trial will evaluate the effect of BION-1301 vs. placebo on proteinuria, eGFR and composite clinical endpoints and key safety measures in adult patients with IgAN at risk of progressive kidney function loss.

Potassium cobalt hexacyanoferrate as a peroxidase mimic for electrochemical immunosensing of Lactobacillus rhamnosus GG

In this paper, the potassium cobalt hexacyanoferrate (II), K2CoFe(CN)6, with peroxidase-like activity was used for the fabrication of a novel label-free Lactobacillus rhamnosus GG (LGG) electrochemical immunosensor. The K2CoFe(CN)6 nanocubes were made by a simple hydrothermal method and followed by low-temperature calcination. In addition to structural characterization, the peroxidase-mimicking catalytic property of the material was confirmed by a chromogenic reaction. It is known that H2O2 can oxidize electroactive thionine molecules under the catalysis of horseradish peroxidase (HRP). In this nanozyme-based electrochemical immunoassay, due to the steric hindrance, the formation of immune-complex of LGG and LGG antibody on the modified GCE inhibits the catalytic activity of the peroxidase mimics of K2CoFe(CN)6 and thus reduced the current signal. Therefore, the developed electrochemical immunosensor achieved quantitative detection of LGG. Under optimal conditions, the linear range of the sensor was obtained from 101 to 106 CFU mL−1 with a minimum detection limit (LOD) of 12 CFU mL−1. Furthermore, the immunosensor was successfully applied in the quantitative detection of LGG in dairy product samples with recoveries ranging from 93.2% to 106.8%. This protocol presents a novel immunoassay method, which provides an alternative implementation pathway for the quantitative detection of microorganisms.

Ras family signaling pathway in immunopathogenesis of inflammatory rheumatic diseases.

The Ras (rat sarcoma virus) is a GTP-binding protein that is considered one of the important members of the Ras-GTPase superfamily. The Ras involves several pathways in the cell that include proliferation, migration, survival, differentiation, and fibrosis. Abnormalities in the expression level and activation of the Ras family signaling pathway and its downstream kinases such as Raf/MEK/ERK1-2 contribute to the pathogenic mechanisms of rheumatic diseases including immune system dysregulation, inflammation, and fibrosis in systemic sclerosis (SSc); destruction and inflammation of synovial tissue in rheumatoid arthritis (RA); and autoantibody production and immune complexes formation in systemic lupus erythematosus (SLE); and enhance osteoblast differentiation and ossification during skeletal formation in ankylosing spondylitis (AS). In this review, the basic biology, signaling of Ras, and abnormalities in this pathway in rheumatic diseases including SSc, RA, AS, and SLE will be discussed.

Label-Free Electrochemical Biosensing Architecture Based on a Protein-Decorated Pt@BSA Nanocomposite for Sensitive Immunoassay of Pathogenic Escherichia coli O157:H7

Electrochemical immunosensors equipped with high specificity, superior analysis sensitivity, low cost, and convenient operations have been widely applied in the food industry, clinical medicine, biomonitor technique, and environmental protection. Because of the pressing requirements for detecting the pathogen Escherichia coli (E. coli) O157:H7 in various practical systems, an efficient and specific immunosensor was delicately designed based on the Pt@BSA nanocomposite with the surface conjugation of affinity-purified antibody molecules to achieve immune recognition. Amplified signal response toward probe molecules and favorable biocompatibility for bacteria attachment could be obtained at the assembled Pt@BSA biosensing interface. With the formation of immune complexes, specific binding of E. coli O157:H7 cells under a certain concentration range onto the antibody-immobilized interface resulted in electrochemical signal responses at different levels, which could be directly measured by electrochemical impedance spectroscopy (EIS) and differential pulse voltammetry (DPV). Herein, E. coli O157:H7 cells were quantitatively monitored by the DPV technique in a linear concentration range from 2.5 × 101 to 5.0 × 108 cfu mL–1 with a low detection limit of 9 cfu mL–1. This fabricated bacteria immunosensor indicated high specificity for E. coli determination, favorable selectivity among the interfering bacteria, excellent storage stability, satisfactory preparation reproducibility with a relative standard deviation (RSD) of 3.5%, and acceptable detection accuracy in practical food samples, providing a promising approach for ultrasensitive monitoring of hazardous substances and disease markers at an extremely low level in clinical diagnosis.

Improved radioimmunodetection of carcinomas with a re-injection of monoclonal antibodies after formation of anti-mouse antibodies.

Scintigraphic imaging was satisfactory in animal experiments, i.e., in the radioimmunodetection with 125J anti-tissue polypeptide antigen monoclonal antibodies and implanted HELA cell carcinomas. Unlabeled anti-mouse antibodies (AMAB), in a surplus of 40:1, 200:1 and 4000:1 compared to the radioactive antibody, were administered five days after administering the 125I anti-TPA antibody (RAAB). In immunoscintigraphies, radioactivity accumulated in the liver immediately after administering the secondary antibody, and the tumor's imaging worsened. It can be expected that imunoscintigraphic imaging might improve when radioimmunodetection is re-performed after the formation of human anti-mouse antibodies (AMAB) and when the ratio of the primary to the secondary antibody is nearly equivalent because, in this ratio, the formation of immune complexes might be accelerated. It is possible to measure the quantity of formed anti-mouse antibodies (AMAB) with immunography measurements. A second administration of diagnostic or therapeutic monoclonal antibodies might lead to the formation of immune complexes if the quantities of the monoclonal antibodies and the anti-mouse antibodies have an equivalent ratio. A second performance of the radioimmunodetection four to eight weeks after the first radioimmunodetection can achieve better tumor imaging because human anti-mouse antibodies (AMAB) can be formed. Immune complexes of the radioactive antibody and the human anti-mouse antibody (AMAB) can be formed to concentrate radioactivity in the tumor.

Systemic lupus erythematosus with diffuse alveolar hemorrhage.

Systemic lupus erythematosus with diffuse alveolar hemorrhage.