IgG Ab Research Articles

Adoptive Cord Blood T Regulatory Cell Therapy Leads to Resolution of Inflammation and Decreased Proteinuria in Lupus Nephritis

Background. Systemic lupus erythematosus (SLE) is associated with widespread inflammation with multi-organ involvement where renal failure is the most dreaded and fatal complication. Adoptive therapy with cord blood (CB) derived T regulatory cells has been shown to improve outcomes in disease driven by inflammation including graft versus host disease (GvHD), inflammatory bone marrow disorder, COVID-19 induced acute respiratory distress syndrome. We hypothesize that CB Treg therapy can treat lupus nephritis. Methods. The suppressive abilities of CB Tregs expressing CD4+CD25highCD127lowFoxP3+ were assayed by human cytokines assay kits (IL-10, IFN-γ, IP-10, TNF-α, IL-6, and IL-17A) in the cell culture supernatants. For examining the efficacy of CB Tregs in vivo, SLE xenograft model was created with female Rag2-/-γc-/- mice transplanted with 3x106 human SLE-peripheral blood mononuclear cells (PBMCs) by intravenous injection on day 0. The mice were allowed to develop disease and on day 30 post-transplant, they were divided into 2 groups: i) control and ii) treatment. 1x107 ex vivo-expanded, cryopreserved, allogeneic, non-HLA matchedCB Tregs were injected into SLE xenografts intravenously once per week for 4 weeks through the tail vein. Serial blood draws were performed for the phenotypic analysis, cytokine assay and anti-double stranded (ds)DNA IgG antibody analysis. Serial examination of the urine samples was performed for creatinine and albumin quantification. Histopathologic examination of the harvested organs was performed at the time of planned euthanasia at 13 weeks. Results. Co-culture of CB Tregs with the pathogenic SLE-PBMCs decreased the secretion of inflammatory cytokines including IFN-γ, IP-10, TNF-α, IL-6, and IL-17A (Figure A) with a reciprocal increase in the secretion of the anti-inflammatory IL-10 cytokine (Figure B). Adoptive therapy with CB Treg cells led to a significant decrease in circulating CD8+ effector T cells and an increase in CD4+ helper T cells (Figure C). CB Treg recipients showed preserved weight gain (Figure D), lower GvHD score (Figure E) and improved overall survival (Figure F). A significant decrease in proteinuria at 9 weeks post-transplant (Figure G) correlated with a decrease in anti-dsDNA IgG Ab levels (Figure H) and soluble CD40 ligand levels (Figure I). Histopathological results from two index cases from each arm (Figure J) demonstrated that CB Treg recipients show reduced T-cells (CD3+) (Figure K) and B-cells (CD20+) (Figure L) in the kidneys, as well as a decrease in the lymphoid infiltration into glomeruli and renal parenchyma as compared to the control arm (Figure M). Conclusion. We are the first to demonstrate the benefit of allogeneic CB Treg cell therapy for treatment of lupus nephritis. We propose to examine such a strategy in the clinical setting. Figure Disclosures Nishimoto: Janssen Pharmaceutical K.K.:: Research Funding; Bayer Yakuhin, Ltd:: Research Funding. Sadeghi:Cellenkos Inc.: Current Employment. Parmar:Cellenkos Inc.: Current equity holder in private company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding.

IgG Immune Complexes Break Immune Tolerance of Human Microglia.

Microglia are phagocytic cells involved in homeostasis of the brain and are key players in the pathogenesis of multiple sclerosis (MS). A hallmark of MS diagnosis is the presence of IgG Abs, which appear as oligoclonal bands in the cerebrospinal fluid. In this study, we demonstrate that myelin obtained post mortem from 8 out of 11 MS brain donors is bound by IgG Abs. Importantly, we show that IgG immune complexes strongly potentiate activation of primary human microglia by breaking their tolerance for microbial stimuli, such as LPS and Poly I:C, resulting in increased production of key proinflammatory cytokines, such as TNF and IL-1β. We identified FcγRI and FcγRIIa as the two main responsible IgG receptors for the breaking of immune tolerance of microglia. Combined, these data indicate that IgG immune complexes potentiate inflammation by human microglia, which may play an important role in MS-associated inflammation and the formation of demyelinating lesions.

Expansion of SARS-CoV-2-Specific Antibody-Secreting Cells and Generation of Neutralizing Antibodies in Hospitalized COVID-19 Patients.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), emerged in late 2019 and has since become a global pandemic. Pathogen-specific Abs are typically a major predictor of protective immunity, yet human B cell and Ab responses during COVID-19 are not fully understood. In this study, we analyzed Ab-secreting cell and Ab responses in 20 hospitalized COVID-19 patients. The patients exhibited typical symptoms of COVID-19 and presented with reduced lymphocyte numbers and increased T cell and B cell activation. Importantly, we detected an expansion of SARS-CoV-2 nucleocapsid protein-specific Ab-secreting cells in all 20 COVID-19 patients using a multicolor FluoroSpot Assay. Out of the 20 patients, 16 had developed SARS-CoV-2-neutralizing Abs by the time of inclusion in the study. SARS-CoV-2-specific IgA, IgG, and IgM Ab levels positively correlated with SARS-CoV-2-neutralizing Ab titers, suggesting that SARS-CoV-2-specific Ab levels may reflect the titers of neutralizing Abs in COVID-19 patients during the acute phase of infection. Last, we showed that IL-6 and C-reactive protein serum concentrations were higher in patients who were hospitalized for longer, supporting the recent observations that IL-6 and C-reactive protein could be used as markers for COVID-19 severity. Altogether, this study constitutes a detailed description of clinical and immunological parameters in 20 COVID-19 patients, with a focus on B cell and Ab responses, and describes tools to study immune responses to SARS-CoV-2 infection and vaccination.

Development of a dendrimer PAMAM‑based gold biochip for rapid and sensitive detection of endogenous IFN‑γ and anti‑IFN‑γ IgG in patients with hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is a rare but severe disease characterized by immune hyperactivation and cytokine storm. Given the high mortality rate of HLH, there is a need for more effective diagnostic tools and treatments. The present study developed a dendrimer-based protein biochip for rapid, sensitive and simultaneous detection of serum interferon (IFN)-γ and endogenous anti-IFN-γ antibody (Ab) in patients with HLH. A gold biochip was modified with 1, 4-phenylene diisothiocyanate (PDITC), polyamidoamine (PAMAM) or PDITC-activated PAMAM. The optimal immobilization concentration for Ab capture and the reaction concentration for detecting Ab on the PDITC-activated PAMAM-modified biochip were 6.25 and 3.12 µg/ml, respectively; the limit of detection of IFN-γ protein was 50 pg/ml. The efficiency of the protein-probed biochip in detecting IFN-γ and anti-IFN-γ Ab in serum samples from 77 patients with HLH was evaluated; the positive rates for IFN-γ and anti-IFN-γ IgG Ab were 63.6% (49/77) and 61.0% (47/77), respectively. The present results demonstrated that the PDITC-activated PAMAM-modified biochip might be a sensitive tool for the specific detection of IFN-γ and anti-IFN-γ Ab in serum, and might have clinical applicability for the diagnosis of HLH.

Mapping Human Monoclonal IgE Epitopes on the Major Dust Mite Allergen Der p 2

IgE Abs drive the symptoms of allergic disease upon cross-linking allergens on mast cells or basophils. If the IgE binding sites on the allergens could be identified, it may be useful for creating new forms of immunotherapy. However, direct knowledge of the human IgE (hIgE) epitopes is limited because of the very low frequency of IgE-producing B cells in blood. A new hybridoma technology using human B cells from house dust mite-allergic patients was used to identify four Der p 2-specific hIgE mAbs. Their relative binding sites were assessed and compared by immunoassays with three previously studied murine IgG mAbs. Immunoassays showed that the recognition of Der p 2 by the first three hIgE was inhibited by a single murine IgG, but the fourth hIgE recognized a different epitope from all the other mAbs. The functional ability of the hIgE that bind different epitopes to cross-link Der p 2 was demonstrated in a mouse model of passive systemic anaphylaxis. Nuclear magnetic resonance analyses of Der p 2 in complex with IgG and IgE Abs were used to identify specific residues in the epitopes. To our knowledge, the combination of immunoassays to distinguish overlapping epitopes and nuclear magnetic resonance analyses to identify specific residues involved in Ab binding provided the first epitope mapping of hIgE mAbs to an allergen. The technologies developed in this study will be useful in high-resolution mapping of human epitopes on other Ags and the design of improved therapeutics.

Influenza Vaccine-Induced CD4 Effectors Require Antigen Recognition at an Effector Checkpoint to Generate CD4 Lung Memory and Antibody Production.

Previously, we discovered that influenza-generated CD4 effectors must recognize cognate Ag at a defined effector checkpoint to become memory cells. Ag recognition was also required for efficient protection against lethal influenza infection. To extend these findings, we investigated if vaccine-generated effectors would have the same requirement. We compared live infection with influenza to an inactivated whole influenza vaccine. Live infection provided strong, long-lasting Ag presentation that persisted through the effector phase. It stimulated effector generation, long-lived CD4 memory generation, and robust generation of Ab-producing B cells. In contrast, immunization with an inactivated virus vaccine, even when enhanced by additional Ag-pulsed APC, presented Ag for 3 d or less and generated few CD4 memory cells or long-lived Ab-producing B cells. To test if checkpoint Ag addition would enhance this vaccine response, we immunized mice with inactivated vaccine and injected Ag-pulsed activated APC at the predicted effector checkpoint to provide Ag presentation to the effector CD4 T cells. This enhanced generation of CD4 memory, especially tissue-resident memory in the lung, long-lived bone marrow Ab-secreting cells, and influenza-specific IgG Ab. All responses increased as we increased the density of peptide Ag on the APC to high levels. This suggests that CD4 effectors induced by inactivated vaccine require high levels of cognate Ag recognition at the effector checkpoint to most efficiently become memory cells. Thus, we suggest that nonlive vaccines will need to provide high levels of Ag recognition throughout the effector checkpoint to optimize CD4 memory generation.

S2281 CMV Colitis Mimicking as IBD Flare-Up in Immunocompetent Patient

INTRODUCTION: Although almost 70 % of human beings are likely to be infected with CMV, the majority, being immunocompetent, are unlikely to manifest. Cytomegalovirus (CMV) colitis rarely occurs in immunocompetent patients. Usually, CMV colitis occurs in immunocompromised individuals but there have been reports in immunocompetent individuals as well. Here, we present a case of a 64 year old female initially presenting as IBD flare-up but later getting diagnosed with CMV colitis. CASE DESCRIPTION/METHODS: A 64-year-old woman, with a past medical history of HTN, type II diabetes, and GERD, IBD - Crohn's type on mesalamine, with recently done EGD and colonoscopy, presented to the hospital for abdominal pain and bloody stools for one month. When she came to the hospital, her WBCs were elevated, and she also had elevated lactate, ferritin but the rest of her labs were unremarkable. Subsequently, a CAT scan of Abdomen and Pelvis showed non-specific colitis. Gastroenterology service recommended starting the patient on Solu-Medrol for a possible Crohn’s Disease Flare Up. Other stool studies were negative. Over the course of her hospitalization, the patient underwent colonoscopy which showed severe inflammation and was also started on Infliximab due to unresponsiveness to treatment. Strongyloides serologies were checked and came back negative as well. The pathology results from colonoscopy came back and the findings were consistent with Cytomegalovirus (CMV) colitis. CMV IgG Ab checked came back positive and viral load came back at 7400 U/L. It was decided to treat the patient with Ganciclovir. The patient reported improvement and was able to tolerate her diet. She was subsequently discharged on valganciclovir. On subsequent follow up reported the resolution of her signs and symptoms. DISCUSSION: CMV colitis occurs in immunocompromised individuals but there have been reports in immunocompetent individuals as well. The mechanism is unclear but it has been hypothesized that CMV superinfects already inflamed mucosa. It is also stated that MHC proteins are upregulated leading to an autoimmune response. The treatment of choice for CMV colitis remains ganciclovir. The association between cytomegalovirus infection and inflammatory bowel disease challenges both the clinicians and the pathologists to establish the correct diagnosis and to prescribe the most appropriate form of therapy, therefore a high suspicion should be there for CMV colitis in Immunocompetent IBD patients as well who are not responding to treatment.Figure 1.: CT Scan of Abdomen and Pelvis showing prominent diffuse colonic wall thickening.

Molecular signatures of anthroponotic cutaneous leishmaniasis in the lesions of patients infected with Leishmania tropica

Anthroponotic cutaneous leishmaniasis (CL) caused by Leishmania tropica (L. tropica) represents a public health challenge in several resource poor settings. We herein employed a systems analysis approach to study molecular signatures of CL caused by L. tropica in the skin lesions of ulcerative CL (UCL) and non-ulcerative CL (NUCL) patients. Results from RNA-seq analysis determined shared and unique functional transcriptional pathways in the lesions of the UCL and NUCL patients. Several transcriptional pathways involved in inflammatory response were positively enriched in the CL lesions. A multiplexed inflammatory protein analysis showed differential profiles of inflammatory cytokines and chemokines in the UCL and NUCL lesions. Transcriptional pathways for Fcγ receptor dependent phagocytosis were among shared enriched pathways. Using L. tropica specific antibody (Ab)-mediated phagocytosis assays, we could substantiate Ab-dependent cellular phagocytosis (ADCP) and Ab-dependent neutrophil phagocytosis (ADNP) activities in the lesions of the UCL and NUCL patients, which correlated with L. tropica specific IgG Abs. Interestingly, a negative correlation was observed between parasite load and L. tropica specific IgG/ADCP/ADNP in the skin lesions of CL patients. These results enhance our understanding of human skin response to CL caused by L. tropica.

A Case of Suspected Covid 19 Related Cardiomyopathy

BackgroundThe novel SARS-CoV-2 virus causing COVID-19 has been associated with diverse cardiovascular pathology. We present a case of cardiomyopathy due to possible COVID-19 resulting in cardiogenic shock.CaseA 54 year-old male presented to the hospital with 4 weeks of progressive dyspnea, leg swelling, and weight gain. His symptoms began 4 weeks after experiencing influenza-like symptoms after a trip to China during the height of their COVID-19 outbreak. He was admitted to the COVID unit in cardiogenic shock and was later intubated for acute hypoxic respiratory failure. Laboratory data demonstrated acute kidney injury, elevated transaminases, lactic acidosis, elevated pro-BNP N-Terminal to 3932pg/mL, and high sensitivity troponin to 72ng/L. Transthoracic echocardiogram showed severe biventricular failure with a LVEF of 10% and a LVIDd 5.2cm. SARS-CoV-2 RNA was negative twice, but SARS-CoV-2 IgG AB and SARS-CoV-2 IgA AB were positive. Urgent right and left heart catheterization was performed demonstrating non-obstructive coronary artery disease and hemodynamics consistent with cardiogenic shock. While supported with an intra-aortic balloon pump (IABP) and norepinephrine, he had a fick cardiac output 3.1 L/min, fick cardiac index 1.6 L/min/m2, pulmonary capillary wedge pressure 37mmHg, right atrial pressure 25mmHg, and pulmonary arterial pressures 65/40mmHg. Given persistent cardiogenic shock on IABP and inotropes, he was later transitioned to Veno-Arterial Extracorporeal Membrane Oxygenation (VA ECMO) and an Impella CP for left ventricular unloading. He continued to have persistent INTERMACS I shock and underwent successful implantation of a HeartMate 3 LVAD with percutaneous temporary right ventricular assist device (RVAD). Pathology of the left ventricular apical core demonstrates polyclonal endocardial infiltration of B-Cells, CD4 and CD8 positive T-Cells, eosinophils, macrophages, and plump reactive endothelial cells (Figure 1). He is currently recovering in the ICU off of vasoactive support with subsequent removal of percutaneous RVAD.ConclusionThis is a suspected case of COVID-19 associated cardiomyopathy presenting as new on-set heart failure with reduced ejection fraction complicated by cardiogenic shock. There is still much to learn about the cardiac manifestations of COVID-19 and further studies are needed to determine appropriate diagnostics and management of such cases.

Diversity Outbred Mice Reveal the Quantitative Trait Locus and Regulatory Cells of HER2 Immunity.

The genetic basis and mechanisms of disparate antitumor immune response was investigated in Diversity Outbred (DO) F1 mice that express human HER2. DO mouse stock samples nearly the entire genetic repertoire of the species. We crossed DO mice with syngeneic HER2 transgenic mice to study the genetics of an anti-self HER2 response in a healthy outbred population. Anti-HER2 IgG was induced by Ad/E2TM or naked pE2TM, both encoding HER2 extracellular and transmembrane domains. The response of DO F1 HER2 transgenic mice was remarkably variable. Still, immune sera inhibited HER2+ SKBR3 cell survival in a dose-dependent fashion. Using DO quantitative trait locus (QTL) analysis, we mapped the QTL that influences both total IgG and IgG2(a/b/c) Ab response to either Ad/E2TM or pE2TM. QTL from these four datasets identified a region in chromosome 17 that was responsible for regulating the response. A/J and NOD segments of genes in this region drove elevated HER2 Ig levels. This region is rich in MHC-IB genes, several of which interact with inhibitory receptors of NK cells. (B6xA/J)F1 and (B6xNOD)F1 HER2 transgenic mice received Ad/E2TM after NK cell depletion, and they produced less HER2 IgG, demonstrating positive regulatory function of NK cells. Depletion of regulatory T cells enhanced response. Using DO QTL analysis, we show that MHC-IB reactive NK cells exert positive influence on the immunity, countering negative regulation by regulatory T cells. This new, to our knowledge, DO F1 platform is a powerful tool for revealing novel immune regulatory mechanisms and for testing new interventional strategies.

Toward defining the immunogenicity of HLA epitopes: Impact of HLA class I eplets on antibody formation during pregnancy.

Eplets are functional units of structural epitopes on donor HLA, potentially recognized by complementarity-determining regions of the paratope of the recipients' B-cell receptors or antibodies (Ab). Their individual immunogenicity is poorly described, yet this feature would be of clinical importance for pretransplant risk assessment. The aim of this study was to determine the relative immunogenicity of HLA class I eplets in the pregnancy setting, where mismatched eplets are present on paternal HLA antigens of the unborn child. One hundred fifty-nine predominantly Caucasian mothers giving birth at the University Hospital Basel and their first newborns were HLA-typed at high-resolution by next-generation sequencing (NGS) (NGSgo Workflow and NGSengine from GenDx; sequencing with a Miseq from Illumina) and eplets were determined using HLAMatchmaker. HLA class I specific IgG Ab was assessed in maternal sera drawn immediately after full-term delivery, by OneLambda LABScreen single antigen ibeads. The Ab profile was subsequently evaluated for eplet-associated patterns. All 72 currently Ab-verified HLA class I eplets were examined for their immunogenicity according to the frequency of child-specific HLA Ab (CSA) directed against their structures. Four hundred twelve of 477 (86.4%) paternal HLA-A, -B or -C alleles were mismatched. CSA were present in 46 mothers (28.9%), directed against 80 (19.4%) of these mismatches. The 10 most immunogenic eplets were 62GK, 145KHA, 144TKH, 62GE, 107W, 80I, 82LR, 41T, 127K, 45KE with immunogenicity rates between 45.8% and 27.3%. This pregnancy study also identified five non-reactive eplets: 62RR, 76ESN, 80TLR, 156DA, 163RW. Based on our results, immunogenic hot and cold spots on the surface of HLA class I molecules were localized and visualized on 3D models. This study strengthens the presumption that different eplets represent different immunogenic potentials. Validation of these results in the clinical transplant setting is an essential next step in identifying those eplets representing a particularly high-risk potential.

Omega-3 fatty acid intake suppresses induction of diverse autoantibody repertoire by crystalline silica in lupus-prone mice

Inhalation of crystalline silica (cSiO2) in the workplace is etiologically linked to lupus and other autoimmune diseases. Exposing lupus-prone NZBWF1 mice to respirable cSiO2 unleashes a vicious cycle of inflammation and cell death in the lung that triggers interferon-regulated gene expression, ectopic lymphoid structure (ELS) development, elevation of local and systemic autoantibodies (AAbs), and glomerulonephritis. However, cSiO2-induced inflammation and onset of autoimmunity can be prevented by inclusion of the ω-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) into the diet of these mice. Since cSiO2 both causes cell death and interferes with efferocytosis, secondary necrosis of residual cell corpses might provide a rich and varied autoantigen (AAg) source in the lung. While it is known that the particle induces anti-nuclear and anti-dsDNA AAbs in NZBWF1 mice, the full extent of the cSiO2-induced AAb response relative to specificity and isotype is not yet understood. The purpose of this study was to test the hypotheses that cSiO2 exposure induces a wide spectrum of AAbs in the pulmonary and systemic compartments, and that dietary DHA intervention prevents these changes. Archived tissue fluid samples were obtained from a prior study in which NZBWF1 mice were fed purified isocaloric diets containing no DHA (control) or DHA corresponding calorically to human doses of 2 and 5 g/day. Mice were intranasally instilled with 1 mg cSiO2 or saline vehicle weekly for 4 weeks, then groups euthanized 1, 5, 9, or 13 weeks post-instillation (PI) of the last cSiO2 dose. Bronchoalveolar lavage fluid (BALF) and plasma from each time point were subjected to AAb profiling using a microarray containing 122 AAgs. cSiO2 triggered robust IgG and IgM AAb responses against lupus-associated AAgs, including DNA, histones, ribonucleoprotein, Smith antigen, Ro/SSA, La/SSB, and complement as early as 1 week PI in BALF and 5 weeks PI in plasma, peaking at 9 and 13 weeks PI, respectively. Importantly, cSiO2 also induced AAbs to AAgs associated with rheumatoid arthritis (collagen II, fibrinogen IV, fibrinogen S, fibronectin, and vimentin), Sjögren’s syndrome (α-fodrin), systemic sclerosis (topoisomerase I), vasculitis (MPO and PR3), myositis (Mi-2, TIF1-γ, MDA5), autoimmune hepatitis (LC-1), and celiac disease (TTG). cSiO2 elicited comparable but more modest IgA AAb responses in BALF and plasma. cSiO2-induced AAb production was strongly associated with time dependent inflammatory/autoimmune gene expression, ELS development, and glomerulonephritis. AAb responses were dose-dependently suppressed by DHA supplementation and negatively correlated with the ω-3 index, an erythrocyte biomarker of ω-3 content in tissue phospholipids. Taken together, these findings suggest that cSiO2 exposure elicits a diverse multi-isotype repertoire of AAbs, many of which have been reported in individuals with lupus and other autoimmune diseases. Furthermore, induction of this broad AAb spectrum could be impeded by increasing ω-3 tissue content via dietary DHA supplementation.

Clinical performance of four immunoassays for antibodies to SARS-CoV-2, including a prospective analysis for the diagnosis of COVID-19 in a real-life routine care setting.

The aim of the present study was to evaluate the clinical performance of four SARS-CoV-2 immunoassays and their contribution in routine care for the diagnosis of COVID-19, in order to benefit of robust data before their extensive use. The clinical performance of Euroimmun ELISA SARS-CoV-2 IgG, Abbott SARS-CoV-2 IgG, Wantai SARS-CoV-2 Ab ELISA, and DiaPro COVID-19 IgG confirmation were evaluated in the context of both a retrospective and a prospective analysis of COVID-19 patients. The retrospective analysis included plasma samples from 63 COVID-19 patients and 89 control (pre-pandemic) patients. The prospective study included 203 patients who tested either negative (n = 181) or positive (n = 22) by RT-PCR before serology sampling. The specificity was 92.1 %, 98.9 %, 100 % and 98.9 % and the sensitivity 14 days after onset of symptoms was 95.6 %, 95.6 %, 97.8 % and 95.6 % for Euroimmun IgG, Abbott IgG, Wantai Ab, and DiaPro IgG confirmation SARS-CoV-2 immunoassays, respectively. The low specificity of Euroimmun IgG (for ratio <5) was not confirmed in routine care setting (98.5 % negative agreement). Serology was complementary to RT-PCR in routine care and lead to identification of false positive (Ct>38, <2 targets detected) and false negative RT-PCR results (>1 month post onset of symptoms). Serology was complementary to RT-PCR for the diagnosis of COVID-19 at least 14 days after onset of symptoms. First line serology testing can be performed with Wantai Ab or Abbott IgG assays, while DiaPro IgG confirmation assay can be used as an efficient confirmation assay.

1945P Distribution of anti-PD1/PDL1 autoantibodies in multiple cancer types and potential biomarker role for anti-PD1 therapy

Antibodies targeting programmed cell death protein 1(PD1) and its ligand (PDL1) have revolutionized cancer therapy. However, little is known about the preexisted anti-PD1/PDL1 antoantibodies (AAbs) IgG and subclass distribution in cancer patients, nor is their potential role for anti-PD1 therapy. We used enzyme linked immune sorbent assay (ELISA) to examine serological anti-PD1/PDL1 IgG AAb and subclasses (IgG1-4) in 189 cancer patients without anti-PD1 therapy, covering 10 types of cancers (lung, breast, esophageal, colorectal, liver, prostatic, cervical, ovarian, gastric cancers and lymphoma). We further tested the anti-PD1/PDL1 AAb IgG, IgG1 and IgG2 in 16 non-small cell lung cancer (NSCLC) baseline plasma samples receiving anti-PD1 therapy, the association of AAb IgG subclasses with clinical parameters and overall survival were analyzed. Anti-PD1 AAb IgG and anti-PDL1 AAb IgG were globally detected in 10 types of cancer patients. Anti-PD1 AAb IgG was highest in breast cancer and lowest in colorectal cancer, while anti-PDL1 AAb IgG was highest in liver cancer and lowest in cervical carcinoma. Of the IgG subclasses, IgG1 and IgG2 were revealed to be the major subtypes for both anti-PD1 and anti-PDL1 AAbs in 10 cancer types. In 16 NSCLC patients receiving anti-PD1 therapy, the anti-PD1 IgG2 and anti-PDL1 IgG2 were significantly associated with white blood cell count(r=0.554, p=0.026; r=0.586, p=0.017), absolute neutrophil count(r=0.577, p=0.019; r=0.626, p=0.009) and absolute monocyte count (r=0.683, p=0.004; r=0.729, p=0.001). Higher anti-PD1 IgG2 was revealed to associate with improved anti-PD1 therapeutic survival, though no statistical significance were reached which may due to limited samples. This study reports for the first time the distribution of preexisted anti-PD1/PDL1 AAb and their subclasses in a wide range of cancer types. Moreover, the PD1 IgG2 were identified to associate with clinical parameters and may serve as potential biomarkers to predict anti-PD1 therapeutic survival in NSCLC and guide anti-PD1 treatment for cancer patients.

Increasing Hepatitis E Virus Seroprevalence in Domestic Pigs and Wild Boar in Bulgaria.

Simple SummaryHepatitis E virus (HEV) is a lesser-known hepatitis virus, but its worldwide spread is undisputed and has increased in recent years. The zoonotic spread of HEV, mainly due to genotype (gt) 3, emerged in developed countries in the past decade. In addition, transmission via contaminated meat from pigs and boars was also established. Detailed analysis of viral dynamics and distribution is needed in order to identify associated risk factors. The aim of the current study is to present new and additional data on the HEV distribution among pigs, and for the first-time, also among the wild boar population in Bulgaria.(1) Background: Hepatitis E virus (HEV) is a causative agent of acute viral hepatitis, predominantly transmitted by the fecal–oral route. In developed countries, HEV is considered to be an emerging pathogen since the number of autochthonous cases is rising. Hepatitis E is a viral disease with a proven zoonotic potential for some of its genotypes. The main viral reservoirs are domestic pigs and wild boar. Consumption of undercooked meat, as well as occupational exposure, are key factors for the spread of HEV. In order to evaluate the risks of future viral evolution, a detailed examination of the ecology and distribution of the virus is needed. The aim of the present study is to investigate the prevalence of anti-HEV IgG Ab in domestic pigs and wild boar in Bulgaria; (2) Methods: In this study, during the period of three years between 2017 and 2019, 433 serum samples from 19 different pig farms and 1 slaughterhouse were collected and analyzed. In addition, 32 samples from wild boar were also collected and analyzed during the 2018–2019 hunting season. All samples were analyzed by commercial indirect ELISA; (3) Results: Overall, HEV seroprevalence was 60% (95% CI 42.7–77.1) in domestic pigs and 12.5% (4/32) in wild boar. The observed seroprevalence of the slaughter-aged pigs was 73.65% (95% Cl 58.7–87.3). Prevalence in domestic pigs was significantly higher in the samples collected during 2019 (98% (95% Cl 96.1–99.9)) compared to those collected during 2017 (45.33% (95% CI 2.7–87.3)) and 2018 (38.46% (95% CI 29.1–49.7.); (4) Conclusions: Our findings suggest that domesticated pigs and wild boar might be the reason for the increased HEV transmission across Bulgaria. The genotypic characterization of HEV found in pigs, wild boar and humans will give a more accurate view of the zoonotic transmission of this virus.

Clinical validation of an immunochromatographic SARS-Cov-2 IgM/IgG antibody assay with Japanese cohort.

The coronavirus disease 2019 (COVID-19) has become a major health threat. To overcome COVID-19, appropriate diagnosis methods are urgently needed. The aim of this study was to clinically evaluate the colloidal gold immunochromatography assay for SARS-Cov-2 IgM/IgG antibody (Ab). Patients confirmed COVID-19 (n = 51) were recruited prospectively from the Musashino Red Cross hospital and Tokyo Medical and Dental University Medical Hospital, between March and May 2020. And the analytical specificity was assessed with serum samples of patients without COVID-19 (n = 100) collected between August to September 2019 before SARS-CoV-2 was first reported in China. Among COVID-19 patients, a total of 87 serum samples were tested for SARS-Cov-2 IgM/IgG Ab assay. IgM was detected 71.0 %, 86.9 %, and 83.3 % at day8-14, 15-28, >29 after symptom onset and IgG was detected in 81.6 %, 87.0 %, and 94.4 %, respectively. The sensitivity of IgM and IgG Ab after day8 assay was significantly higher than before day7, respectively (p=0.0016, 0.0003). There were no positive results in 100 serum samples from patients without COVID-19. The SARS-Cov-2 IgM/IgG Ab assay had 79.7% / 86.1% sensitivity (the 8 days after from onset) and 100% specificity in this population.

Optimal Aspergillus fumigatus and Asp f 1 serum IgG cut-offs for the diagnosis of allergic bronchopulmonary aspergillosis

BackgroundThe presence of IgG antibodies (Abs) to Aspergillus fumigatus (Af) is a crucial diagnostic criterion for allergic bronchopulmonary aspergillosis (ABPA). Although precipitation is traditionally used to document IgG Abs, anti-Af serum IgG levels can also be measured by enzyme immunoassay (EIA). However, there are insufficient data on the optimal cut-offs to assess diagnostic performance of the EIA method. This study aimed to determine cut-off levels of IgG binding crude Af extracts or recombinant Asp f 1 (by ImmunoCAP®) and to compare their efficacy for ABPA diagnosis with Af-precipitating Abs. MethodsThe age distribution of levels of IgG to crude extracts of Af (Af-IgG) and recombinant Asp f 1 (Asp f 1-IgG) was established using sera from 694 healthy controls (HC). Receiver operating characteristic analysis for Af-IgG and Asp f 1-IgG levels for the purpose of ABPA diagnosis was performed in 306 Af-sensitized asthma patients (including 49 ABPA), and cut-offs were determined. ResultsAn age-dependent decline in the levels of Af-IgG was observed in HC. Thus, cut-offs for Af-IgG levels were determined separately by age as 60 mg/L for patients aged <55 years, and 45 mg/L for those aged ≥55 years. For Asp f 1-IgG, 6.6 mg/L was set as the cut-off regardless of age. Although such IgG testing by EIA allowed a sufficiently good diagnostic performance, Af-precipitating Abs had better diagnostic applicability for ABPA. ConclusionsWe determined cut-offs for Af-IgG and Asp f 1-IgG measured by EIA, which can be useful in clinical settings where precipitating Abs are unavailable.

Autoantibodies against specific nuclear antigens are present in psoriatic disease and are diminished by secukinumab

Anti-nuclear antibodies (ANA) are frequently detected in patients with psoriasis (Ps) and psoriatic arthritis (PsA), but their target autoantigens remain unknown. We assessed antibody (ab) reactivity against 23 known nuclear antigens in patients with Ps and PsA and assess the effects of secukinumab (anti-IL17A) treatment on ANA levels. A total of 201 patients, 101 with Ps and 100 with PsA, and 50 ANA-negative healthy controls (HCs) were tested for ANAs by a line immunoassay testing reactivity to 23 nuclear antigens. Ab reactivity to at least 1 antigen was found in 20.4% psoriatic disease patients (25.7% Ps and 15% PsA) compared to 8% HCs (p = ns), the most frequent being against dense fine speckled 70 (DFS70) (6.5%). In Ps and PsA patients with secukinumab-induced remission, anti-DFS70 and other antigen-specific autoantibodies were diminished over time. No decline was noted for IgG abs against antigens from pathogens such as cytomegalovirus, Epstein–Barr virus and Helicobacter pylori. Autoantibody decrease was associated with significant reduction of plasmablasts, follicular B and follicular T cells. In conclusion, one third of antigen-specific ANA patients with psoriatic disease recognize DFS70. Secukinumab decreases nuclear antigen autoreactivity, plasmablasts, follicular B and follicular T cells, highlighting a new mechanism of its action.

Association of the Sialylation of Antibodies Specific to the HCV E2 Envelope Glycoprotein with Hepatic Fibrosis Progression and Antiviral Therapy Efficacy.

The E2 envelope glycoprotein of the hepatitis C virus (HCV) is a major target of broadly neutralizing antibodies that are closely related to a spontaneous cure of HCV infection. There is still no data about the diversity of E2-specific antibodies (Abs) glycosylation. The aim of this study was to analyze the level and sialylation of E2 IgG Abs, the relation of the respective changes to hepatic fibrosis (F) progression and their possible association with the efficacy of interferon-α-2a plus ribavirin (IFN-RBV) antiviral therapy. One hundred three HCV infected treatment-naive patients were examined using ELISA with E2 recombinant protein as antigen and sialic acid-specific Sambucus nigra agglutinin. The efficacy of the IFN-RBV treatment of patients with HCV dominant 1b and 3a genotypes (GT) was evaluated. A significant decrease of E2 Abs sialylation in the late stages of fibrosis was found irrespective of HCV genotype. On this basis, the F4 stage of fibrosis can be discriminated from its F0 or F1-3 stage by an about 75-79% accuracy. HCV infection of 1b genotype is associated with the production of lower sialylated E2 Abs, a higher frequency of F4 stage fibrosis, and a worse response to antiviral therapy. The increased SNA reactivity of E2 Abs was observed in patients with a sustained virological response (SVR). The proportion of SVR responders was significantly higher among patients with 3a genotype. However, for both dominant HCV genotypes (3a and 1b), an increased sialylation of E2 IgG was associated with a higher rate of patients with sustained virological response to antiviral therapy. Thus, the association of alterations of anti-E2 IgG Abs sialylation with hepatic fibrosis stage, HCV genotype, and the efficacy of antiviral therapy enables using these changes as novel noninvasive predictive biomarkers. The clinical potential of these findings is discussed.

Intravenous Infusion of Cortisol, Adrenaline, or Noradrenaline Alters Porcine Immune Cell Numbers and Promotes Innate over Adaptive Immune Functionality.

Despite the importance of pigs (Sus scrofa domestica) in livestock production and their increasing role as a model organism for human physiology, knowledge about the porcine immune system under the influence of stress hormones is fragmentary. Exceptionally little is known about the effects of catecholamines. Therefore, the aim of this study was to examine the in vivo effects of adrenaline, noradrenaline, and cortisol on number and functionality of porcine blood immune cells. Castrated male pigs (n = 34) were treated with physiological doses of either adrenaline, noradrenaline, or cortisol via i.v. infusion for 48 h. Blood samples were collected before treatment (-24 h, -22 h, 0 h), during treatment (+2 h, +24 h, +48 h), and at 72 h postinfusion. Immune cell numbers and phagocytic activity were evaluated by flow cytometry and lymphocyte proliferation by 3H-thymidine incorporation. Total IgG and IgM Ab levels were determined via ELISA. Pigs receiving cortisol showed strongly decreased adaptive immune cell numbers and increased neutrophils, accompanied by hampered lymphocyte proliferation but increased monocyte phagocytosis. Catecholamine effects on immune cell numbers were mostly similar to cortisol in direction but smaller in intensity and duration. Lymphocyte proliferation was inhibited after 2 h of noradrenaline infusion, and both catecholamines promoted monocyte and neutrophil phagocytosis. These findings indicate a shift from adaptive to innate immunity in stressful situations. This study is the first (to our knowledge) to systematically investigate specific glucocorticoid and catecholamine actions on the porcine immune system in this level of detail and confirms many similarities to humans, thus strengthening the pig as a human model in psychoneuroimmunology.