1.
Nonstructural protein NS1 immunodominant epitope detected specifically in dengue virus infected material by a SELDI-TOF/MS based assay.
Source
Biomarker Department, BioMerieux SA, Chemin de l'Orme, Marcy l'Etoile, France.
Abstract
Dengue virus (DV) infection is the most common mosquito-born viral disease of public health significance. Though most patients only suffer from flu-like symptoms, a small group of patients experiences more severe forms of the disease. The viral nonstructural protein 1 (NS1), a secreted protein correlating with viremia, is a key element used for dengue diagnosis with potential implications in severe dengue prognosis. Capture-ELISAs for the early detection of the NS1 protein in the sera during the acute febrile stage are commonly used in routine by diagnostic laboratories. In this study, the detection of NS1 protein in DV-infected material was assessed by an alternative method combining a single NS1-directed monoclonal antibody and the SELDI-TOF/MS technology. According to the epitope mapping, the antibodiesused are mainly directed against an immuno-dominant peptide located on the C-terminal part of the protein. The NS1 SELDI-TOF assay is specific, has a sensitivity level close to capture-ELISAs and is potentially useful for a coupled serotyping/detection assay or for the detection of subtle post-translational modifications on the protein. J. Med. Virol. 84:490-499, 2012. © 2011 Wiley Periodicals, Inc.
Copyright © 2012 Wiley Periodicals, Inc.
- PMID:
- 22246837
- [PubMed - in process]
Neuroplasticity of sensory and sympathetic nerve fibers in the painful arthritic joint.
Source
Research Service, VA Medical Center, One Veterans Drive, Minneapolis, MN 55417, USA.
Abstract
OBJECTIVE.: Many forms of arthritis are accompanied by significant chronic joint pain. Here we studied whether there is significant sprouting of sensory and sympathetic nerve fibers in the painful arthritic knee joint and whether nerve growth factor (NGF) drives this pathological reorganization. METHODS.: A painful arthritic knee joint was produced by injection of complete Freund's adjuvant (CFA) into the knee joint of young adult mice. CFA-injected mice were then treated systemically with vehicle or anti-NGF antibody. Pain behaviors were assessed and at 28 days following the initial CFA injection, the knee joints were processed for immunohistochemistry using antibodies raised against calcitonin gene-related peptide (CGRP; sensory nerve fibers), neurofilament 200 kDa (NF200; sensory nerve fibers), growth associated protein-43 (GAP43; sprouted nerve fibers), tyrosine hydroxylase (TH; sympathetic nerve fibers), CD31 (endothelial cells) or CD68 (monocytes/macrophages). RESULTS.: In CFA-injected mice, but not vehicle-injected mice, there was a significant increase in the density of CD68(+) macrophages, CD31(+) blood vessels, CGRP(+) , NF200(+) , GAP43(+) , and TH(+) nerve fibers in the synovium as well as joint pain-related behaviors. Administration of anti-NGF reduced these pain-related behaviors and the ectopic sprouting of nerve fibers, but had no significant effect on the increase in density of CD31(+) blood vessels or CD68(+) macrophages. CONCLUSIONS.: Ectopic sprouting of sensory and sympathetic nerve fibers occurs in the painful arthritic joint and may be involved in the generation and maintenance of arthritic pain.
Copyright © 2012 by the American College of Rheumatology.
- PMID:
- 22246649
- [PubMed - as supplied by publisher]
One-step immunopurification and lectinochemical characterization of the Duffy atypical chemokine receptor from human erythrocytes.
Source
Department of Immunochemistry, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, R. Weigla 12, 53-114, Wrocław, Poland.
Abstract
Duffy antigen/receptor for chemokines (DARC) is a glycosylated seven-transmembrane protein acting as a blood group antigen, a chemokine binding protein and a receptor for Plasmodium vivax malaria parasite. It is present on erythrocytes and endothelial cells of postcapillary venules. The N-terminal extracellular domain of the Duffy glycoprotein carries Fy(a)/Fy(b) blood group antigens and Fy6 linear epitope recognized by monoclonal antibodies. Previously, we have shown that recombinant Duffy protein expressed in K562 cells has three N-linked oligosaccharide chains, which are mainly of complex-type. Here we report a one-step purification method of Duffy protein from human erythrocytes. DARC was extracted from erythrocyte membranes in the presence of 1% n-dodecyl-β-D-maltoside (DDM) and 0.05% cholesteryl hemisuccinate (CHS) and purified by affinity chromatography using immobilized anti-Fy6 2C3 mouse monoclonal antibody. Duffy glycoprotein was eluted from the column with synthetic DFEDVWN peptide containing epitope for 2C3 monoclonal antibody. In this single-step immunoaffinity purification method we obtained highly purified DARC, which migrates in SDS-polyacrylamide gel as a major diffuse band corresponding to a molecular mass of 40-47 kDa. In ELISA purified Duffy glycoprotein binds anti-Duffy antibodies recognizing epitopes located on distinct regions of the molecule. Results of circular dichroism measurement indicate that purified DARC has a high content of α-helical secondary structure typical for chemokine receptors. Analysis of DARC glycans performed by means of lectin blotting and glycosidase digestion suggests that native Duffy N-glycans are mostly triantennary complex-type, terminated with α2-3- and α2-6-linked sialic acid residues with bisecting GlcNAc and α1-6-linked fucose at the core.
- PMID:
- 22246380
- [PubMed - as supplied by publisher]
Disruption of Platelet-derived Chemokine Heteromers Prevents Neutrophil Extravasation in Acute Lung Injury.
Source
RWTH Aachen, Aachen, Germany.
Abstract
RATIONALE:
Acute lung injury (ALI) causes high mortality, but its molecular mechanisms and therapeutic options remain ill-defined. Gram-negative bacterial infections are the main cause of ALI, leading to lung neutrophil infiltration, permeability increases, deterioration of gas exchange, and lung damage. Platelets are activated during ALI, but insights into their mechanistic contribution to neutrophil accumulation in the lung are elusive.
OBJECTIVES:
To determine mechanisms of platelet-mediated neutrophil recruitment in ALI.
METHODS:
Interference with platelet-neutrophil interactions using antagonists to P-selectin and glycoprotein IIb/IIIa or a small peptide antagonist disrupting platelet chemokine heteromer formation in mouse models of ALI. MEASUREMENTS AND
MAIN RESULTS:
In a murine model of LPS-induced ALI, we uncover important roles for neutrophils and platelets in permeability changes and subsequent lung damage. Furthermore, platelet depletion abrogated lung neutrophil infiltration, suggesting a sequential participation of platelets and neutrophils. Whereas antagonists to P-selectin and glycoprotein IIb/IIIa had no effects on LPS-mediated ALI, antibodies to the platelet-derived chemokines CCL5 and CXCL4 strongly diminished neutrophil eflux and permeability changes. The two chemokines were found to form heteromers in human and murine ALI samples, positively correlating with leukocyte influx into the lung. Disruption of CCL5-CXCL4 heteromers in LPS-, acid-, and sepsis-induced ALI abolished lung edema, neutrophil infiltration, and tissue damage, thereby revealing a causal contribution.
CONCLUSIONS:
Taken together, our data identify a novel function of platelet-derived chemokine heteromers during ALI and demonstrate means for therapeutic interference.
- PMID:
- 22246174
- [PubMed - as supplied by publisher]
Positron emission tomography evaluation of somatostatin receptor targeted (64)Cu-TATE-liposomes in a human neuroendocrine carcinoma mouse model.
Source
Technical University of Denmark, DTU Nanotech, Department of Micro- and Nanotechnology, Center for Nanomedicine and Theranostics, Building 423, 2800 Lyngby, Denmark; Technical University of Denmark, Hevesy Laboratory, Risø National Laboratory for Sustainable Energy, Frederiksborgvej 399, 4000 Roskilde, Denmark.
Abstract
Targeted therapeutic and diagnostic nanocarriers functionalized with antibodies, peptides or other targeting ligands that recognize over-expressed receptors or antigens on tumor cells have potential in the diagnosis and therapy of cancer. Somatostatin receptors (SSTRs) are over-expressed in a variety of cancers, particularly neuroendocrine tumors (NETs) and can be targeted with somatostatin peptide analogs such as octreotate (TATE). In the present study we investigate liposomes that target SSTR in a NET xenograft mouse model (NCI-H727) by use of TATE. TATE was covalently attached to the distal end of DSPE-PEG(2000) on PEGylated liposomes with an encapsulated positron emitter (64)Cu that can be utilized for positron emission tomography (PET) imaging. The biodistribution and pharmacokinetics of the (64)Cu-loaded PEGylated liposomes with and without TATE was investigated and their ability to image NETs was evaluated using PET. Additionally, the liposome accumulation and imaging capability was compared with free radiolabelled TATE peptide administered as (64)Cu-DOTA-TATE. The presence of TATE on the liposomes resulted in a significantly faster initial blood clearance in comparison to control-liposomes without TATE. PEGylated liposomes with or without TATE accumulated at significantly higher quantities in NETs (5.1±0.3 and 5.8±0.2 %ID/g, respectively) than the free peptide (64)Cu-DOTA-TATE (1.4±0.3 %ID/g) 24h post-injection. Importantly, (64)Cu-loaded PEGylated liposomes with TATE showed significantly higher tumor-to-muscle (T/M) ratio (12.7±1.0) than the control-liposomes without TATE (8.9±0.9) and the (64)Cu-DOTA-TATE free peptide (7.2±0.3). The higher T/M ratio of the PEGylated liposomes with TATE suggests some advantage of active targeting of NETs, although no absolute benefit in tumor accumulation over the non-targeted liposomes was observed. Collectively, these data showed that (64)Cu-loaded PEGylated liposomes with TATE conjugated to the surface could be promising new imaging agents for visualizing tumor tissue and especially NETs using PET.
Copyright © 2011. Published by Elsevier B.V.
- PMID:
- 22245688
- [PubMed - as supplied by publisher]
Measuring and evaluating the role of ATP-Sensitive K(+) channels in cardiac muscle.
Source
Department of Pediatrics, NYU School of Medicine, New York, NY 10016, USA.
Abstract
Since ion channels move electrical charge during their activity, they have traditionally been studied using electrophysiological approaches. This was sometimes combined with mathematical models, for example with the description of the ionic mechanisms underlying the initiation and propagation of action potentials in the squid giant axon by Hodgkin and Huxley. The methods for studying ion channels also have strong roots in protein chemistry (limited proteolysis, the use of antibodies, etc.). The advent of the molecular cloning and the identification of genes coding for specific ion channel subunits in the late 1980s introduced a multitude of new techniques with which to study ion channels and the field has been rapidly expanding ever since (e.g. antibody development against specific peptidesequences, mutagenesis, the use of gene targeting in animal models, determination of their protein structures) and new methods are still in development. This review focuses on techniques commonly employed to examine ion channel function in an electrophysiological laboratory. The focus is on the K(ATP) channel, but many of the techniques described are also used to study other ion channels.
Copyright © 2011. Published by Elsevier Ltd.
- PMID:
- 22245446
- [PubMed - as supplied by publisher]
Penaeus monodon tropomyosin induces CD4 T-cell proliferation in shrimp-allergic patients.
Source
Department of Dermatology, University of Utah, Salt Lake City, Utah, USA.
Abstract
Shellfish allergy affects approximately 2% of the population and can cause immediate hypersensitivity reactions such as urticaria, swelling, difficulty breathing, and, in some cases, anaphylaxis. Tropomyosin is the major shrimp allergen and binds IgE in two-thirds of patients. A total of 38 shrimp-allergic patients and 20 negative control subjects were recruited and evaluated on the basis of history, skin prick testing, specific immunoglobulin E (IgE) levels, and peripheral blood mononuclear cell proliferation in response to shrimp tropomyosin or shrimp tropomyosin-derived peptides. Of the classically allergic patients by history, 59%tested positive for serum shrimp IgE antibodies. Of patients with shrimp-specific IgE in sera, 70% also had significant IgE levels specific for shrimp tropomyosin. Peripheral blood mononuclear cells from classically shrimp-allergic patients proliferated in a dose-dependent manner in response to to tropomyosin. In addition, a T-cell line derived from a shrimp-allergic patient proliferated specifically in response to tropomyosin-derivedpeptides. These studies suggest a strategy for immunotherapy using a tropomyosin-derived T-cell epitope vaccination.
Copyright © 2012. Published by Elsevier Inc.
- PMID:
- 22244920
- [PubMed - as supplied by publisher]
Inhibition of Amyloid Formation.
Abstract
Amyloid is aggregated protein in the form of insoluble fibrils. Amyloid deposition in human tissue-amyloidosis-is associated with a number of diseases including all common dementias and type II diabetes. Considerable progress has been made to understand the mechanisms leading to amyloid formation. It is, however, not yet clear by which mechanisms amyloid and protein aggregates formed on the path to amyloid are cytotoxic. Strategies to prevent protein aggregation and amyloid formation are nevertheless, in many cases, promising and even successful. This review covers research on intervention of amyloidosis and highlights several examples of how inhibition of protein aggregation and amyloid formation has been achieved in practice. For instance, rational design can provide drugs that stabilize a native folded state of a protein, protein engineering can provide new binding proteins that sequester monomeric peptides from aggregation, small molecules and peptides can be designed to block aggregation or direct it into non-cytotoxic paths, and monoclonal antibodies have been developed for therapies towards neurodegenerative diseases based on amyloid inhibition and clearance.
Copyright © 2012. Published by Elsevier Ltd.
- PMID:
- 22244855
- [PubMed - as supplied by publisher]
A General LC-MS/MS Method Approach to Quantify Therapeutic MonoclonalAntibodies Using a Common Whole Antibody Internal Standard with Application to Preclinical Studies.
Abstract
Ligand binding assays (LBAs) are widely used for monoclonal antibody (mAb) Ligand binding assays (LBAs) are widely used for therapeutic monoclonal antibody (mAb) quantification in biological samples. Major limitations are long method development times, reagent procurement, and matrix effects. LC-MS/MS methods using signature peptides are emerging as an alternative approach, which typically use a stable isotope labeled signature peptide as the internal standard (IS). However, a new IS has to be generated for every candidate, and the IS may not correct for variations at all processing steps. We have developed a general LC-MS/MS method approach employing a uniformly heavy-isotope labeled common whole mAb IS and a common immunocapture for sample processing. The method was streamlined with automation for consistency and throughput. Method qualification of four IgG2 and four IgG1 mAbs showed sensitivity of 0.1 µg/mL and linearity of 0.1-15 µg/mL. Quality control (QC) data of these eight mAbs were accurate and precise. The QC performance of the whole molecule labeled IS was better than those of synthetic labeled IS peptidestested. The pharmacokinetic results of two mAbs (an IgG2 and IgG1 candidate) dosed in rats were comparable to those of LBA. The general LC-MS/MS method approach overcomes the limitations of current methods to reduce time and resources required for preclinical studies.
- PMID:
- 22243404
- [PubMed - as supplied by publisher]
[Progress in the study of allergic disease drugs targeting on IgE/FcepsilonRI signaling pathway].
Source
Key Laboratory for Drug Quality Control and Analysis of Hebei Province, Baoding 071002, China. liuzc@hbu.edu.cn
Abstract
Allergic diseases have become global social health problems. The binding of IgE with its high affinity receptor FcepsilonRI plays a key step in I-type allergy. Recently, more and more key molecules on the IgE/FcepsilonRI signaling transduction pathway were to be the drug candidates against allergic diseases, with in-depth study of FcepsilonRI signal pathway gradually. The main drugs include molecule antibodies, peptides, vaccines, fusion proteins, small molecules, and other drugs related to IgE/FcepsilonRI. The recent progress in the study of mechanisms of representative drugs targeting on IgE/FcepsilonRI signaling pathway was reviewed in this article.
- PMID:
- 22242444
- [PubMed - in process]
Interleukin 15 levels in serum may predict a severe disease course in patients with early arthritis.
Source
Rheumatology Service, Hospital Universitario de La Princesa, IIS Princesa, Madrid, Spain.
Abstract
BACKGROUND:
Interleukin-15 (IL-15) is thought to be involved in the physiopathological mechanisms of RA and it can be detected in the serum and the synovial fluid of inflamed joints in patients with RA but not in patients with osteoarthritis or other inflammatory joint diseases. Therefore, the objective of this work is to analyse whether serum IL-15 (sIL-15) levels serve as a biomarker of disease severity in patients with early arthritis (EA).
METHODOLOGY AND RESULTS:
Data from 190 patients in an EA register were analysed (77.2% female; median age 53 years; 6-month median disease duration at entry). Clinical and treatment information was recorded systematically, especially the prescription of disease modifying anti-rheumatic drugs. Two multivariate longitudinal analyses were performed with different dependent variables: 1) DAS28 and 2) a variable reflecting intensive treatment. Both included sIL-15 as predictive variable and other variables associated with disease severity, including rheumatoid factor (RF) and anti-cyclic citrullinated peptide antibodies (ACPA). Of the 171 patients (638 visits analysed) completing the follow-up, 71% suffered rheumatoid arthritis and 29% were considered as undifferentiated arthritis. Elevated sIL-15 was detected in 29% of this population and this biomarker did not overlap extensively with RF or ACPA. High sIL-15 levels (β Coefficient [95% confidence interval]: 0.12 [0.06-0.18]; p<0.001) or ACPA (0.34 [0.01-0.67]; p = 0.044) were significantly and independently associated with a higher DAS28 during follow-up, after adjusting for confounding variables such as gender, age and treatment. In addition, those patients with elevated sIL-15 had a significantly higher risk of receiving intensive treatment (RR 1.78, 95% confidence interval 1.18-2.7; p = 0.007).
CONCLUSIONS:
Patients with EA displaying high baseline sIL-15 suffered a more severe disease and received more intensive treatment. Thus, sIL-15 may be a biomarker for patients that are candidates for early and more intensive treatment.
Peptidyl-arginine deiminase: an additional marker of rheumatoid arthritis.
Source
Biotechnology Department, Heritage Institute of Technology, Chowbagha Road, Anandapur, Kolkata, India. psbasu.heritage@gmail.com
Abstract
BACKGROUND:
Antibodies against cyclic citrullinated peptide (anti-CCP) were thought to be more specific than rheumatoid factor (RF) for the diagnosis of rheumatoid arthritis (RA). The determination of anti-CCP in addition to RF could be helpful in the serological diagnosis and monitoring of patients with RA. Citrullination of proteins involves the enzymatic conversion of protein containing arginine residues to citrulline residues by the enzyme peptidylarginine deiminase (PAD). The present investigation was undertaken to estimate serum PAD enzyme activity in RA patients with a view to find its importance as a new diagnosis marker in a rheumatology clinic.
METHODS:
The activity of the PAD enzyme was measured by spectrophotometric method at 530 nm in sera of control subjects and in patients of RA (Group I: RF negative and CCP positive: Group II: both RF and CCP positive) in terms of citrulline formation using benzoyl-arginine ethyl ester (BAEE) as substrate. Anti-CCP and RF were also estimated in two groups by enzyme immunoassay and immunoturbidimetry for comparison. Clinical variables (duration of morning stiffness, swollen and tender joint counts, patient's assessment of pain) and C-reactive protein were also evaluated.
RESULTS:
A marked increase in PAD enzyme activity (p < 0.001) was noted in RA patients in comparison to controls and the level diminished appreciably along with two known serological markers (anti-CCP and RF) after six months of disease modifying antirheumatic drug (DMARD) treatment. The Group II RA patients showed much higher enzyme activity than Group I RA patients. However, clinical variables did not differ significantly between the two Groups of RA patients.
CONCLUSIONS:
We conclude that determination of PAD enzyme activity may be used as an additional marker for monitoring disease progression and regression along with anti-CCP and RF in patients with RA. Moreover, this method is rapid, sensitive, and inexpensive and can be adopted in a laboratory having modest facilities.
- PMID:
- 22239037
- [PubMed - in process]
Neutralizing Epitopes in the MPER of HIV-1 gp41 Are Influenced by the Transmembrane Domain and the Plasma Membrane.
Source
Dept. of Molecular Biology and Biochemistry.
Abstract
Failure to elicit broadly (b) neutralizing (Nt) antibodies (Abs) against the membrane proximal external region of HIV-1 gp41 (MPER), reflects the difficulty of mimicking its neutralization-competent structure (NCS). Here, we analyzed MPER antigenicity in the context of the plasma membrane, and identified a role for the gp41 transmembrane domain (TM) in exposing the epitopes of three bNt monoclonal (M)Abs (2F5, 4E10, Z13e1). We transiently expressed DNA constructs encoding gp41 ectodomain fragments fused to either the TM of the platelet-derived growth factor receptor (PDGFR), or the gp41 TM and cytoplasmic tail domain (CT). Constructs encoding the MPER tethered to the gp41 TM followed by a 27-residue CT fragment (MPER-TM1) produced optimal MAb binding. Critical binding residues for the three Nt MAbs were identified using a panel of 24 MPER-TM1 mutants bearing single amino-acid substitutions in the MPER; many were previously shown to affect MAb-mediated viral neutralization. Moreover, non-Nt mutants of MAbs 2F5 and 4E10 exhibited a reduction in binding to MPER-TM1, yet maintained binding to synthetic MPER peptides, indicating that MPER-TM1 better approximates the MPER NCS than peptides. Substitution of the gp41 TM and CT of MPER-TM1 with the PDGFR TM reduced binding by MAb 4E10, but not 2F5, indicating the gp41 TM plays a pivotal role in orienting the 4E10 epitope, and more globally, in affecting MPER exposure.
Evaluation of Biological Sample Preparation for Immunosignature Based Diagnostics.
Source
Center for Innovations in Medicine, The Biodesign Institute, Arizona State University, Tempe, AZ 85287-5901.
Abstract
To address the need for a universal system to assess health status, we previously described a method termed immunosignaturing which splays the entire humoral antibody repertoire across a peptide microarray. Two important issues relative to the potential broad use of immunosignatures are sample preparation and stability. In the present study we compared the immunosignatures developed from serum, plasma, saliva and antibodies eluted from blood dried onto filter paper. We found serum and plasma provide identical immunosignatures. Dried blood also correlated well with non-dried serum from the same individual. Immunosignatures derived from dried blood were capable of distinguishing naïve mice from those infected with influenza. Saliva was applied to the arrays and the IgA immunosignature correlated strongly with that from dried blood. Finally, we demonstrate that dried blood retains immunosignature information even when exposed to high temperature. This work expands the potential diagnostic uses for immunosignatures. These features suggest that different forms of archival samples can be used for diagnosis development and that in prospective studies samples can be easily procured.
Anti-cytokine auto-vaccinations as tools for the analysis of cytokine function in vivo.
Source
Ludwig Institute for Cancer Research, Brussels Branch, Brussels, Belgium; Cellular Genetics Unit, Christian de Duve Institute of Cellular Pathology, Université Catholique de Louvain, Brussels, Belgium.
Abstract
Braking B cell tolerance to generate antibodies against autologous cytokines or chemokines offers an alternative to gene inactivation for functional analysis of these factors in vivo. It is clearly less potent than the genetic approach but offers the advantage of extreme flexibility. The basic principle is to enable a self-reactive B cell to attract T cell help by presenting foreign peptides, a process we called "deceptive" antigen presentation. We here review the different auto-vaccine procedures that are currently used and provide several examples of functional information acquired by this procedure or by mAbs derived from auto-vaccinated mice.
Copyright © 2011 Elsevier Ltd. All rights reserved.
Immunolocalization of sprouty-1 and sprouty-2 in developing rat lung.
Source
Department of Pathology, University of Pittsburgh School of Medicine, Pittsburgh, Pa., USA.
Abstract
Objective: Sprouty, a common antagonist of fibroblast growth factor (FGF) and epidermal growth factor signaling, is a key player regulating tracheal branching and eye development in Drosophila. Four Sprouty homologs have been identified in vertebrates and all share a cysteine-rich region. However, the physiological function(s) of the individual Sprouty homologs is unknown. mRNA of Sprouty homologs is expressed during mouse lung development. In the present study, we investigated the immunolocalization of Sprouty proteins in rat lung at different stages of development. Methods: Rabbit antibodies were raised against peptides derived from rat Sprouty-1 and Sprouty-2 and were used in Western blot analysis to determine Sprouty distribution in subcellular fractions (pellets and supernatant centrifuged at 5,000 and 20,000 g) and bronchoalveolar lavage fluid (BAL) from adult rat lungs or used in immunohistochemistry. Results: Western blot analysis revealed a 30-kDa Sprouty-1 band and a 34-kDa Sprouty-2 band in the supernatant and pellet fractions centrifuged at 20,000 g. BAL contained a band of approximately 16 kDa with Sprouty-1 antibody derived from proteolytic fragmentation of Sprouty-1. In embryonic day (E) 14 and E16 lungs, Sprouty-1 and Sprouty-2 were expressed both in epithelial and peripheral mesenchymal cells. In adult rat lung, bronchiolar and alveolar type II epithelial cells showed staining for both Sprouty-1 and Sprouty-2. Sprouty-1 expression was also seen in alveolar type I epithelial cells. Conclusion: In light of the proximity of the distribution of Sprouty to that of FGF-10 (peripheral mesenchyme) and its receptor FGFR2IIIb (distal tubular epithelium) in lung development, and the finding that FGF-9, which is expressed in mesothelial cells, upregulates FGF-10, it appears that Sprouty expression in epithelial and mesenchymal cells during branching morphogenesis is closely related to signaling by FGF-9 and FGF-10.
Copyright © 2012 S. Karger AG, Basel.
Clinical Relevance of Anti-exenatide Antibodies: Safety, Efficacy, and Cross-reactivity with Long-term Treatment.
Source
Elcelyx Therapeutics, Inc., San Diego, CA Eli Lilly and Company, Indianapolis, IN, USA Diabetes Center, VU University Medical Center, Amsterdam, the Netherlands American Diabetes Association, Alexandria, VA, USA Amylin Pharmaceuticals, Inc., San Diego, CA.
Abstract
Aims Antibody formation to therapeutic peptides is common. This analysis characterizes the time-course and cross-reactivity of anti-exenatide antibodies and potential effects on efficacy and safety. Materials and Methods Data from ITT patients in 12 controlled (n=2225, 12-52wks) and 5 uncontrolled (n=1538, up to 3yrs) exenatide BID trials and 4 controlled (n=653, 24-30wks) exenatide once weekly (QW) trials with 1 uncontrolled period (n=128, 52wks) were analyzed. Results Mean titers peaked early (6-22wks), and subsequently declined. At 30wks, 36.7% of exenatide BID patients were antibody-positive; 31.7% exhibited low titers (≤125) and 5.0% had higher titers (≥625). Antibody incidence declined to 16.9% (1.4% higher titer) at 3yrs. Similarly, 56.8% of exenatide QW patients were antibody-positive (45.0% low/11.8% higher titer) at 24-30wks, declining to 45.4% positive (9.2% higher titer) at 52wks. Treatment-emergent anti-exenatide antibodies from a subset of patients tested did not cross-react with human GLP-1 or glucagon. Other than injection-site reactions, adverse event rates in antibody-positive and antibody-negative patients were similar. Efficacy was robust in both antibody-negative and antibody-positive patients (mean HbA(1c) : -1.0% and -0.9%, respectively, exenatide BID; -1.6% and -1.3% exenatide QW). No correlation between change in HbA(1c) and titer was observed for exenatide BID, although mean reductions were attenuated in the small subset of patients (5%) with higher titers. A significant correlation was observed for exenatide QW, with no difference between antibody negative and low-titer patients but an attenuated mean reduction in the subset of patients (12%) with higher titers. Conclusions Low titer anti-exenatide antibodies were common with exenatide treatment (32% exenatide BID, 45% exenatide QW patients), but had no apparent effect on efficacy. Higher titer antibodies were less common (5% exenatide BID, 12% exenatide QW) and within that titer group, increasing antibody titer was associated with reduced average efficacy that was statistically significant for exenatide QW. Other than injection-site reactions, anti-exenatide antibodies did not impact the safety of exenatide.
Copyright © 2012 John Wiley & Sons A/S.
Circumscribing the Conformational Peptide Epitope Landscape.
Source
Department of Biochemistry and Molecular Biology, University of Bari, Via Orabona 4, 70126 Bari, Italy. d.kanduc@biologia.uniba.it.
Abstract
The development of vaccines for new and re-emerging pathologies and infections is based on the ability to define immunogenic epitopes. An immunogenic B-cell peptide epitope is a specific restricted antigen region that is capable of eliciting a humoral immune response and of combining with a specific site on antibodies. Using a number of experimental models and based on data from several literature reports, we identified low levels of sequence similarity to the host proteome as one of the main factors modulating the B-cell epitope repertoire in the humoral immune response. In point of fact, a low level of sequence identity to the host proteins is a common denominator unifying the composite, disparate assembly of linear peptide B-cell epitopes that has been experimentally validated and described in the literature. Here, we explore the proteomic similarity of conformational epitopes experimentally validated and described in published reports. Again, discontinuous epitopic structures formed by non-contiguous amino acid residues were found to define immunological peptide units with a low level of similarity to the host. The present meta-analysis adds further significance to the immunological low-similarity theory and its clinical implications. Potentially, low-similarity peptideepitopes pave the way for novel effective vaccines in cancer, autoimmunity, and infectious diseases.
- PMID:
- 22236129
- [PubMed - as supplied by publisher]
Epitope specificity of anti-HA2 antibodies induced in humans during influenza infection.
Source
Institute of Virology, Slovak Academy of Sciences, Bratislava, Slovak Republic Public Health Authority of the Slovak Republic, National Influenza Centre, Bratislava, Slovak Republic.
Abstract
Please cite this paper as: Stanekováet al. (2012) Epitope specificity of anti-HA2 antibodies induced in humans during influenza infection. Influenza and Other Respiratory Viruses DOI: 10.1111/j.1750-2659.2011.00328.x Background The conserved, fusion-active HA2 glycopolypeptide (HA2) subunit of influenza A hemagglutinin comprises four distinct antigenic sites. Monoclonal antibodies (MAbs) recognizing three of these sites are broadly cross-reactive and protective. Objectives This study aimed to establish whether antibodies specific to these three antigenic sites were elicited during a natural influenza infection or by vaccination of humans. Methods Forty-five paired acute and convalescent sera from individuals with a confirmed influenza A (subtype H3) infection were examined for the presence of HA2-specificantibodies. The fraction of antibodies specific to three particular antigenic sites (designated IIF4, FC12, and CF2 here) was investigated using competitive enzyme immunoassay. Results Increased levels of antibodies specific to an ectodomain of HA2 (EHA2: N-terminal residues 23-185 of HA2) were detected in 73% of tested convalescent sera (33/45), while an increased level of antibodies specific to the HA2 fusion peptide (N-terminal residues 1-38) was induced in just 15/45 individuals (33%). Competitive assays confirmed that antibodies specific to the IIF4 epitope (within HA2 residues 125-175) prevailed in 86% (13/15) over those specific to the other two epitopes during infection. However, only a negligible increase in HA2-specific antibodies was detectable following vaccination with a current subunit vaccine. Conclusions We observed that the antigenic site localized within N-terminal HA2 residues 125-175 was more immunogenic than that within residues 1-38 (HA2 fusion protein), although both are weak natural immunogens. We suggest that new anti-influenza vaccines should include HA2 (or specific epitopes localized within this glycopolypeptide) to enhance their cross-protective efficacy.
© 2012 Blackwell Publishing Ltd.
- PMID:
- 22236105
- [PubMed - as supplied by publisher]
CD40/APC-specific antibodies with three T-cell epitopes loaded in the constant domains induce CD4+ T-cell responses.
Source
Department of Molecular Biosciences, Centre for Immune Regulation, University of Oslo, PO Box 1041, Blindern, NO-0316 Oslo, Norway.
Abstract
CD4(+) T lymphocytes play a central role in the orchestration and maintenance of the adaptive immune response. Targeting of antigen to antigen presenting cells (APCs) increases peptide loading of major histocompatibility complex (MHC) class II molecules and CD4(+) T-cell activation. APCs have been targeted by APC-specific recombinantantibodies (rAbs) with single T-cell epitopes integrated in the constant region of the heavy chain (C(H)). However, the strategy may be improved if several T-cell epitopes could be delivered simultaneously by one rAb. We here demonstrate that a single rAb can be loaded with multiple identical or different T-cell epitopes, integrated as loops between β-strands in C(H) domains. One epitope was inserted in C(H)1, while two were placed in C(H)2 of IgG. T-cell proliferation assays showed that all three peptides were excised from loops and presented on MHC class II to T-cells. Induction of T-cell activation by each epitope in the multi-peptide rAb was as good, or even better, than that elicited by corresponding single-peptide rAbs. Furthermore, following DNA vaccination of mice with plasmids that encode CD40-specific rAbs loaded with either one or three peptides, T-cell responses were induced. Thus, integration of multiple epitopes in C(H) region loops of APC-specific rAbs is feasible and may be utilized in design of multi-vaccines.
No comments:
Post a Comment