Disease-specific Antigen Research Articles

FASTMAP-a flexible and scalable immunopeptidomics pipeline for HLA- and antigen-specific T-cell epitope mapping based on artificial antigen-presenting cells.

The study of peptide repertoires presented by major histocompatibility complex (MHC) molecules and the identification of potential T-cell epitopes contribute to a multitude of immunopeptidome-based treatment approaches. Epitope mapping is essential for the development of promising epitope-based approaches in vaccination as well as for innovative therapeutics for autoimmune diseases, infectious diseases, and cancer. It also plays a critical role in the immunogenicity assessment of protein therapeutics with regard to safety and efficacy concerns. The main challenge emerges from the highly polymorphic nature of the human leukocyte antigen (HLA) molecules leading to the requirement of a peptide mapping strategy for a single HLA allele. As many autoimmune diseases are linked to at least one specific antigen, we established FASTMAP, an innovative strategy to transiently co-transfect a single HLA allele combined with a disease-specific antigen into a human cell line. This approach allows the specific identification of HLA-bound peptides using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Using FASTMAP, we found a comparable spectrum of endogenous peptides presented by the most frequently expressed HLA alleles in the world's population compared to what has been described in literature. To ensure a reliable peptide mapping workflow, we combined the HLA alleles with well-known human model antigens like coagulation factor VIII, acetylcholine receptor subunit alpha, protein structures of the SARS-CoV-2 virus, and myelin basic protein. Using these model antigens, we have been able to identify a broad range of peptides that are in line with already published and in silico predicted T-cell epitopes of the specific HLA/model antigen combination. The transient co-expression of a single affinity-tagged MHC molecule combined with a disease-specific antigen in a human cell line in our FASTMAP pipeline provides the opportunity to identify potential T-cell epitopes/endogenously processed MHC-bound peptides in a very cost-effective, fast, and customizable system with high-throughput potential.

Treatment with an antigen-specific dual microparticle system reverses advanced multiple sclerosis in mice

Antigen-specific therapies hold promise for treating autoimmune diseases such as multiple sclerosis while avoiding the deleterious side effects of systemic immune suppression due to delivering the disease-specific antigen as part of the treatment. In this study, an antigen-specific dual-sized microparticle (dMP) treatment reversed hind limb paralysis when administered in mice with advanced experimental autoimmune encephalomyelitis (EAE). Treatment reduced central nervous system (CNS) immune cell infiltration, demyelination, and inflammatory cytokine levels. Mechanistic insights using single-cell RNA sequencing showed that treatment impacted the MHC II antigen presentation pathway in dendritic cells, macrophages, B cells, and microglia, not only in the draining lymph nodes but also strikingly in the spinal cord. CD74 and cathepsin S were among the common genes down-regulated in most antigen presenting cell (APC) clusters, with B cells also having numerous MHC II genes reduced. Efficacy of the treatment diminished when B cells were absent, suggesting their impact in this therapy, in concert with other immune populations. Activation and inflammation were reduced in both APCs and T cells. This promising antigen-specific therapeutic approach advantageously engaged essential components of both innate and adaptive autoimmune responses and capably reversed paralysis in advanced EAE without the use of a broad immunosuppressant.

MRNA as a Therapeutics: Understanding mRNA Vaccines.

Vaccination is one of the important approaches in the prevention and control of diseases. Although the capacity to present antigens other than the disease-specific antigen in the traditional vaccine composition provides a potential benefit by increasing its protective efficacy, many components that are not needed for the related disease are also transferred. These components can reduce vaccine activity by lowering immunity against protective antigens. The reasons such as the low effectiveness of traditional vaccines and the high cost of production and time-consuming reasons show that it is necessary to develop a new vaccine method for our world, which is struggling with epidemics almost every year. Among nucleic acids, mRNA has many advantages, such as genomic integration, induction of anti-DNA autoantibodies, and immune tolerance induced by long-term antigen expression. mRNA vaccines have become a therapeutic target for reasons such as efficacy, safety, fast and non-expensive production. The fact that mRNA triggers both humoral and cellular immunity and goes only to the cytoplasm, not to the nucleus, makes it highly efficient. The mRNA must cross the lipid bilayer barrier and entry to the cytoplasm where it is translated into protein. There are two main ways of mRNA vaccine delivery for this: ex vivo loading of mRNA into dendritic cells (DCs) and direct injection of mRNA with or without a carrier. Studies continue to understand which delivery system is therapeutically more efficient. Preclinical and clinical trials showed that mRNA vaccines trigger a long-lasting and safe immune response.

Disease-specific antigen presentation on MHC class II is inhibited by a small molecule in Sjögren’s Syndrome

Abstract Sjogren’s Syndrome (SjS) is one of the more common autoimmune diseases that has a strong genetic linkage. To date, no vaccine or therapeutic exists to cure SjS, and patients must rely on lifelong therapies to treat symptoms. Class II MHC are the primary susceptibility loci that form the genetic basis for many auto-immune disorders including SjS. I-Ag7 is the MHC in NOD mice that is used for the presentation of autoantigens and plays a critical role in the adaptive immune response. HLA-DQ8 the MHC in humans, is analogous to I-Ag7, in the NOD mouse model and is an important model for the onset of SjS. Using an in-silico molecular docking program a chemical library was screened that defined small molecules that were capable of occupying specific structural pockets along the I-Ag7 binding groove, with the objective of influencing auto-antigen peptide presentation to T cells. In this study we show, that the small molecule TATD can inhibit TCR signaling based on the structural pocket targeting mechanism. TATD previously proven to inhibit the onset of diabetes by blocking the auto-antigen presentation on I-Ag7, was analyzed to observe effects of prevention of SjS by, preventing the accumulation of B and T cells within the salivary gland of NOD mice. Treated mice also showed reduction in the incidence of inflammatory cell infiltration into lacrimal and salivary glands as compared to control mice with notable reduction in of the hallmark SjS anti-nuclear antibodies of Ro52, Ro60 and La. As a result this study presents a novel method for identifying small molecules capable of inhibiting T cell responses, with potentially therapeutic applications.

Multipathogen Analysis of IgA and IgG Antigen Specificity for Selected Pathogens in Milk Produced by Women From Diverse Geographical Regions: The INSPIRE Study.

Breastfeeding provides defense against infectious disease during early life. The mechanisms underlying this protection are complex but likely include the vast array of immune cells and components, such as immunoglobulins, in milk. Simply characterizing the concentrations of these bioactives, however, provides only limited information regarding their potential relationships with disease risk in the recipient infant. Rather, understanding pathogen and antigen specificity profiles of milk-borne immunoglobulins might lead to a more complete understanding of how maternal immunity impacts infant health and wellbeing. Milk produced by women living in 11 geographically dispersed populations was applied to a protein microarray containing antigens from 16 pathogens, including diarrheagenic E. coli, Shigella spp., Salmonella enterica serovar Typhi, Staphylococcus aureus, Streptococcus pneumoniae, Mycobacterium tuberculosis and other pathogens of global health concern, and specific IgA and IgG binding was measured. Our analysis identified novel disease-specific antigen responses and suggests that some IgA and IgG responses vary substantially within and among populations. Patterns of antibody reactivity analyzed by principal component analysis and differential reactivity analysis were associated with either lower-to-middle-income countries (LMICs) or high-income countries (HICs). Antibody levels were generally higher in LMICs than HICs, particularly for Shigella and diarrheagenic E. coli antigens, although sets of S. aureus, S. pneumoniae, and some M. tuberculosis antigens were more reactive in HICs. Differential responses were typically specific to canonical immunodominant antigens, but a set of nondifferential but highly reactive antibodies were specific to antigens possibly universally recognized by antibodies in human milk. This approach provides a promising means to understand how breastfeeding and human milk protect (or do not protect) infants from environmentally relevant pathogens. Furthermore, this approach might lead to interventions to boost population-specific immunity in at-risk breastfeeding mothers and their infants.

Design of biodegradable nanoparticles to modulate phenotypes of antigen-presenting cells for antigen-specific treatment of autoimmune disease

Current therapeutic options for autoimmune diseases, such as multiple sclerosis (MS), often require lifelong treatment with immunosuppressive drugs, yet strategies for antigen-specific immunomodulation are emerging. Biodegradable particles loaded with disease-specific antigen, either alone or with immunomodulators, have been reported to ameliorate disease. Herein, we hypothesized that the carrier could impact polarization of the immune cells that associate with particles and the subsequent disease progression. Single injection of three polymeric carriers, 50:50 poly (DL-lactide-co-glycolide) (PLG) with two molecular weights (Low, High) and poly (DL-lactide) (PLA), loaded with the disease-specific antigen, proteolipid protein (PLP139-151), were investigated for the ability to attenuate clinical scores in experimental autoimmune encephalomyelitis (EAE), a mouse model of MS. At a low particle dose, mice treated with PLA-based particles had significantly lower clinical scores at the chronic stage of the disease over 200 days post immunization, while neither PLG-based particles nor OVA control particles reduced the clinical scores. Compared to PLG-based particles, PLA-based particles were largely associated with Kupffer cells and liver sinusoidal endothelial cells, which had a reduced co-stimulatory molecule expression that correlated with a reduction of CD4+ T-cell populations in the central nervous system. Delivery of PLA-based particles encapsulated with higher levels of PLP139-151 at a reduced dose were able to completely ameliorate EAE over 200 days along with inhibition of Th1 and Th17 polarization. Collectively, our study demonstrates that the carrier properties and antigen loading determine phenotypes of immune cells in the peripheral organs, influencing the amelioration of both acute and chronic stages of autoimmunity.

Rationally designed small molecules to prevent type 1 diabetes.

To review the recent findings that small 'drug-like' compounds block disease-specific human leukocyte antigen (HLA) molecules in type 1 diabetes (T1D). The predominant genetic risk for developing T1D, the immune-mediated form of diabetes, is conferred through HLA genes. One such gene, termed HLA-DQ8, is present in 50-60% of patients with T1D and those at-risk. DQ8 presents disease-relevant peptides to T cells, which mediate tissue-specific destruction of pancreatic islets. Using a structure-based approach to evaluate the 'druggability' of the DQ8 molecule, methyldopa, a clinically well-established oral antihypertensive agent, was discovered to bind DQ8. Methyldopa blocked the activation of DQ8-specific T cells responding to self-antigens such as insulin but not influenza. In a proof-of-concept clinical trial (NCT01883804), methyldopa was administered to recent-onset T1D patients with the DQ8 gene that confirmed the mechanism of action and diminished inflammatory T cell responses toward insulin. Methyldopa blocks the diabetes-specific function of HLA-DQ8, which represents a personalized medicine approach to treat the underlying autoimmunity in T1D. Clinical trials are warranted and underway to evaluate methyldopa in potentially preserving residual β-cell function in those with new onset and at risk for T1D.

Soluble antigen arrays disarm antigen-specific B cells to promote lasting immune tolerance in experimental autoimmune encephalomyelitis

Autoreactive lymphocytes that escape central immune tolerance may be silenced via an endogenous peripheral tolerance mechanism known as anergy. Antigen-specific therapies capable of inducing anergy may restore patients with autoimmune diseases to a healthy phenotype while avoiding deleterious side effects associated with global immunosuppression. Inducing anergy in B cells may be a particularly potent intervention, as B cells can contribute to autoimmune diseases through multiple mechanisms and offer the potential for direct antigen-specific targeting through the B cell receptor (BCR). Our previous results suggested autoreactive B cells may be silenced by multivalent ‘soluble antigen arrays’ (SAgAs), which are polymer conjugates displaying multiple copies of autoantigen with or without a secondary peptide that blocks intracellular cell-adhesion molecule-1 (ICAM-1). Here, key therapeutic molecular properties of SAgAs were identified and linked to the immunological mechanism through comprehensive cellular and in vivo analyses. We determined non-hydrolyzable ‘cSAgAs’ displaying multivalent ‘click’-conjugated antigen more potently suppressed experimental autoimmune encephalomyelitis (EAE) compared to hydrolyzable SAgAs capable of releasing conjugated antigen. cSAgAs restored a healthy phenotype in disease-specific antigen presenting cells (APCs) by inducing an anergic response in B cells and a subset of B cells called autoimmune-associated B cells (ABCs) that act as potent APCs in autoimmune disease. Accompanied by a cytokine response skewed towards a Th2/regulatory phenotype, this generated an environment of autoantigenic tolerance. By identifying key therapeutic molecular properties and an immunological mechanism that drives SAgA efficacy, this work guides the design of antigen-specific immunotherapies capable of inducing anergy.

Proteomic approach to profiling immune complex antigens in cerebrospinal fluid samples from patients with central nervous system autoimmune diseases

BackgroundImmune complexes (ICs) may clearly reflect immunological abnormalities caused by disease, especially for autoimmune diseases. Although ICs have been detected in cerebrospinal fluid (CSF) from patients with CNS autoimmune diseases, identities of antigens in such ICs have not been comprehensively determined. MethodsWe used immune complexome analysis, in which nano-liquid chromatography-tandem mass spectrometry is employed to comprehensively identify antigens incorporated into ICs in biological fluids, to characterize ICs in CSF samples from patients with CNS autoimmune diseases, and to find disease-specific IC antigen to a certain CNS autoimmune disease. Also, we compared the IC antigens we identified with the reported CSF proteome or with the published plasma proteome to examine if the method is distinguished from the conventional CSF proteome analysis. ResultsWe identified 176 antigens in 78 CSF samples. We then assessed the overlaps among these antigens, the CSF proteome, and the plasma proteome; 140 of the 176 antigens were found to be exclusively detected by our method. Notably, IC-associated suprabasin in CSF was 100% specific to neuropsychiatric systemic lupus erythematosus (NPSLE). ConclusionsThis report is the first to comprehensively identify the antigens incorporated into ICs in CSF. There was limited overlap between the antigens we identified and the CSF proteome or the plasma proteome; therefore, our method can be distinguished from the conventional CSF proteome analysis. Although the sensitivity of disease-specific IC-antigens detected in immune complexome analysis screening, the sensitivity may be improved by developing an ELISA method specifically for detecting the ICs. Immune complexome analysis of CSF may be a new and promising path to biomarker discovery for diagnosis and study for CNS autoimmune diseases.

Cross-phenotype analysis of Immunochip data identifies KDM4C as a relevant locus for the development of systemic vasculitis

ObjetiveSystemic vasculitides represent a heterogeneous group of rare complex diseases of the blood vessels with a poorly understood aetiology. To investigate the shared genetic component underlying their predisposition, we performed...

Evolution of the magic bullet: Single chain antibody fragments for the targeted delivery of immunomodulatory proteins.

Immunocytokines are fusion proteins that combine the specific antigen binding capacities of an antibody or derivative thereof and the potent bioactivity of a cytokine partner. These novel biopharmaceuticals have been directed to various targets of oncological as well as non-oncological origin and a handful of promising constructs are currently advancing in the clinical trial pipeline. Several factors such as the choice of a disease specific antigen, the antibody format and the modulatory nature of the payload are crucial, not only for therapeutic efficacy and safety but also for the commercial success of such a product. In this review, we provide an overview of the basic principles and obstacles in immunocytokine design with a specific focus on single chain antibody fragment-based constructs that employ interleukins as the immunoactive component. Impact statement Selective activation of the immune system in a variety of malignancies represents an attractive approach when existing strategies have failed to provide adequate treatment options. Immunocytokines as a novel class of bifunctional protein therapeutics have emerged recently and generated promising results in preclinical and clinical studies. In order to harness their full potential, multiple different aspects have to be taken into consideration. Several key points of these fusion constructs are discussed here and should provide an outline for the development of novel products based on an overview of selected formats.

Carbon nanotubes based biosensor for detection of cancer antigens (CA-125) under shear flow condition

The detection and determination of the cancer biomarkers is very crucial to diagnose at the early stages. We implement to design the biosensor which can improve the detection by reducing the process time, cost and space. The research approach mainly is focused on developing a surface modification protocol for better sensitivity under the shear flow rate conditions using carbon nanotubes (CNTs). The interdigitated electrode transducer was modified using functionalized CNTs for signal enhancement. The biosensor was integrated with PDMS microfluidic channels for controlled self-driven flow. In this article, the role and functionality of the CNTs for signal enhancement are summarized. The experimental results provide the evidence of disease-specific antigen (CA-125) detection from a micro volume of biofluid sample using the CNTs modified interdigitated electrodes under capillary flow condition.

In situ programming of leukaemia-specific T cells using synthetic DNA nanocarriers.

An emerging approach for treating cancer involves programming patient-derived T cells with genes encoding disease-specific chimeric antigen receptors (CARs), so that they can combat tumour cells once they are reinfused. Although trials of this therapy have produced impressive results, the in vitro methods they require to generate large numbers of tumour-specific T cells are too elaborate for widespread application to treat cancer patients. Here, we describe a method to quickly program circulating T cells with tumour-recognizing capabilities, thus avoiding these complications. Specifically, we demonstrate that DNA-carrying nanoparticles can efficiently introduce leukaemia-targeting CAR genes into T-cell nuclei, thereby bringing about long-term disease remission. These polymer nanoparticles are easy to manufacture in a stable form, which simplifies storage and reduces cost. Our technology may therefore provide a practical, broadly applicable treatment that can generate anti-tumour immunity 'on demand' for oncologists in a variety of settings.

Abstract 3133: Glypican-1 is a potential marker of prognosis and involved in chemoresistance of cisplatin in esophageal squamous cell cancer

Abstract [Introduction]Despite recent improvements in multi-treatment approaches, including surgery, radiotherapy and chemotherapy, the prognosis for patients with Esophageal squamous cell cancer (ESCC) remains unsatisfactory. In addition, predicting a prognosis and treatment response for ESCC had been constantly being required but remains difficult, even when using the TNM system. We identified Glypican-1(GPC1) as a novel disease specific antigen of ESCC by quantitative proteomic analysis focused on cell surface membrane proteins. GPC1 is one of the cell-surface heparan sulfate proteoglycans which has been reported as a prognostic factor in pancreatic cancer. [Objectives]This study aimed to clarify correlations between GPC1 expression and the prognosis in patient with ESCC and also reveal the association of GPC1 with the drug resistance to chemotherapeutic agents in squamous cancer cell lines and esophageal cancer patients treated with chemotherapy. [Methods]ESCC tissues were obtained from 175 patients who underwent esophagectomy at Osaka University Hospital(Osaka, Japan), from 2001 to 2013 and were subjected to immunohistochemistry. We performed immunohistochemical (IHC) assessment using anti-GPC1 antibody and investigated the association between GPC1 expression and the clinicopathological parameters. We divided the patients into two groups such as high expression group (HG) and low expression group (LG). Furthermore, we investigated associations between GPC1 expression and clinical response of neo-adjuvant chemotherapy for patients with ESCC. We established GPC1 knocked out cells of TE-14 using Crisper-Cas9 system and stable GPC1-transfected cells of LK-2(Lung squamous cell carcinoma cell line) using pcDNA3.1-GPC1 plasmid transfection. To investigate the relationship between GPC1 expression and sensitivity to cisplatin (CDDP), we examined sensitivity to CDDP by the WST-8 assay. [Results]The patients characteristics as follows, age (median) = 64yrs(39-80), gender (Male: Female) = 155:20. By the IHC analysis, 99 patients were classified into HG and 76 were LG. HG patients exhibited a poorer prognosis compared with the LG patients. (p<0.001) In multivariate analyses pT, pN, and GPC1 high expression were became significant prognostic factors (p<0.01). We clarified there was a significant negative correlation between histological responses and GPC1 expression in patient with neoadjuvant chemo therapy. Furthermore, in vitro experiments showed that the IC50 values for CDDP of GPC1 expressing cells (TE-14 and GPC1 transfected LK-2) were significantly higher than that of GPC1 no-expressing cells (GPC1 knocked out TE-14 and LK-2). [Conclusion]Our study showed that GPC1 was an independent prognostic factor in esophageal cancer. Furthermore, we showed that GPC1 was a critical molecule for chemoresistance to cisplatin. Citation Format: Takahiko Nishigaki, Tsuyoshi Takahashi, Satishi Serada, Minoru Fujimoto, Hisashi Hara, Yasuhiro Miyazaki, Tomoki Makino, Yukinori Kurokawa, Makoto Yamasaki, Kiyokazu Nakajima, Shuji Takiguchi, Masaki Mori, Yuichiro Doki, Tetsuji Naka. Glypican-1 is a potential marker of prognosis and involved in chemoresistance of cisplatin in esophageal squamous cell cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3133.

Advancement and applications of peptide phage display technology in biomedical science.

Combinatorial phage library is a powerful research tool for high-throughput screening of protein interactions. Of all available molecular display techniques, phage display has proven to be the most popular approach. Screening phage-displayed random peptide libraries is an effective means of identifying peptides that can bind target molecules and regulate their function. Phage-displayed peptide libraries can be used for (i) B-cell and T-cell epitope mapping, (ii) selection of bioactive peptides bound to receptors or proteins, disease-specific antigen mimics, peptides bound to non-protein targets, cell-specific peptides, or organ-specific peptides, and (iii) development of peptide-mediated drug delivery systems and other applications. Targeting peptides identified using phage display technology may be useful for basic research and translational medicine. In this review article, we summarize the latest technological advancements in the application of phage-displayed peptide libraries to applied biomedical sciences.

Sexual dimorphism in an animal model of Sjögren's syndrome: a potential role for Th17 cells.

ABSTRACTSjögren's syndrome is a complex autoimmune disease with an array of diverse immunological, genetic and environmental etiologies, making identification of the precise autoimmune mechanism difficult to define. One of the most distinctive aspects of Sjögren's syndrome is the high sexual dimorphism with women affected 10-20 times more than men. It is nearly impossible to study the sexual dimorphic development of Sjögren's syndrome in human patients; therefore it is pertinent to develop an appropriate animal model which resembles human disease. The data indicated that female C57BL/6.NOD-Aec1Aec2 mice developed an earlier onset of sialadenitis with a higher composition of CD3+ T cells and a 10-fold increase in glandular infiltration of Th17 cells at the onset of clinical disease compared to male mice. Inflammatory Th17 cells of female mice exhibited a stronger proliferation in response to disease-specific antigen than their male counterpart. At the clinical disease stage, altered autoantibody patterns can be detected in females whereas they are seldom observed in male C57BL/6.NOD-Aec1Aec2 mice. Interestingly, male C57BL/6.NOD-Aec1Aec2 mice developed an earlier loss of secretory function, despite the fact that female C57BL/6.NOD-Aec1Aec2 mice exhibited a more rapid secretory loss. This data indicates the strong sexual dimorphism in the SjS-susceptible C57BL/6.NOD-Aec1Aec2 animal model, making it an appropriate animal model to examine human disease.

Transglutaminase 6 Antibodies in the Serum of Patients With Amyotrophic Lateral Sclerosis.

Celiac disease is an autoimmune disorder triggered by gluten in genetically predisposed individuals. Gluten sensitivity can cause neurologic manifestations, such as ataxia or neuropathy, with or without gastrointestinal symptoms. Many patients with gluten ataxia produce antibodies toward the newly identified neuronal transglutaminase 6 (TG6). Two case reports described patients initially diagnosed with amyotrophic lateral sclerosis (ALS) and ultimately with celiac disease who improved with a strict gluten-free diet. To evaluate the prevalence of celiac disease-related antibodies and HLA antigen alleles, as well as TG6 antibodies, in patients with ALS and healthy individuals serving as controls to determine whether a neurologic presentation of a gluten-related disorder mimicking ALS might occur in some patients. In a case-control study conducted in an ALS tertiary center, we measured serum levels of total IgA antibodies, IgA antibodies to transglutaminase 2 (TG2) and endomysium, as well as IgA and IgG antibodies to deamidated gliadine peptide and TG6 and performed HLA antigen genotyping in 150 consecutive patients with ALS and 115 healthy volunteers of similar age and sex. Participants did not have any known autoimmune or gastroenterologic disorder and were not receiving any immunomodulatory medications. The study was conducted from July 1, 2010, to December 31, 2012. Antibody levels and frequency of individuals with abnormal antibody values as well as frequency of HLA antigen alleles were compared between patient and control groups. All patients and control group participants were seronegative to IgA antibodies to TG2, endomysium, and deamidated gliadine peptide. Twenty-three patients (15.3%) were seropositive to TG6 IgA antibodies as opposed to only 5 controls (4.3%) (P = .004). The patients seropositive for TG6 showed a classic picture of ALS, similar to that of seronegative patients. Fifty patients and 20 controls were tested for celiac disease-specific HLA antigen alleles; 13 of 22 TG6 IgA seropositive individuals (59.1%) were seropositive for celiac disease-related alleles compared with 8 (28.6%) of the 28 seronegative individuals (P = .04). Mean (SD) levels of IgA antibodies to TG2 were 1.78 (0.73) in patients and 1.58 (0.68) in controls (normal, <10). In a subset of study participants, mean levels of deamidated gliadin peptide autoantibodies were 7.46 (6.92) in patients and 6.08 (3.90) in controls (normal, <16). Mean levels of IgA antibodies to TG6 were 29.3 (30.1) in patients and 21.0 (27.4) in controls (P = .02; normal, <26). The data from this study indicate that, in certain cases, an ALS syndrome might be associated with autoimmunity and gluten sensitivity. Although the data are preliminary and need replication, gluten sensitivity is potentially treatable; therefore, this diagnostic challenge should not be overlooked.

Deriving benefit of early detection from biomarker-based prognostic models.

Many prognostic models for cancer use biomarkers that have utility in early detection. For example, in prostate cancer, models predicting disease-specific survival use serum prostate-specific antigen levels. These models typically show that higher marker levels are associated with poorer prognosis. Consequently, they are often interpreted as indicating that detecting disease at a lower threshold of the biomarker is likely to generate a survival benefit. However, lowering the threshold of the biomarker is tantamount to early detection. For survival benefit to not be simply an artifact of starting the survival clock earlier, we must account for the lead time of early detection. It is not known whether the existing prognostic models imply a survival benefit under early detection once lead time has been accounted for. In this article, we investigate survival benefit implied by prognostic models where the predictor(s) of disease-specific survival are age and/or biomarker level at disease detection. We show that the benefit depends on the rate of biomarker change, the lead time, and the biomarker level at the original date of diagnosis as well as on the parameters of the prognostic model. Even if the prognostic model indicates that lowering the threshold of the biomarker is associated with longer disease-specific survival, this does not necessarily imply that early detection will confer an extension of life expectancy.

Identification of HnRNP-A2/B1 as a Target Antigen of Anti-Endothelial Cell IgA Antibody in Behçet's Disease

Behçet's disease (BD) is a chronic, multisystemic vasculitis that theoretically affects all sizes and types of blood vessels. Although pathogenesis remains enigmatic, endothelial cells are believed to be the primary target in this disease. We detected the target protein using western blotting and immunoprecipitation and determined the amino-acid sequence of the peptide by liquid chromatography-matrix assisted laser desorption/ionization-tandem time-of-flight analysis (LC-MALDI-TOF/TOF). Serum reactivity against the recombinant target protein was analyzed by immunoblotting. Serum reactivity against streptococcal 65-kD heat shock protein (hsp-65) and the recombinant target protein was investigated by ELISA. The 36-40-kD protein band that was obtained from immunoprecipitation, which was analyzed by LC-MALDI-TOF/TOF, exhibited the amino-acid sequences of heterogeneous nuclear ribonucleoproteins A2/B1 (hnRNP-A2/B1). Reactivity of serum IgA against human recombinant hnRNP-A2/B1 was detected in 25 of 30 BD patients (83.3%), 4 of 30 systemic lupus erythematosus patients (13.3%), 8 of 30 rheumatoid arthritis patients (26.7%), 9 of 30 Takayasu's arteritis patients (30%), 6 of 30 healthy controls (20%), and none of 30 IgA nephropathy patients. Optical densities obtained from ELISAs against the recombinant human hnRNP-A2/B1 were correlated with those against the recombinant streptococcal hsp-65.JID JOURNAL CLUB ARTICLE: For questions, answers, and open discussion about this article, please go to http://www.nature.com/jid/journalclub.

The Application of CD Antigen Proteomics to Pharmacogenomics

The advent of multiplexing technologies has raised the possibility that disease states can be defined using discrete genomic and proteomic patterns or signatures. However, this emerging area has been limited by the 'content problem', arising from the uncertainty of which molecules to focus on. The human cluster of differentiation (CD) antigens are expressed on cells of the human immune system (leukocytes) and on other cell types. These heterogeneous molecules perform a host of roles essential to immune function and to the physiology of other lineages. The 339 defined CD antigens and their, as yet, undefined counterparts constitute key components of the expressed human cell surface proteome. We propose that CD antigen expression patterns will form the basis of a rational, discrete and generalized diagnostic and prognostic system. Furthermore, disease-specific CD antigen proteomic signatures are likely to be more robust than corresponding genomic signatures and will also help to identify molecular targets for therapeutic intervention.