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Discover how PEPperPRINT Peptide Microarray products have been used in different fields of research.

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Anti-PfGARP activates programmed cell death of parasites and reduces severe malaria

Raj, Dipak K.; Das Mohapatra, Alok; Jnawali, Anup; Zuromski, Jenna; Jha, Ambrish; Cham-Kpu, Gerald; Sherman, Brett; Rudlaff, Rachel M.; Nixon, Christina E.; Hilton, Nicholas; Oleinikov, Andrew V.; Chesnokov, Olga; Merritt, Jordan; Pond-Tor, Sunthorn; Burns, Lauren; Jolly, Grant; Ben Mamoun, Choukri; Kabyemela, Edward; Muehlenbachs, Atis; Lambert, Lynn; Orr-Gonzalez, Sachy; Gnädig, Nina F.; Fidock, David A.; Park, Sangshin; Dvorin, Jeffrey D.; Pardi, Norbert; Weissman, Drew; Mui, Barbara L.; Tam, Ying K.; Friedman, Jennifer F.; Fried, Michal; Duffy, Patrick E.; Kurtis, Jonathan D.
Nature.
Apr 2020
10.1038/s41586-020-2220-1
Malaria caused by Plasmodium falciparum remains the leading single-agent cause of mortality in children1, yet the promise of an effective vaccine has not been fulfilled. Here, using our previously described differential screening method to analyse the proteome of blood-stage P. falciparum parasites2, we identify P. falciparum glutamic-acid-rich protein (PfGARP) as a parasite antigen that is recognized by antibodies in the plasma of children who are relatively resistant—but not those who are susceptible—to malaria caused by P. falciparum. PfGARP is a parasite antigen of 80 kDa that is expressed on the exofacial surface of erythrocytes infected by early-to-late-trophozoite-stage parasites. We demonstrate that antibodies against PfGARP kill trophozoite-infected erythrocytes in culture by inducing programmed cell death in the parasites, and that vaccinating non-human primates with PfGARP partially protects against a challenge with P. falciparum. Furthermore, our longitudinal cohort studies showed that, compared to individuals who had naturally occurring anti-PfGARP antibodies, Tanzanian children without anti-PfGARP antibodies had a 2.5-fold-higher risk of severe malaria and Kenyan adolescents and adults without these antibodies had a twofold-higher parasite density. By killing trophozoite-infected erythrocytes, PfGARP could synergize with other vaccines that target parasite invasion of hepatocytes or the invasion of and egress from erythrocytes.

On‐Chip Neo‐Glycopeptide Synthesis for Multivalent Glycan Presentation

Mende, Marco; Tsouka, Alexandra; Heidepriem, Jasmin; Paris, Grigori; Mattes, Daniela S.; Eickelmann, Stephan; Bordoni, Vittorio; Wawrzinek, Robert; Fuchsberger, Felix F.; Seeberger, Peter H.; Rademacher, Christoph; Delbianco, Martina; Mallagaray, Alvaro; Loeffler, Felix F
Chem. Eur. J..
Apr 2020
10.1002/chem.202001291
Single glycan–protein interactions are often weak, such that glycan binding partners commonly utilize multiple, spatially defined binding sites to enhance binding avidity and specificity. Current array technologies usually neglect defined multivalent display. Laser-based array synthesis technology allows for flexible and rapid on-surface synthesis of different peptides. By combining this technique with click chemistry, neo-glycopeptides were produced directly on a functionalized glass slide in the microarray format. Density and spatial distribution of carbohydrates can be tuned, resulting in well-defined glycan structures for multivalent display. The two lectins concanavalin A and langerin were probed with different glycans on multivalent scaffolds, revealing strong spacing-, density-, and ligand-dependent binding. In addition, we could also measure the surface dissociation constant. This approach allows for a rapid generation, screening, and optimization of a multitude of multivalent scaffolds for glycan binding.

Autoantibody Signature in Cardiac Arrest

Maguy, Ange; Tardif, Jean-Claude; Busseuil, David; Ribi, Camillo; Li, Jin
Circulation.
Apr 2020
10.1161/CIRCULATIONAHA.119.044408
Background: Cardiac arrest is a tragic event that causes one death roughly every 90 seconds worldwide. Survivors generally undergo a work-up to identify the etiology of arrest. However, 5-10% of cardiac arrest remain unexplained. As cardiac arrhythmias mostly underlie cardiac arrest and increasing evidence strongly supports the involvement of autoantibodies in arrhythmogenesis, a large-panel autoantibody screening was performed in cardiac arrest patients. Methods: This is an observational, cross-sectional study of patients from the Montreal Heart Institute (MHI) hospital cohort, a single center registry of participants. A peptide microarray was designed to screen for IgG targeting epitopes from all known cardiac ion channels with extracellular domains. Plasma samples from 23 patients with unexplained cardiac arrest were compared to 22 cardiac arrest cases of ischemic origin and a group of 29 age-, sex- and BMI-matched healthy subjects. The false discovery rate (FDR), LASSO logistic regression and random forest methods were jointly carried out to find significant differential IgG responses. Results: The autoantibody against the pore domain of the L-type voltage-gated calcium channel (Ca v 1.2) was consistently identified as a biomarker of idiopathic cardiac arrest (P=0.002, FDR=0.007, classification accuracies ≥0.83). Functional studies on human induced pluripotent stem cell-derived cardiomyocytes demonstrated that the anti-Ca v 1.2 IgG purified from patients with idiopathic cardiac arrest is proarrhythmogenic by reducing the action potential duration through calcium channel inhibition. Conclusions: The present report addresses the concept of autoimmunity and cardiac arrest. Hitherto unknown autoantibodies targeting extracellular sequences of cardiac ion channels were detected. Moreover, the study identified an autoantibody signature specific to patients with cardiac arrest.

Plasmodium Falciparum and Plasmodium Vivax Vaccine

Werner, Ekkehard
Apr 2020
The present invention relates to a vaccine V comprising (A) at least one isolated polypeptide strand P comprising or consisting of at least nine consecutive amino acid moieties of the repetitive organellar protein, putative of Plasmodium falciparum or the hypothetical protein PVNG_04523 of Plasmodium vivax or a polynucleotide strand encoding for such polypeptide; and (B) at least one pharmaceutically acceptable carrier or excipient. Furthermore, the present invention refers to an antibody binding to the repetitive organellar protein,putative of Plasmodium falciparumor the hypothetical protein PVNG_04523 of Plasmodium vivax or a polynucleotide strand encoding therefor, to a method of generating such antibody and uses thereof.

Distinct early IgA profile may determine severity of COVID-19 symptoms: an immunological case series

Dahlke, Christine; Heidepriem, Jasmin; Kobbe, Robin; Santer, Rene; Koch, Till; Fathi, Anahita; Ly, My L.; Schmiedel, Stefan; Seeberger, Peter H.; ID-UKE COVID-19 study group; Addo, Marylyn M.; Loeffler, Felix F.
Apr 2020
10.1101/2020.04.14.20059733
SARS-CoV-2 is the causative agent of COVID-19 and is a severe threat to global health. Patients infected with SARS-CoV-2 show a wide range of symptoms and disease severity, while limited data is available on its immunogenicity. Here, the kinetics of the development of SARS-CoV-2-specific antibody responses in relation to clinical features and dynamics of specific B-cell populations are reported. Immunophenotyping of B cells was performed by flow cytometry with longitudinally collected PBMCs. In parallel, serum samples were analyzed for the presence of SARS-CoV-2-specific IgA, IgG, and IgM antibodies using whole proteome peptide microarrays. Soon after disease onset in a mild case, we observed an increased frequency of plasmablasts concomitantly with a strong SARS-CoV-2-specific IgA response. In contrast, a case with more severe progression showed a delayed, but eventually very strong and broad SARS-CoV-2-specific IgA response. This case study shows that determining SARS-CoV-2-specific antibody epitopes can be valuable to monitor the specificity and magnitude of the early B-cell response, which could guide the development of vaccine candidates. Follow-up studies are required to evaluate whether the kinetics and strength of the SARS-CoV-2-specific IgA response could be potential prognostic markers of viral control.

Mapping the epitopes of Schistosoma japonicum esophageal gland proteins for incorporation into vaccine constructs

Li, Xiao-Hong; Vance, Gillian M.; Cartwright, Jared; Cao, Jian-Ping; Wilson, R Alan; Castro-Borges, William
PLoS ONE.
Feb 2020
10.1371/journal.pone.0229542
Background The development of a schistosome vaccine has proved challenging but we have suggested that characterisation of the self-cure mechanism in rhesus macaques might provide a route to an effective product. The schistosome esophagus is a complex structure where blood processing is initiated by secretions from anterior and posterior glands, achieved by a mixture of ~40 unique proteins. The mechanism of self-cure in macaques involves cessation of feeding, after which worms slowly starve to death. Antibody coats the esophagus lumen and disrupts the secretory processes from the glands, potentially making their secretions ideal vaccine targets. Methodology/Principal findings We have designed three peptide arrays comprising overlapping 15-mer peptides encompassing 32 esophageal gland proteins, and screened them for reactivity against 22-week infection serum from macaques versus permissive rabbit and mouse hosts. There was considerable intra- and inter-species variation in response and no obvious unique target was associated with self-cure status, which suggests that self-cure is achieved by antibodies reacting with multiple targets. Some immuno-dominant sequences/regions were evident across species, notably including: MEGs 4.1C, 4.2, and 11 (Array 1); MEG-12 and Aspartyl protease (Array 2); a Tetraspanin 1 loop and MEG-n2 (Array 3). Responses to MEGs 8.1C and 8.2C were largely confined to macaques. As proof of principle, three synthetic genes were designed, comprising several key targets from each array. One of these was expressed as a recombinant protein and used to vaccinate rabbits. Higher antibody titres were obtained to the majority of reactive regions than those elicited after prolonged infection. Conclusions/Significance It is feasible to test simultaneously the additive potential of multiple esophageal proteins to induce protection by combining their most reactive regions in artificial constructs that can be used to vaccinate suitable hosts. The efficacy of the approach to disrupt esophageal function now needs to be tested by a parasite challenge.

Immunization with full-length Plasmodium falciparum merozoite surface protein 1 is safe and elicits functional cytophilic antibodies in a randomized first-in-human trial

Blank, Antje; Fürle, Kristin; Jäschke, Anja; Mikus, Gerd; Lehmann, Monika; Hüsing, Johannes; Heiss, Kirsten; Giese, Thomas; Carter, Darrick; Böhnlein, Ernst; Lanzer, Michael; Haefeli, Walter E.; Bujard, Hermann
npj Vaccines.
Jan 2020
10.1038/s41541-020-0160-2
A vaccine remains a priority in the global fight against malaria. Here, we report on a single-center, randomized, double-blind, placebo and adjuvant-controlled, dose escalation phase 1a safety and immunogenicity clinical trial of full-length Plasmodium falciparum merozoite surface protein 1 (MSP1) in combination with GLA-SE adjuvant. Thirty-two healthy volunteers were vaccinated at least three times with MSP1 plus adjuvant, adjuvant alone, or placebo (24:4:4) to evaluate the safety and immunogenicity. MSP1 was safe, well tolerated and immunogenic, with all vaccinees sero-converting independent of the dose. The MSP1-specific IgG and IgM titers persisted above levels found in malaria semi-immune humans for at least 6 months after the last immunization. The antibodies were variant- and strain-transcending and stimulated respiratory activity in granulocytes. Furthermore, full-length MSP1 induced memory T-cells. Our findings encourage challenge studies as the next step to evaluate the efficacy of full-length MSP1 as a vaccine candidate against falciparum malaria (EudraCT 2016-002463-33).

Molecular mimicry, genetic homology, and gene sharing proteomic “molecular fingerprints” using an EBV (Epstein-Barr virus)-derived microarray as a potential diagnostic method in autoimmune disease

Dreyfus, David H.; Farina, Antonella; Farina, Giuseppina Alessandra
Immunol Res.
Dec 2018
10.1007/s12026-018-9045-0
EBV (Epstein-Barr Virus) and other human DNA viruses are associated with autoimmune syndromes in epidemiologic studies. In this work, immunoglobulin G response to EBV-encoded proteins which share regions with human immune response proteins from the human host including ZEBRA (BZLF-1 encoded protein), BALF-2 recombinase expressed primarily during the viral lytic replication cycle, and EBNA-1 (Epstein-Barr Virus Nuclear Antigen) expressed during the viral latency cycle respectively were characterized using a laser-printed micro-array (PEPperprint.com). IgG response to conserved “A/T hooks” in EBV-encoded proteins such as EBNA-1 and the BALF-2 recombinase related to host DNA-binding proteins including RAG-1 recombinase and histones, and EBV-encoded virokines such as the IL-10 homologue BCRF-1 suggest further directions for clinical research. The author suggests that proteomic “molecular fingerprints” of the immune response to viral proteins shared with human immune response genes are potentially useful in early diagnosis and monitoring of autoantibody production and response to therapy in EBV-related autoimmune syndromes.

Soybean Allergy Related Epitopes

Kern, Karolin; Spiegel, Holger; Havenith, Heide; Szardenings, Michael
Nov 2018
The invention relates to a compilation comprising at least five different peptides, each peptide comprising at least one sequence element corresponding to an epitope selected from the group consisting of SEQ ID NO.: 1-354, wherein at least five different epitopes are represented. The invention further relates to an in vitro method for determining a patient’s immune status to soybean allergens, to a method for detecting at least one soybean allergen in a substance and to a method for determining the allergenicity of a soybean variety. Additionally, the invention relates to a kit comprising at least one composition containing a compound comprising at least five different sequence elements each corresponding to an epitope selected from the group consisting of SEQ ID NO.: 1-354, wherein at least five different epitopes are represented. Furthermore, the invention relates to the use of a peptide comprising a sequence element corresponding to an epitope for providing a molecule binding to a protein or peptide comprising the epitope.

Linear epitopes in Onchocerca volvulus vaccine candidate proteins and excretory-secretory proteins

Lagatie, Ole; Verheyen, Ann; Van Dorst, Bieke; Batsa Debrah, Linda; Debrah, Alex; Stuyver, Lieven J.
Parasite Immunol.
Nov 2018
10.1111/pim.12587
In our previous study, a proteome-wide screen was conducted to identify linear epitopes in this parasite’s proteome, resulting in the discovery of three immunodominant motifs. Here, we investigated whether such antigenic peptides were found in proteins that were already known as vaccine candidates and excretome/secretome proteins for Onchocerca volvulus This approach led to the identification of 71 immunoreactive stretches in 46 proteins. A deep-dive into the immunoreactivity profiles of eight vaccine candidates that were chosen as most promising candidates for further development (Ov-CPI-2, Ov-ALT-1, Ov-RAL-2, Ov-ASP-1, Ov-103, Ov-RBP-1, Ov-CHI-1, and Ov-B20), resulted in the identification of a poly-glutamine stretch in Ov-RAL-2 that has properties for use as a serodiagnostic marker for O. volvulus infection. A peptide ELISA was developed, and the performance of this assay was evaluated. Based on this assessment, it was found that this assay has a sensitivity of 75.0% [95% CI: 64.9%-83.5%] and a specificity of 98.5% [95% CI: 94.6%-99.8%]. Furthermore, 8.7% reactivity in Asian parasite-infected individuals (8 out of 92) was observed. Besides this identification of a linear epitope marker, the information on the presence of linear epitopes in vaccine candidate proteins might be useful in the study of vaccines for river blindness.

Potential therapeutic antibodies targeting specific adiponectin isoforms in rheumatoid arthritis

Lee, Yeon-Ah; Hahm, Dae-Hyun; Kim, Jung Yeon; Sur, Bonjun; Lee, Hyun Min; Ryu, Chun Jeih; Yang, Hyung-In; Kim, Kyoung Soo
Arthritis Res Ther.
Oct 2018
10.1186/s13075-018-1736-3
Background Different adiponectin isoforms appear to be differentially involved in the pathogenesis of various diseases. The purpose of this study was to generate monoclonal antibodies (mAbs) specific to different adiponectin isoforms and investigate whether these mAbs have potential as therapeutic agents for such diseases. Methods Hybridoma cells producing monoclonal antibodies were generated and screened using enzyme-linked immunosorbent assay and Western blotting for the production of mAbs recognizing human adiponectin isoforms. Results The mAb from hybridoma clone KH7–41 recognized both the middle molecular weight (MMW) (hexamer) and low molecular weight (LMW) (trimer) isoforms of adiponectin in human serum, whereas the KH7–33 mAb detected only MMW (hexamer) adiponectin. The KH4–8 clone recognized both the high molecular weight (HMW) (multimer) and MMW adiponectin isoforms. However, in mouse and rat sera, the abovementioned antibodies recognized only the MMW isomer. These mAbs also recognized adiponectin in various human tissues, such as lung, kidney, and adipose tissues, although the three mAbs had different staining intensities. The mAb from clone KH4–8 effectively inhibited increases in interleukin-6 (IL-6) and IL-8 expression in recombinant adiponectin-stimulated human osteoblasts and human umbilical vein endothelial cells. Also, the mAbs KH7–33 and KH4–8 significantly ameliorated rheumatic symptoms in a collagen-induced arthritis mouse model. This result suggests that these mAb treatments may ameliorate adiponectin-mediated inflammatory response. Conclusions mAbs against human adiponectin isomers can potentially be developed as therapeutic antibodies to target specific detrimental isoforms of adiponectin while maintaining the functions of beneficial isoforms.

Cytotoxic anti-circumsporozoite antibodies target malaria sporozoites in the host skin

Aliprandini, Eduardo; Tavares, Joana; Panatieri, Raquel Hoffmann; Thiberge, Sabine; Yamamoto, Marcio Massao; Silvie, Olivier; Ishino, Tomoko; Yuda, Masao; Dartevelle, Sylvie; Traincard, François; Boscardin, Silvia Beatriz; Amino, Rogerio
Nat Microbiol.
Oct 2018
10.1038/s41564-018-0254-z
The circumsporozoite protein (CSP) is the major surface protein of malaria sporozoites (SPZs), the motile and invasive parasite stage inoculated in the host skin by infected mosquitoes. Antibodies against the central CSP repeats of different plasmodial species are known to block SPZ infectivity, but the precise mechanism by which these effectors operate is not completely understood. Here, using a rodent Plasmodium yoelii malaria model, we show that sterile protection mediated by anti-P. yoelii CSP humoral immunity depends on the parasite inoculation into the host skin, where antibodies inhibit motility and kill P. yoelii SPZs via a characteristic ‘dotty death’ phenotype. Passive transfer of an anti-repeat monoclonal antibody (mAb) recapitulates the skin inoculation-dependent protection, in a complement- and Fc receptor γ-independent manner. This purified mAb also decreases motility and, notably, induces the dotty death of P. yoelii SPZs in vitro. Cytotoxicity is species-transcendent since cognate anti-CSP repeat mAbs also kill Plasmodium berghei and Plasmodium falciparum SPZs. mAb cytotoxicity requires the actomyosin motor-dependent translocation and stripping of the protective CSP surface coat, rendering the parasite membrane susceptible to the SPZ pore-forming-like protein secreted to wound and traverse the host cell membrane. The loss of SPZ fitness caused by anti-P. yoelii CSP repeat antibodies is thus a dynamic process initiated in the host skin where SPZs either stop moving, or migrate and traverse cells to progress through the host tissues at the eventual expense of their own life.

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