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1.
Proc Natl Acad Sci U S A ; 118(37)2021 09 14.
Article in English | MEDLINE | ID: covidwho-1768982

ABSTRACT

Adaptive immune recognition is mediated by the binding of peptide-human leukocyte antigen complexes by T cells. Positive selection of T cells in the thymus is a fundamental step in the generation of a responding T cell repertoire: only those T cells survive that recognize human peptides presented on the surface of cortical thymic epithelial cells. We propose that while this step is essential for optimal immune function, the process results in a defective T cell repertoire because it is mediated by self-peptides. To test our hypothesis, we focused on amino acid motifs of peptides in contact with T cell receptors. We found that motifs rarely or not found in the human proteome are unlikely to be recognized by the immune system just like the ones that are not expressed in cortical thymic epithelial cells or not presented on their surface. Peptides carrying such motifs were especially dissimilar to human proteins. Importantly, we present our main findings on two independent T cell activation datasets and directly demonstrate the absence of naïve T cells in the repertoire of healthy individuals. We also show that T cell cross-reactivity is unable to compensate for the absence of positively selected T cells. Additionally, we show that the proposed mechanism could influence the risk for different infectious diseases. In sum, our results suggest a side effect of T cell positive selection, which could explain the nonresponsiveness to many nonself peptides and could improve the understanding of adaptive immune recognition.


Subject(s)
Adaptive Immunity/immunology , Self Tolerance/immunology , T-Lymphocytes/immunology , Databases, Factual , Humans , Lymphocyte Activation/immunology , Peptides/immunology , Peptides/metabolism , Receptors, Antigen, T-Cell/immunology , T-Lymphocytes/metabolism
2.
Int J Mol Sci ; 23(5)2022 Feb 28.
Article in English | MEDLINE | ID: covidwho-1715409

ABSTRACT

We propose a new hypothesis that explains the maintenance and evolution of MHC polymorphism. It is based on two phenomena: the constitution of the repertoire of naive T lymphocytes and the evolution of the pathogen and its impact on the immune memory of T lymphocytes. Concerning the latter, pathogen evolution will have a different impact on reinfection depending on the MHC allomorph. If a mutation occurs in a given region, in the case of MHC allotypes, which do not recognize the peptide in this region, the mutation will have no impact on the memory repertoire. In the case where the MHC allomorph binds to the ancestral peptides and not to the mutated peptide, that individual will have a higher chance of being reinfected. This difference in fitness will lead to a variation of the allele frequency in the next generation. Data from the SARS-CoV-2 pandemic already support a significant part of this hypothesis and following up on these data may enable it to be confirmed. This hypothesis could explain why some individuals after vaccination respond less well than others to variants and leads to predict the probability of reinfection after a first infection depending upon the variant and the HLA allomorph.


Subject(s)
COVID-19/immunology , HLA Antigens/immunology , Polymorphism, Genetic/immunology , SARS-CoV-2/immunology , T-Lymphocytes/immunology , COVID-19/epidemiology , COVID-19/virology , Evolution, Molecular , Gene Frequency , HLA Antigens/genetics , HLA Antigens/metabolism , Humans , Immunity, Innate/genetics , Immunity, Innate/immunology , Mutation/genetics , Mutation/immunology , Pandemics , Peptides/immunology , Peptides/metabolism , Polymorphism, Genetic/genetics , SARS-CoV-2/physiology , T-Lymphocytes/cytology , T-Lymphocytes/metabolism
3.
ACS Appl Bio Mater ; 5(3): 905-944, 2022 03 21.
Article in English | MEDLINE | ID: covidwho-1705996

ABSTRACT

This review discusses peptide epitopes used as antigens in the development of vaccines in clinical trials as well as future vaccine candidates. It covers peptides used in potential immunotherapies for infectious diseases including SARS-CoV-2, influenza, hepatitis B and C, HIV, malaria, and others. In addition, peptides for cancer vaccines that target examples of overexpressed proteins are summarized, including human epidermal growth factor receptor 2 (HER-2), mucin 1 (MUC1), folate receptor, and others. The uses of peptides to target cancers caused by infective agents, for example, cervical cancer caused by human papilloma virus (HPV), are also discussed. This review also provides an overview of model peptide epitopes used to stimulate non-specific immune responses, and of self-adjuvanting peptides, as well as the influence of other adjuvants on peptide formulations. As highlighted in this review, several peptide immunotherapies are in advanced clinical trials as vaccines, and there is great potential for future therapies due the specificity of the response that can be achieved using peptide epitopes.


Subject(s)
Vaccines, Subunit/administration & dosage , Adjuvants, Immunologic/administration & dosage , Animals , Antigens/immunology , Cancer Vaccines/administration & dosage , Communicable Disease Control , Humans , Neoplasms/therapy , Peptides/immunology
4.
Viruses ; 14(2)2022 01 27.
Article in English | MEDLINE | ID: covidwho-1667340

ABSTRACT

Coronavirus disease 2019 (COVID-19), the pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is characterized by symptoms such as fever, fatigue, a sore throat, diarrhea, and coughing. Although various new vaccines against COVID-19 have been developed, early diagnostics, isolation, and prevention remain important due to virus mutations resulting in rapid and high disease transmission. Amino acid substitutions in the major diagnostic target antigens of SARS-CoV-2 may lower the sensitivity for the detection of SARS-CoV-2. For this reason, we developed specific monoclonal antibodies that bind to epitope peptides as antigens for the rapid detection of SARS-CoV-2 NP. The binding affinity between antigenic peptides and monoclonal antibodies was investigated, and a sandwich pair for capture and detection was employed to develop a rapid biosensor for SARS-CoV-2 NP. The rapid biosensor, based on a monoclonal antibody pair binding to conserved epitopes of SARS-CoV-2 NP, detected cultured virus samples of SARS-CoV-2 (1.4 × 103 TCID50/reaction) and recombinant NP (1 ng/mL). Laboratory confirmation of the rapid biosensor was performed with clinical specimens (n = 16) from COVID-19 patients and other pathogens. The rapid biosensor consisting of a monoclonal antibody pair (75E12 for capture and the 54G6/54G10 combination for detection) binding to conserved epitopes of SARS-CoV-2 NP could assist in the detection of SARS-CoV-2 NP under the circumstance of spreading SARS-CoV-2 variants.


Subject(s)
Antibodies, Monoclonal/metabolism , Antibodies, Viral/metabolism , Biosensing Techniques/methods , Epitopes/metabolism , Nucleocapsid Proteins/metabolism , SARS-CoV-2/immunology , Viral Proteins/metabolism , Animals , Antibodies, Monoclonal/immunology , Antibodies, Viral/immunology , Epitopes/genetics , Epitopes/immunology , Humans , Immunoassay , Mice , Mice, Inbred BALB C , Nucleocapsid Proteins/genetics , Nucleocapsid Proteins/immunology , Peptides/immunology , Peptides/metabolism , Protein Binding , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , Viral Proteins/immunology
5.
Int J Mol Sci ; 23(3)2022 Feb 01.
Article in English | MEDLINE | ID: covidwho-1667197

ABSTRACT

An Escherichia coli (E. coli) production of the receptor-binding domain (RBD) of the SARS-CoV-2 (isolate Wuhan-Hu-1) spike protein would significantly accelerate the search for anti-COVID-19 therapeutics because of its versatility and low cost. However, RBD contains four disulfide bonds and its expression in E. coli is limited by the formation of aberrant disulfide bonds resulting in inclusion bodies. Here, we show that a solubility-enhancing peptide (SEP) tag containing nine arginine residues (RBD-C9R) attached at the C-terminus can overcome this problem. The SEP-tag increased the expression in the soluble fraction and the final yield by five times (2 mg/L). The folding properties of the E. coli expressed RBD-C9R were demonstrated with biophysical characterization using RP-HPLC, circular dichroism, thermal denaturation, fluorescence, and light scattering. A quartz crystal microbalance (QCM) analysis confirmed the binding activity of RBD-C9R with ACE2, the host cell's receptor. In addition, RBD-C9R elicited a Th-2 immune response with a high IgG titer in Jcl: ICR mice. The RBD-C9R antisera interacted with both itself and the mammalian-cell expressed spike protein (S1), as demonstrated by ELISA, indicating that the E. coli expressed RBD-C9R harbors native-like epitopes. Overall, these results emphasize the potential of our SEP-tag for the E. coli production of active multi-disulfide-bonded RBD.


Subject(s)
Antibodies, Viral/blood , Escherichia coli/growth & development , Peptides/administration & dosage , Spike Glycoprotein, Coronavirus/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Animals , Cloning, Molecular , Disulfides/metabolism , Escherichia coli/genetics , Female , Humans , Immune Sera/metabolism , Immunization , Mice , Mice, Inbred ICR , Peptides/genetics , Peptides/immunology , Protein Domains , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology , Th2 Cells/metabolism
6.
Front Immunol ; 12: 812176, 2021.
Article in English | MEDLINE | ID: covidwho-1662586

ABSTRACT

Although not being the first viral pandemic to affect humankind, we are now for the first time faced with a pandemic caused by a coronavirus. The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has been responsible for the COVID-19 pandemic, which caused more than 4.5 million deaths worldwide. Despite unprecedented efforts, with vaccines being developed in a record time, SARS-CoV-2 continues to spread worldwide with new variants arising in different countries. Such persistent spread is in part enabled by public resistance to vaccination in some countries, and limited access to vaccines in other countries. The limited vaccination coverage, the continued risk for resistant variants, and the existence of natural reservoirs for coronaviruses, highlight the importance of developing additional therapeutic strategies against SARS-CoV-2 and other coronaviruses. At the beginning of the pandemic it was suggested that countries with Bacillus Calmette-Guérin (BCG) vaccination programs could be associated with a reduced number and/or severity of COVID-19 cases. Preliminary studies have provided evidence for this relationship and further investigation is being conducted in ongoing clinical trials. The protection against SARS-CoV-2 induced by BCG vaccination may be mediated by cross-reactive T cell lymphocytes, which recognize peptides displayed by class I Human Leukocyte Antigens (HLA-I) on the surface of infected cells. In order to identify potential targets of T cell cross-reactivity, we implemented an in silico strategy combining sequence-based and structure-based methods to screen over 13,5 million possible cross-reactive peptide pairs from BCG and SARS-CoV-2. Our study produced (i) a list of immunogenic BCG-derived peptides that may prime T cell cross-reactivity against SARS-CoV-2, (ii) a large dataset of modeled peptide-HLA structures for the screened targets, and (iii) new computational methods for structure-based screenings that can be used by others in future studies. Our study expands the list of BCG peptides potentially involved in T cell cross-reactivity with SARS-CoV-2-derived peptides, and identifies multiple high-density "neighborhoods" of cross-reactive peptides which could be driving heterologous immunity induced by BCG vaccination, therefore providing insights for future vaccine development efforts.


Subject(s)
BCG Vaccine/immunology , COVID-19/immunology , Cross Reactions/immunology , Peptides/immunology , SARS-CoV-2/immunology , T-Lymphocytes/immunology , Viral Vaccines/immunology , Humans , Pandemics/prevention & control , Vaccination/methods
7.
J Clin Invest ; 132(5)2022 03 01.
Article in English | MEDLINE | ID: covidwho-1642991

ABSTRACT

Recent studies have shown that vaccinated individuals harbor T cells that can cross-recognize SARS-CoV-2 and endemic human common cold coronaviruses. However, it is still unknown whether CD4+ T cells from vaccinated individuals recognize peptides from bat coronaviruses that may have the potential of causing future pandemics. In this study, we identified a SARS-CoV-2 spike protein epitope (S815-827) that is conserved in coronaviruses from different genera and subgenera, including SARS-CoV, MERS-CoV, multiple bat coronaviruses, and a feline coronavirus. Our results showed that S815-827 was recognized by 42% of vaccinated participants in our study who received the Pfizer-BioNTech (BNT162b2) or Moderna (mRNA-1273) COVID-19 vaccines. Using T cell expansion and T cell receptor sequencing assays, we demonstrated that S815-827-reactive CD4+ T cells from the majority of responders cross-recognized homologous peptides from at least 6 other diverse coronaviruses. Our results support the hypothesis that the current mRNA vaccines elicit T cell responses that can cross-recognize bat coronaviruses and thus might induce some protection against potential zoonotic outbreaks. Furthermore, our data provide important insights that inform the development of T cell-based pan-coronavirus vaccine strategies.


Subject(s)
/immunology , CD4-Positive T-Lymphocytes/immunology , COVID-19/immunology , Epitopes, T-Lymphocyte/immunology , Receptors, Antigen, T-Cell/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , /administration & dosage , COVID-19/prevention & control , Female , Humans , Male , Peptides/immunology
8.
ACS Appl Mater Interfaces ; 14(4): 4811-4822, 2022 Feb 02.
Article in English | MEDLINE | ID: covidwho-1641826

ABSTRACT

Canonical immunoassays rely on highly sensitive and specific capturing of circulating biomarkers by interacting biomolecular baits. In this frame, bioprobe immobilization in spatially discrete three-dimensional (3D) spots onto analytical surfaces by hydrogel encapsulation was shown to provide relevant advantages over conventional two-dimensional (2D) platforms. Yet, the broad application of 3D systems is still hampered by hurdles in matching their straightforward fabrication with optimal functional properties. Herein, we report on a composite hydrogel obtained by combining a self-assembling peptide (namely, Q3 peptide) with low-temperature gelling agarose that is proved to have simple and robust application in the fabrication of microdroplet arrays, overcoming hurdles and limitations commonly associated with 3D hydrogel assays. We demonstrate the real-case scenario feasibility of our 3D system in the profiling of Covid-19 patients' serum IgG immunoreactivity, which showed remarkably improved signal-to-noise ratio over canonical assays in the 2D format and exquisite specificity. Overall, the new two-component hydrogel widens the perspectives of hydrogel-based arrays and represents a step forward towards their routine use in analytical practices.


Subject(s)
COVID-19/diagnosis , Immunoassay/methods , Immunoglobulin G/blood , SARS-CoV-2/isolation & purification , Biomarkers/blood , COVID-19/blood , COVID-19/immunology , COVID-19/virology , Humans , Hydrogels/chemistry , Immunoglobulin G/immunology , Peptides/chemistry , Peptides/immunology , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Sepharose
9.
J Immunol ; 208(4): 979-990, 2022 02 15.
Article in English | MEDLINE | ID: covidwho-1631932

ABSTRACT

Calprotectin is released by activated neutrophils along with myeloperoxidase (MPO) and proteases. It plays numerous roles in inflammation and infection, and is used as an inflammatory biomarker. However, calprotectin is readily oxidized by MPO-derived hypohalous acids to form covalent dimers of its S100A8 and S100A9 subunits. The dimers are susceptible to degradation by proteases. We show that detection of human calprotectin by ELISA declines markedly because of its oxidation by hypochlorous acid and subsequent degradation. Also, proteolysis liberates specific peptides from oxidized calprotectin that is present at inflammatory sites. We identified six calprotectin-derived peptides by mass spectrometry and detected them in the bronchoalveolar lavage fluid of children with cystic fibrosis (CF). We assessed the peptides as biomarkers of neutrophilic inflammation and infection. The content of the calprotectin peptide ILVI was related to calprotectin (r = 0.72, p = 0.01, n = 10). Four of the peptides were correlated with the concentration of MPO (r > 0.7, p ≤ 0.01, n = 21), while three were higher (p < 0.05) in neutrophil elastase-positive (n = 14) than -negative samples (n = 7). Also, five of the peptides were higher (p < 0.05) in the bronchoalveolar lavage fluid from children with CF with infections (n = 21) than from non-CF children without infections (n = 6). The specific peptides liberated from calprotectin will signal uncontrolled activity of proteases and MPO during inflammation. They may prove useful in tracking inflammation in respiratory diseases dominated by neutrophils, including coronavirus disease 2019.


Subject(s)
Bronchoalveolar Lavage Fluid/immunology , Cystic Fibrosis/immunology , Inflammation/immunology , Leukocyte L1 Antigen Complex/metabolism , Neutrophils/immunology , Peptides/metabolism , Respiratory System/metabolism , Child , Child, Preschool , Cystic Fibrosis/diagnosis , Female , Humans , Inflammation/diagnosis , Leukocyte L1 Antigen Complex/genetics , Leukocyte L1 Antigen Complex/immunology , Male , Neutrophil Activation , Oxidation-Reduction , Peptides/genetics , Peptides/immunology , Proteolysis
10.
Nat Commun ; 13(1): 19, 2022 01 10.
Article in English | MEDLINE | ID: covidwho-1616981

ABSTRACT

T cells play a vital role in combatting SARS-CoV-2 and forming long-term memory responses. Whereas extensive structural information is available on neutralizing antibodies against SARS-CoV-2, such information on SARS-CoV-2-specific T-cell receptors (TCRs) bound to their peptide-MHC targets is lacking. Here we determine the structures of a public and a private TCR from COVID-19 convalescent patients in complex with HLA-A2 and two SARS-CoV-2 spike protein epitopes (YLQ and RLQ). The structures reveal the basis for selection of particular TRAV and TRBV germline genes by the public but not the private TCR, and for the ability of the TCRs to recognize natural variants of RLQ but not YLQ. Neither TCR recognizes homologous epitopes from human seasonal coronaviruses. By elucidating the mechanism for TCR recognition of an immunodominant yet variable epitope (YLQ) and a conserved but less commonly targeted epitope (RLQ), this study can inform prospective efforts to design vaccines to elicit pan-coronavirus immunity.


Subject(s)
COVID-19/immunology , Epitopes, T-Lymphocyte/immunology , HLA-A2 Antigen/immunology , Receptors, Antigen, T-Cell/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD4-Positive T-Lymphocytes/virology , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/virology , COVID-19/virology , Epitopes, T-Lymphocyte/metabolism , HLA-A2 Antigen/chemistry , HLA-A2 Antigen/metabolism , Humans , Immunodominant Epitopes/immunology , Immunodominant Epitopes/metabolism , Jurkat Cells , K562 Cells , Peptides/chemistry , Peptides/immunology , Peptides/metabolism , Protein Binding , Protein Conformation , Receptors, Antigen, T-Cell/chemistry , Receptors, Antigen, T-Cell/metabolism , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Spike Glycoprotein, Coronavirus/metabolism , Surface Plasmon Resonance/methods
11.
Pathog Dis ; 80(1)2022 02 09.
Article in English | MEDLINE | ID: covidwho-1612517

ABSTRACT

Given the emergence of SARS-CoV-2 virus as a life-threatening pandemic, identification of immunodominant epitopes of the viral structural proteins, particularly the nucleocapsid (NP) protein and receptor-binding domain (RBD) of spike protein, is important to determine targets for immunotherapy and diagnosis. In this study, epitope screening was performed using a panel of overlapping peptides spanning the entire sequences of the RBD and NP proteins of SARS-CoV-2 in the sera from 66 COVID-19 patients and 23 healthy subjects by enzyme-linked immunosorbent assay (ELISA). Our results showed that while reactivity of patients' sera with reduced recombinant RBD protein was significantly lower than the native form of RBD (P < 0.001), no significant differences were observed for reactivity of patients' sera with reduced and non-reduced NP protein. Pepscan analysis revealed weak to moderate reactivity towards different RBD peptide pools, which was more focused on peptides encompassing amino acids (aa) 181-223 of RBD. NP peptides, however, displayed strong reactivity with a single peptide covering aa 151-170. These findings were confirmed by peptide depletion experiments using both ELISA and western blotting. Altogether, our data suggest involvement of mostly conformational disulfide bond-dependent immunodominant epitopes in RBD-specific antibody response, while the IgG response to NP is dominated by linear epitopes. Identification of dominant immunogenic epitopes in NP and RBD of SARS-CoV-2 could provide important information for the development of passive and active immunotherapy as well as diagnostic tools for the control of COVID-19 infection.


Subject(s)
Antibodies, Viral/immunology , COVID-19/immunology , Immunodominant Epitopes/immunology , Nucleocapsid/immunology , Receptors, Virus/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Aged , Amino Acid Motifs , Antibodies, Viral/blood , COVID-19/virology , Enzyme-Linked Immunosorbent Assay/methods , Female , Humans , Immunodominant Epitopes/chemistry , Iran , Male , Middle Aged , Pandemics , Peptides/immunology , Protein Binding , SARS-CoV-2/chemistry , Spike Glycoprotein, Coronavirus/chemistry , Viral Proteins/immunology
12.
Anal Bioanal Chem ; 414(5): 1949-1962, 2022 Feb.
Article in English | MEDLINE | ID: covidwho-1607761

ABSTRACT

Recently, numerous diagnostic approaches from different disciplines have been developed for SARS-CoV-2 diagnosis to monitor and control the COVID-19 pandemic. These include MS-based assays, which provide analytical information on viral proteins. However, their sensitivity is limited, estimated to be 5 × 104 PFU/ml in clinical samples. Here, we present a reliable, specific, and rapid method for the identification of SARS-CoV-2 from nasopharyngeal (NP) specimens, which combines virus capture followed by LC-MS/MS(MRM) analysis of unique peptide markers. The capture of SARS-CoV-2 from the challenging matrix, prior to its tryptic digestion, was accomplished by magnetic beads coated with polyclonal IgG-α-SARS-CoV-2 antibodies, enabling sample concentration while significantly reducing background noise interrupting with LC-MS analysis. A sensitive and specific LC-MS/MS(MRM) analysis method was developed for the identification of selected tryptic peptide markers. The combined assay, which resulted in S/N ratio enhancement, achieved an improved sensitivity of more than 10-fold compared with previously described MS methods. The assay was validated in 29 naive NP specimens, 19 samples were spiked with SARS-CoV-2 and 10 were used as negative controls. Finally, the assay was successfully applied to clinical NP samples (n = 26) pre-determined as either positive or negative by RT-qPCR. This work describes for the first time a combined approach for immuno-magnetic viral isolation coupled with MS analysis. This method is highly reliable, specific, and sensitive; thus, it may potentially serve as a complementary assay to RT-qPCR, the gold standard test. This methodology can be applied to other viruses as well.


Subject(s)
COVID-19 Testing/methods , COVID-19/diagnosis , Chromatography, Liquid/methods , Immunomagnetic Separation/methods , SARS-CoV-2/genetics , Tandem Mass Spectrometry/methods , Amino Acid Sequence , Antibodies, Viral/chemistry , Biomarkers/chemistry , COVID-19/immunology , COVID-19/virology , COVID-19 Testing/instrumentation , COVID-19 Testing/standards , Chromatography, Liquid/instrumentation , Chromatography, Liquid/standards , Humans , Immunomagnetic Separation/instrumentation , Immunomagnetic Separation/standards , Nasopharynx/virology , Peptides/chemistry , Peptides/immunology , SARS-CoV-2/immunology , Sensitivity and Specificity , Tandem Mass Spectrometry/instrumentation , Tandem Mass Spectrometry/standards
13.
Cell Immunol ; 371: 104454, 2022 01.
Article in English | MEDLINE | ID: covidwho-1509640

ABSTRACT

Immune dysregulation is commonly observed in patients with coronavirus disease 2019 (COVID-19). Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces severe lung inflammation and innate immune cell dysregulation. However, the precise interaction between SARS-CoV-2 and the innate immune system is currently unknown. To understand the interaction between SARS-CoV-2 and natural killer (NK) cells, several SARS-CoV-2 S protein peptides capable of binding to the NKG2D receptor were screened by in silico analysis. Among them, two peptides, cov1 and cov2, bound to NK cells and NKG2D receptors. These cov peptides increased NK cytotoxicity toward lung cancer cells, stimulated interferon gamma (IFN-γ) production by NK cells, and likely mediated these responses through the phosphorylation of Vav1, a key downstream-signaling molecule of NKG2D and NK activation genes. The direct interaction between SARS-CoV-2 and NK cells is a novel finding, and modulation of this interaction has potential clinical application as a therapeutic target for COVID-19.


Subject(s)
COVID-19/immunology , Killer Cells, Natural/immunology , NK Cell Lectin-Like Receptor Subfamily K/immunology , Peptides/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/immunology , Amino Acid Sequence , COVID-19/metabolism , COVID-19/virology , Cell Line, Tumor , Cytotoxicity, Immunologic/immunology , Humans , Interferon-gamma/immunology , Interferon-gamma/metabolism , Killer Cells, Natural/metabolism , Lung/immunology , Lung/pathology , Lung/virology , Lymphocyte Activation/immunology , NK Cell Lectin-Like Receptor Subfamily K/metabolism , Peptides/metabolism , Protein Binding , SARS-CoV-2/metabolism , SARS-CoV-2/physiology , Signal Transduction/immunology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
14.
J Biol Chem ; 297(5): 101329, 2021 11.
Article in English | MEDLINE | ID: covidwho-1474696

ABSTRACT

Population genetic variability in immune system genes can often underlie variability in immune responses to pathogens. Cytotoxic T-lymphocytes are emerging as critical determinants of both severe acute respiratory syndrome coronavirus 2 infection severity and long-term immunity, after either recovery or vaccination. A hallmark of coronavirus disease 2019 is its highly variable severity and breadth of immune responses between individuals. To address the underlying mechanisms behind this phenomenon, we analyzed the proteolytic processing of S1 spike glycoprotein precursor antigenic peptides across ten common allotypes of endoplasmic reticulum aminopeptidase 1 (ERAP1), a polymorphic intracellular enzyme that can regulate cytotoxic T-lymphocyte responses by generating or destroying antigenic peptides. We utilized a systematic proteomic approach that allows the concurrent analysis of hundreds of trimming reactions in parallel, thus better emulating antigen processing in the cell. While all ERAP1 allotypes were capable of producing optimal ligands for major histocompatibility complex class I molecules, including known severe acute respiratory syndrome coronavirus 2 epitopes, they presented significant differences in peptide sequences produced, suggesting allotype-dependent sequence biases. Allotype 10, previously suggested to be enzymatically deficient, was rather found to be functionally distinct from other allotypes. Our findings suggest that common ERAP1 allotypes can be a major source of heterogeneity in antigen processing and through this mechanism contribute to variable immune responses in coronavirus disease 2019.


Subject(s)
Aminopeptidases/immunology , Antigens, Viral/immunology , Immunoglobulin Allotypes/immunology , Minor Histocompatibility Antigens/immunology , Peptides/immunology , SARS-CoV-2/chemistry , Spike Glycoprotein, Coronavirus/immunology , Aminopeptidases/chemistry , Antigen Presentation/immunology , Humans , Minor Histocompatibility Antigens/chemistry , Peptides/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/immunology , SARS-CoV-2/immunology , Spike Glycoprotein, Coronavirus/chemistry
15.
Molecules ; 26(20)2021 Oct 13.
Article in English | MEDLINE | ID: covidwho-1470934

ABSTRACT

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2, the causative agent of coronavirus disease (COVID-19)) has caused relatively high mortality rates in humans throughout the world since its first detection in late December 2019, leading to the most devastating pandemic of the current century. Consequently, SARS-CoV-2 therapeutic interventions have received high priority from public health authorities. Despite increased COVID-19 infections, a vaccine or therapy to cover all the population is not yet available. Herein, immunoinformatics and custommune tools were used to identify B and T-cells epitopes from the available SARS-CoV-2 sequences spike (S) protein. In the in silico predictions, six B cell epitopes QTGKIADYNYK, TEIYQASTPCNGVEG, LQSYGFQPT, IRGDEVRQIAPGQTGKIADYNYKLPD, FSQILPDPSKPSKRS and PFAMQMAYRFNG were cross-reacted with MHC-I and MHC-II T-cells binding epitopes and selected for vaccination in experimental animals for evaluation as candidate vaccine(s) due to their high antigenic matching and conserved score. The selected six peptides were used individually or in combinations to immunize female Balb/c mice. The immunized mice raised reactive antibodies against SARS-CoV-2 in two different short peptides located in receptor binding domain and S2 region. In combination groups, an additive effect was demonstrated in-comparison with single peptide immunized mice. This study provides novel epitope-based peptide vaccine candidates against SARS-CoV-2.


Subject(s)
COVID-19 Vaccines/chemistry , COVID-19/prevention & control , Epitopes, B-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/chemistry , SARS-CoV-2/metabolism , Amino Acid Sequence , Animals , Antibodies, Viral/blood , Antibodies, Viral/immunology , COVID-19/virology , COVID-19 Vaccines/administration & dosage , Epitopes, B-Lymphocyte/immunology , Epitopes, B-Lymphocyte/metabolism , Epitopes, T-Lymphocyte/immunology , Epitopes, T-Lymphocyte/metabolism , Female , Humans , Immunization , Mice , Mice, Inbred BALB C , Peptides/chemistry , Peptides/immunology , Peptides/metabolism , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
16.
Int J Mol Sci ; 22(20)2021 Oct 18.
Article in English | MEDLINE | ID: covidwho-1470893

ABSTRACT

SputnikV is a vaccine against SARS-CoV-2 developed by the Gamaleya National Research Centre for Epidemiology and Microbiology. The vaccine has been shown to induce both humoral and cellular immune responses, yet the mechanisms remain largely unknown. Forty SputnikV vaccinated individuals were included in this study which aimed to demonstrate the location of immunogenic domains of the SARS-CoV-2 S protein using an overlapping peptide library. Additionally, cytokines in the serum of vaccinated and convalescent COVID-19 patients were analyzed. We have found antibodies from both vaccinated and convalescent sera bind to immunogenic regions located in multiple domains of SARS-CoV-2 S protein, including Receptor Binding Domain (RBD), N-terminal Domain (NTD), Fusion Protein (FP) and Heptad Repeats (HRs). Interestingly, many peptides were recognized by immunized and convalescent serum antibodies and correspond to conserved regions in circulating variants of SARS-CoV-2. This breadth of reactivity was still evident 90 days after the first dose of the vaccine, showing that the vaccine has induced a prolonged response. As evidenced by the activation of T cells, cellular immunity strongly suggests the high potency of the SputnikV vaccine against SARS-CoV-2 infection.


Subject(s)
COVID-19 Vaccines/administration & dosage , COVID-19/prevention & control , Immunity, Cellular , Immunity, Humoral , Adult , Amino Acid Sequence , COVID-19/immunology , COVID-19/virology , COVID-19 Vaccines/immunology , Cytokines/metabolism , Female , Humans , Male , Peptides/chemistry , Peptides/immunology , Principal Component Analysis , Recombinant Fusion Proteins/biosynthesis , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/immunology , SARS-CoV-2/isolation & purification , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/immunology , T-Lymphocytes/cytology , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Vaccination
17.
Clin Transl Gastroenterol ; 12(10): e00411, 2021 10 06.
Article in English | MEDLINE | ID: covidwho-1456553

ABSTRACT

INTRODUCTION: The adherence to a gluten-free diet (GFD) is a trending topic in the management of celiac disease. The aim of our study was to evaluate the diagnostic performance of urinary gluten immunogenic peptides (GIP) determination to detect gluten contamination of the GFD. METHODS: In study A, 25 healthy adults on a standard GFD performed 6 gluten challenges (0, 10, 50, 100, 500, and 1,000 mg) with quantification of urinary GIP before (T0) and during the following 24 hours. In study B, 12 participants on a gluten contamination elimination diet underwent urinary GIP determination at T0 and after challenge with 5 or 10 mg gluten. Urine GIP concentration was determined by an immunochromatographic assay. RESULTS: In study A, 51 of 150 baseline urine samples were GIP+ on GFD and 7 of 17 were GIP+ after the zero-gluten challenge, whereas only 55 of 81 were GIP+ after the 10-1,000 mg gluten challenges. There was no significant change in the 24-hour urinary GIP when increasing gluten from 10 to 1,000 mg. In study B, 24 of 24 baseline urine samples were GIP-, whereas 8 of 24 were GIP+ after 5 or 10 mg of gluten. DISCUSSION: Traces of gluten in the standard GFD may cause positivity of urinary GIP determination, whereas a false negativity is common after a gluten intake of 10-1,000 mg. Owing to the impossibility of standardizing the test in normal conditions, it seems unlikely that urinary GIP determination may represent a reliable tool to assess the compliance to the GFD of patients with celiac disease or other gluten-related disorders.


Subject(s)
Celiac Disease/diet therapy , Celiac Disease/urine , Diet, Gluten-Free , Glutens/urine , Patient Compliance , Peptides/urine , Adult , Celiac Disease/immunology , Double-Blind Method , Female , Glutens/immunology , Humans , Immunoglobulin A/blood , Male , Peptides/immunology , Transglutaminases/immunology
18.
Immunity ; 54(9): 2133-2142.e3, 2021 09 14.
Article in English | MEDLINE | ID: covidwho-1433401

ABSTRACT

SARS-CoV-2 mRNA vaccines have shown remarkable clinical efficacy, but questions remain about the nature and kinetics of T cell priming. We performed longitudinal antigen-specific T cell analyses on healthy SARS-CoV-2-naive and recovered individuals prior to and following mRNA prime and boost vaccination. Vaccination induced rapid antigen-specific CD4+ T cell responses in naive subjects after the first dose, whereas CD8+ T cell responses developed gradually and were variable in magnitude. Vaccine-induced Th1 and Tfh cell responses following the first dose correlated with post-boost CD8+ T cells and neutralizing antibodies, respectively. Integrated analysis revealed coordinated immune responses with distinct trajectories in SARS-CoV-2-naive and recovered individuals. Last, whereas booster vaccination improved T cell responses in SARS-CoV-2-naive subjects, the second dose had little effect in SARS-CoV-2-recovered individuals. These findings highlight the role of rapidly primed CD4+ T cells in coordinating responses to the second vaccine dose in SARS-CoV-2-naive individuals.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , COVID-19 Vaccines/immunology , COVID-19/immunology , SARS-CoV-2/physiology , Th1 Cells/immunology , Adult , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Antigens, CD/metabolism , Antigens, Differentiation, T-Lymphocyte/metabolism , Female , Humans , Immunity, Cellular , Immunity, Humoral , Immunization, Secondary , Immunologic Memory , Lectins, C-Type/metabolism , Lymphocyte Activation , Male , Middle Aged , Peptides/immunology , Spike Glycoprotein, Coronavirus/immunology , Vaccination , Young Adult
19.
Front Immunol ; 12: 724060, 2021.
Article in English | MEDLINE | ID: covidwho-1430701

ABSTRACT

Thirty-five peptides selected from functionally-relevant SARS-CoV-2 spike (S), membrane (M), and envelope (E) proteins were suitably modified for immunising MHC class II (MHCII) DNA-genotyped Aotus monkeys and matched with HLA-DRß1* molecules for use in humans. This was aimed at producing the first minimal subunit-based, chemically-synthesised, immunogenic molecules (COLSARSPROT) covering several HLA alleles. They were predicted to cover 48.25% of the world's population for 6 weeks (short-term) and 33.65% for 15 weeks (long-lasting) as they induced very high immunofluorescent antibody (IFA) and ELISA titres against S, M and E parental native peptides, SARS-CoV-2 neutralising antibodies and host cell infection. The same immunological methods that led to identifying new peptides for inclusion in the COLSARSPROT mixture were used for antigenicity studies. Peptides were analysed with serum samples from patients suffering mild or severe SARS-CoV-2 infection, thereby increasing chemically-synthesised peptides' potential coverage for the world populations up to 62.9%. These peptides' 3D structural analysis (by 1H-NMR acquired at 600 to 900 MHz) suggested structural-functional immunological association. This first multi-protein, multi-epitope, minimal subunit-based, chemically-synthesised, highly immunogenic peptide mixture highlights such chemical synthesis methodology's potential for rapidly obtaining very pure, highly reproducible, stable, cheap, easily-modifiable peptides for inducing immune protection against COVID-19, covering a substantial percentage of the human population.


Subject(s)
COVID-19 Vaccines/immunology , COVID-19/immunology , Coronavirus Envelope Proteins/immunology , Coronavirus M Proteins/immunology , Spike Glycoprotein, Coronavirus/immunology , Vaccines, Subunit/immunology , Animals , Antibodies, Neutralizing/blood , Antibodies, Neutralizing/immunology , Antibodies, Viral/blood , Antibodies, Viral/immunology , Aotidae , COVID-19/prevention & control , HLA-DRB1 Chains/genetics , Humans , Peptides/immunology , SARS-CoV-2/immunology
20.
Front Immunol ; 12: 728936, 2021.
Article in English | MEDLINE | ID: covidwho-1413272

ABSTRACT

The use of minimal peptide sets offers an appealing alternative for design of vaccines and T cell diagnostics compared to conventional whole protein approaches. T cell immunogenicity towards peptides is contingent on binding to human leukocyte antigen (HLA) molecules of the given individual. HLA is highly polymorphic, and each variant typically presents a different repertoire of peptides. This polymorphism combined with pathogen diversity challenges the rational selection of peptide sets with broad immunogenic potential and population coverage. Here we propose PopCover-2.0, a simple yet highly effective method, for resolving this challenge. The method takes as input a set of (predicted) CD8 and/or CD4 T cell epitopes with associated HLA restriction and pathogen strain annotation together with information on HLA allele frequencies, and identifies peptide sets with optimal pathogen and HLA (class I and II) coverage. PopCover-2.0 was benchmarked on historic data in the context of HIV and SARS-CoV-2. Further, the immunogenicity of the selected SARS-CoV-2 peptides was confirmed by experimentally validating the peptide pools for T cell responses in a panel of SARS-CoV-2 infected individuals. In summary, PopCover-2.0 is an effective method for rational selection of peptide subsets with broad HLA and pathogen coverage. The tool is available at https://services.healthtech.dtu.dk/service.php?PopCover-2.0.


Subject(s)
Epitopes, T-Lymphocyte/immunology , HLA Antigens/genetics , HLA Antigens/immunology , Peptides/immunology , Alleles , Allergy and Immunology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , COVID-19/immunology , COVID-19/prevention & control , Genotype , HLA Antigens/classification , Humans , Immunogenicity, Vaccine , Immunologic Techniques , Peptides/classification , SARS-CoV-2/immunology
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