Immunodominant Epitope Peptides Research Articles

Mapping of conserved immunodominant epitope peptides in the outer membrane porin (Omp) L of prominent Enterobacteriaceae pathogens associated with gastrointestinal infections

BackgroundMembers of Enterobacteriaceae such as Escherichia coli O 157:H7, Salmonella sp., Shigella sp., Klebsiella sp., and Citrobacter freundii are responsible for the outbreak of serious foodborne illness and other mucosal infections across the globe. The outer membrane proteins (OMPs) of Enterobacteriaceae are highly immunogenic in eliciting immune responses against pathogens. Moreover, the OMPs are highly conserved in the Enterobacteriaceae family. Sequence homology in the OMPs will ensure the presence of conserved immunodominant regions with predominant epitopes. The OmpL is such an immunogen that is highly conserved among the Enterobacteriaceae pathogens. In this study, we performed computational analysis on the outer membrane porin (Omp) L of prominent Enterobacteriaceae pathogens. ResultsMultiple sequence and structural alignment analysis have revealed that the OmpL protein is highly conserved among the selected Enterobacteriaceae pathogens. This amount of sequence and structural homology uncovered the conserved antibody binding B-cell epitopes in the OmpL protein. The B-cell epitopes predicted in the OmpL of Salmonella typhimurium are highly conserved among the other Enterobacteriaceae pathogens. ConclusionIn conclusion, these conserved B-cell epitopes will vouch for the generation of heterologous humoral immune response in conferring cross protection against the Enterobacteriaceae pathogens and control their outbreaks across the globe.

Identification of a Novel Linear B Cell Epitope on the Sao Protein of Streptococcus suis Serotype 2

Surface antigen one (Sao) protein is a bacterial surface protein identified in the important zoonotic pathogen Streptococcus suis serotype 2 (S. suis 2) during an extensive search for functional proteins. The Sao protein is anchored to the bacterial cell wall by the LPVTG motif and is widely distributed in many S. suis serotypes. In this paper, we present the immunodominant epitope peptide of the Sao protein that is recognized by BALB/c antibodies against the Sao protein: 355SEKQMPSVVNENAVTPEKQMTNKENDNIET384 (location Sao355−384). To determine the core epitope recognized by antibodies, we prepared truncation peptide libraries. Analyses of the immunoreactivity of truncation peptides with anti-Sao355−384 serum revealed that the most immunoreactive sequence was 355SEKQMPSVVNENAVTPEK372 (location Sao355−372). Moreover, we observed that this core epitope also showed good specificity based on the ratio of reactivity with serum from S. suis–positive patients compared to serum from S. suis–negative patients. Our results point to the potential of using the Sao355−372 peptide in diagnostic assays to determine S. suis infection in humans.

Intranasal immunization with immunodominant epitope peptides derived from HpaA conjugated with CpG adjuvant protected mice against Helicobacter pylori infection

HpaA is considered to be an effective protective antigen for vaccination against Helicobacter pylori (H. pylori) infection. Oral immunization with HpaA significantly decreases bacterial colonization in H. pylori infected mice. In this study, we investigated whether subcutaneous or intranasal immunization with HpaA could protect against H. pylori infection. Mice immunized subcutaneously with HpaA in Complete Freund’s adjuvant, but not mice intranasally immunized with HpaA in CpG adjuvant acquired protection against H. pylori infection. However, intranasal immunization with immunodominant epitope peptides in CpG adjuvant protected mice against H. pylori infection, and immunodominant epitope peptides stimulated stronger Th1 responses and mediated more robust protection against H. pylori infection than subdominant ones. Our results suggest that the length of a candidate antigen is critical for particular vaccination routes, and that immunodominant epitope peptides are promising candidates for protection against H. pylori infection through nasal vaccination.

Development of a M cell-targeted microparticulate platform, BSK02™, for oral immunization against the ovarian cancer antigen, sperm protein 17.

Although it only accounts for approximately 5% of all female cancer cases, ovarian cancer (OC) ranks as the fifth leading cause of death due to cancer in women. We have evaluated the potential of an orally administered microparticulate vaccine incorporating an immunodominant epitope peptide derived from the cancer/testis antigen sperm protein 17 (SP17) aberrantly expressed in OC, to retard the progression of the disease. The peptide antigen and the immune-stimulatory toll-like receptor 9 ligand CpG oligonucleotide were incorporated into spray dried microparticles composed of enteric and sustained release polymers together with the Aleuria aurantia lectin targeting microfold cells present in the gut-associated lymphoid tissue. These particles were administered via oral route to mice challenged week prior with SP17-expressing ID8 OC cells. Analysis of splenocytes harvested from vaccinated mice revealed strong activation of IFN-γ+/CD8+ lymphocytes in response to re-stimulation with the SP17 antigen. Moreover, vaccinated animals showed significant retardation of ascites/tumor volume in comparison to placebo-treated animals four weeks after the tumor challenge (p = 0.005). Taken together, our results suggest that vaccination against SP17 using orally administered microparticles could potentially be used as an effective consolidation strategy for OC patients with residual tumor or high probability for relapse following first-line treatments. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 107B: 29-36, 2019.

Novel immunodominant epitopes derived from MAGE-A3 and its significance in serological diagnosis of gastric cancer

To evaluate the significance of MAGE-A3 novel immunodominant epitopes in serological diagnosis of gastric cancer. B cell, CTL, and Th epitopes of MAGE-A3 were analyzed using computer-assisted techniques. Three possible immunodominant epitope peptides located at 5aa-23aa (QRSQHCKPEEGLEARGEAL), 112aa-131aa (KVAELVHFLLLKYRAREPVT), and 232aa-246aa (EGREDSILGDPKKLL) with potential B cell-dominant epitope, high-score HLA-A2 and A24 restriction CTL epitope, and HLA-DRB restriction Th epitope were selected. After optimized by prokaryotic codon, these genes were expressed as Trx-His-tag recombinant proteins in Escherichia coli and purified by Ni-NTA agarose beads. Three recombinant proteins were identified by Western blotting using His-tag monoclonal antibody and the serum antibodies from the patient of gastric cancer. The level of specific antibodies in the sera from 210 patients with gastric cancer, 56 patients with chronic gastritis, and 116 healthy controls was further analyzed by indirect ELISA. Three MAGE-A3 epitope recombinant proteins about 20kDa molecular weight were specifically recognized by His-tag monoclonal antibody and the serum of gastric cancer patients. ELISA based on the epitope recombinant protein indicated that gastric cancer patients had significantly higher reactivity to these immunodominant epitope proteins compared with chronic gastritis and healthy individuals (P<0.05). Furthermore, the serum antibody positive rate in the gastric cancer group was also significantly higher than that in the chronic gastritis patients and healthy controls (P<0.05), while there was no significant difference in gastritis group and the healthy control group (P>0.05). These study results demonstrated that these three predictive epitopes may be potential targets for applications in the design of serological diagnosis tools for gastric cancer.

Stability of Interferon-Gamma and Interleukin-10 Responses to Plasmodium falciparum Liver Stage Antigen 1 and Thrombospondin-Related Adhesive Protein Immunodominant Epitopes in a Highland Population from Western Kenya

Long-term planning to prevent malaria epidemics requires in-depth understanding of immunity to Plasmodium falciparum in areas of unstable transmission. Cytokine responses to immunodominant epitope peptides from liver stage antigen 1 (LSA-1) and thrombospondin-related adhesive protein (TRAP) were evaluated over a nine-month interval in adults and children in Kenya from a malaria epidemic-prone highland area after several years of low transmission. The proportion and magnitude of interferon-gamma ELISPOT responses and the proportion of interleukin-10 responders to LSA-1 and TRAP peptides tended to be higher in adults than children. Frequencies of interferon-gamma responders to these peptides were similar at the two time points, but responses were not consistently generated by the same persons. These results suggest that T cell memory to pre-erythrocytic stage malaria antigens is maintained but may be unavailable for consistent detection in peripheral blood, and that these antigens induce both pro-inflammatory and anti-inflammatory cytokine responses in this population.

Characterization of T Cell Responses to the RgpA-Kgp Proteinase-Adhesin Complexes of Porphyromonas gingivalis in BALB/c Mice

Porphyromonas gingivalis is a Gram-negative bacterium strongly associated with chronic periodontitis, an inflammatory oral disease. A major virulence factor common to all characterized strains of P. gingivalis is the RgpA-Kgp proteinase-adhesin complexes (RgpA-Kgp complexes). In this study, we investigated T cell proliferative and cytokine responses to the RgpA-Kgp complexes and identified T cell epitopes in BALB/c mice utilizing Pepscan methodology. T cell proliferative responses were found to be predominantly directed toward the proteinase catalytic domains. Eleven T cell epitopes were identified using RgpA-Kgp-primed lymph node T cells (IL-4 dominant) and 21 using an RgpA-Kgp-specific T cell line (IFN-gamma dominant), with 5 T cell epitopes, including the immunodominant epitope peptide 22, common to both T cell populations. Peptide 22 ((439)ANYTAHGSETAWADP(453)) from the Kgp proteinase catalytic domain induced a Th2 cytokine response in mice, and peptide 22-primed T cells had a Th2 cytokine profile when stimulated with the RgpA-Kgp complexes. Truncation and alanine scanning of peptide 22 identified the minimum epitope ((442)TAHGSETAWA(451)), and residues His(444), Glu(447), and Trp(450) as critical for T cell proliferation. With a view to vaccine development, peptide 22 was incorporated into a synthetic peptide polymer. Peptide 22 polymer induced strong T cell proliferation and crossreactivity to native RgpA-Kgp complexes. In conclusion, we have identified a major T cell epitope of P. gingivalis and established that antigenicity of the T cell epitope is retained when delivered as a peptide polymer. The strategies employed here may have potential in the development of a synthetic peptide vaccine for P. gingivalis.

Gender-Dependent HLA-DR-Restricted Epitopes Identified from Herpes Simplex Virus Type 1 Glycoprotein D

In recent clinical trials, a herpes simplex virus (HSV) recombinant glycoprotein D (gD) vaccine was more efficacious in woman than in men. Here we report six HLA-DR-restricted T-cell gD epitope peptides that bind to multiple HLA-DR (DR1, DR4, DR7, DR13, DR15, and DRB5) molecules that represent a large proportion of the human population. Four of these peptides recalled naturally primed CD4(+) T cells in up to 45% of the 46 HSV-seropositive, asymptomatic individuals studied. For the gD(49-82), gD(77-104), and gD(121-152) peptides, the CD4(+) T-cell responses detected in HSV-seropositive, asymptomatic women were higher and more frequent than the responses detected in men. Immunization of susceptible DRB1*0101 transgenic mice with a mixture of three newly identified, gender-dependent, immunodominant epitope peptides (gD(49-82), gD(77-104), and gD(121-152)) induced a gender- and CD4(+) T-cell-dependent immunity against ocular HSV type 1 challenge. These results revealed a gender-dependent T-cell response to a discrete set of gD epitopes and suggest that while a T-cell epitope-based HSV vaccine that targets a large percentage of the human population may be feasible with a limited number of immunodominant promiscuous HLA-DR-restricted epitopes, gender should be taken into account during evaluations of such vaccines.

Human population-based identification of CD4 + T-cell peptide epitope determinants

A human cell-based method to identify functional CD4 + T-cell epitopes in any protein has been developed. Proteins are tested as synthetic 15-mer peptides offset by three amino acids. Percent responses within a large donor population are tabulated for each peptide in the set. Peptide epitope regions are designated by difference in response frequency from the overall background response rate for the compiled dataset. Epitope peptide responses are reproducible, with a median coefficient of variance of 21% when tested on multiple random-donor sets. The overall average response rate within the dataset increases with increasing putative human population antigenic exposure to a given protein. The background rate was high for HPV16 E6, and was low for human-derived cytokine proteins. The assay identified recall epitope regions within the donor population for the protein staphylokinase. For an industrial protease with minimal presumed population exposure, immunodominant epitope peptides were identified that were found to bind promiscuously to many HLA class II molecules in vitro. The peptide epitope regions identified in presumably unexposed donors represent a subset of the total recall epitopes. Finally, as a negative control, the assay found no peptide epitope regions in human β2-microglobulin. This method identifies functional CD4 + T-cell epitopes in any protein without pre-selection for HLA class II, suggests whether a donor population is pre-exposed to a protein of interest, and does not require sensitized donors for in vitro testing.

Cryptic epitopes on the nicotinic acetylcholine receptor are recognized by autoreactive CD4+ cells.

Experimental autoimmune myasthenia gravis is induced in C57BL/6 mice by injection of Torpedo nicotinic acetylcholine receptor (TAChR). We investigated here the presence of cryptic CD4+ epitopes on the TAChR molecule, and their relationship with potentially autoreactive CD4+ cells, which survived clonal deletion. CD4+ cells from C57BL/6 mice immunized with native or denatured TAChR were challenged in vitro with overlapping synthetic peptides, 20-residue long, screening the sequences of TAChR alpha, gamma, and delta subunits. Only three epitopes on the alpha subunit were recognized consistently. Mice immunized with large doses (nanomoles) of TAChR clearly recognized only the immunodominant sequence T alpha 150-169. Anti-TAChR CD4+ cells did not cross-react with murine alpha subunit sequences, or with any synthetic sequence of human gamma and delta subunits, which are very similar to the corresponding murine subunits. To facilitate recognition of cryptic epitopes, we injected mice with pools of synthetic peptides corresponding to the sequences of TAChR alpha, gamma, and delta subunits. In addition to the three immunodominant alpha subunit epitopes, other epitopes were recognized by CD4+ cells within the sequences T alpha 304-322, T gamma 105-124, T gamma 120-139, T gamma 401-420, T gamma 357-376, T delta 16-35, T delta 61-80, T delta 121-140, and T delta 301-320. CD4+ cells thus sensitized cross-reacted with the mammalian sequences alpha 304-322, gamma 105-124, gamma 120-139, and delta 301-320. Mice were immunized with large doses (approximately 40 nmol) of individual TAChR synthetic cryptic epitopes. CD4+ cells sensitized to five cryptic epitopes (the ones listed above plus delta 121-140) cross-reacted with autologous sequences. We determined the dose dependence of the sensitization of CD4+ cells in vivo to the strongly immunodominant epitope peptide T alpha 150-169 and to the cryptic epitope peptides T gamma 120-139 and T delta 301-320 by immunizing mice with increasing doses of peptide (approximately 1.2 to approximately 20 nmol), and testing the in vitro anti-peptide response of the CD4+ cells. No difference was found for the epitopes tested. Doses of 3 to 10 micrograms induced a strong CD4+ sensitization, and the dose dependence of the in vitro response of the sensitized cells to the relevant peptide was comparable. Production of cryptic epitopes upon in vitro TAChR processing was investigated by testing peptide-sensitized CD4+ cells with native TAChR: only two cryptic epitopes were produced.