Development of a Trivalent Construct Omp18/AhpC/FlgH Multi Epitope Peptide Vaccine Against Campylobacter jejuni

Campylobacter jejuni (C. jejuni) is one of the major pathogens contributing to the enteritis in humans. Infection can lead to numerous complications, including but not limited to Guillain-Barre syndrome, reactive arthritis, and Reiter’s syndrome. Over the past two decades, joint efforts have been made toward developing a proper strategy of limiting the transmission of C. jejuni to humans. Nevertheless, except for biosecurity measures, no available vaccine has been developed so far. Judging from the research findings, Omp18, AhpC outer membrane protein, and FlgH flagellin subunits of C. jejuni could be adopted as surface protein antigens of C. jejuni for screening dominant epitope thanks to their strong antigenicity, expression of varying strains, and conservative sequence. In this study, bioinformatics technology was adopted to analyze the T-B antigenic epitopes of Omp18, AhpC, and FlgH in C. jejuni strain NCTC11168. Both ELISA and Western Blot methods were adopted to screen the dominant T-B combined epitope. GGS (GGCGGTAGC) sequence was adopted to connect the dominant T-B combined epitope peptides and to construct the prokaryotic expression system of tandem repeats of antigenic epitope peptides. The mouse infection model was adopted to assess the immunoprotective effect imposed by the trivalent T-B combined with antigen epitope peptide based on Omp18/AhpC/FlgH. In this study, a tandem epitope AhpC-2/Omp18-1/FlgH-1 was developed, which was composed of three epitopes and could effectively enhance the stability and antigenicity of the epitope while preserving its structure. The immunization of BALB/c mice with a tandem epitope could induce protective immunity accompanied by the generation of IgG2a antibody response through the in vitro synthesis of IFN-γ cytokines. Judging from the results of immune protection experiments, the colonization of C. jejuni declined to a significant extent, and it was expected that AhpC-2/Omp18-1/FlgH-1 could be adopted as a candidate antigen for genetic engineering vaccine of C. jejuni MAP.

Myelin Peptide–Mannan Conjugate Multiple Sclerosis Vaccines: Conjugation Efficacy and Stability of Vaccine Ingredient

Myelin peptide–mannan conjugates have been shown to be potential vaccines in the immunotherapy of multiple sclerosis. The conjugates are comprised from the epitope peptide and the polysaccharide mannan which transfers as a carrier the antigenic peptide to dendritic cells that process and present antigenic peptides at their surface in complex with MHC class I or class II resulting in T-cell stimulation. The conjugation of antigenic peptide with mannan occurs through the linker (Lys–Gly)5, which connects the peptide with the oxidized mannose units of mannan. This study describes novel methods for the quantification of the vaccine ingredient peptide within the conjugate, a prerequisite for approval of clinical trials in the pursuit of multiple sclerosis therapeutics. Myelin peptides, such as MOG35–55, MBP83–99, and PLP131–145 in linear or cyclic form, as altered peptide ligands or conjugated to appropriate carriers, possess immunomodulatory properties in experimental models and are potential candidates for clinical trials.

A Pencil-Lead Immunosensor for the Rapid Electrochemical Measurement of Anti-Diphtheria Toxin Antibodies

Diphtheria is a vaccine-preventable disease, yet immunization can wane over time to non-protective levels. We have developed a low-cost, miniaturized electroanalytical biosensor to quantify anti-diphtheria toxin (DTx) immunoglobulin G (anti-DTx IgG) antibody to minimize the risk for localized outbreaks. Two epitopes specific to DTx and recognized by antibodies generated post-vaccination were selected to create a bi-epitope peptide, biEP, by synthesizing the epitopes in tandem. The biEP peptide was conjugated to the surface of a pencil-lead electrode (PLE) integrated into a portable electrode holder. Captured anti-DTx IgG was measured by square wave voltammetry from the generation of hydroquinone (HQ) from the resulting immunocomplex. The performance of the biEP reagent presented high selectivity and specificity for DTx. Under the optimized working conditions, a logarithmic calibration curve showed good linearity over the concentration range of 10−5–10−1 IU mL−1 and achieved a limit of detection of 5 × 10−6 IU mL−1. The final device proved suitable for interrogating the immunity level against DTx in actual serum samples. Results showed good agreement with those obtained from a commercial enzyme-linked immunosorbent assay. In addition, the flexibility for conjugating other capture molecules to PLEs suggests that this technology could be easily adapted to the diagnoses of other pathogens.

Reversion analysis reveals the in vivo immunogenicity of a poorly MHC I-binding cancer neoepitope

High-affinity MHC I-peptide interactions are considered essential for immunogenicity. However, some neo-epitopes with low affinity for MHC I have been reported to elicit CD8 T cell dependent tumor rejection in immunization-challenge studies. Here we show in a mouse model that a neo-epitope that poorly binds to MHC I is able to enhance the immunogenicity of a tumor in the absence of immunization. Fibrosarcoma cells with a naturally occurring mutation are edited to their wild type counterpart; the mutation is then re-introduced in order to obtain a cell line that is genetically identical to the wild type except for the neo-epitope-encoding mutation. Upon transplantation into syngeneic mice, all three cell lines form tumors that are infiltrated with activated T cells. However, lymphocytes from the two tumors that harbor the mutation show significantly stronger transcriptional signatures of cytotoxicity and TCR engagement, and induce greater breadth of TCR reactivity than those of the wild type tumors. Structural modeling of the neo-epitope peptide/MHC I pairs suggests increased hydrophobicity of the neo-epitope surface, consistent with higher TCR reactivity. These results confirm the in vivo immunogenicity of low affinity or ‘non-binding’ epitopes that do not follow the canonical concept of MHC I-peptide recognition.

A novel design of multi-epitope peptide vaccine against Pseudomonas aeruginosa

Background: As an opportunistic pathogen, Pseudomonas aeruginosa causes many different hazardous infections. The high mortality rate resulting from infection with this antibiotic-resistant pathogen has made it a major challenge in clinical treatment; it has been listed as the most harmful bacterium to humans by the WHO. So far, no vaccine has been approved for P. aeruginosa. Objective: Infections performed by bacterial attachment and colonization with type IV pili (T4P), known as the most essential adhesive vital for adhesion, while pilQ is necessary for the biogenesis of T4P, also outer membrane proteins of a pathogen is also effective in stimulating the immune system; in this regard, pilQ, OprF, and OprI, are excellent candidate antigens for production of an effective vaccine against P. aeruginosa. Methods: In this research, various bioinformatics methods were employed in order to design a new multi-epitope peptide vaccine versus P. aeruginosa. Since T CD4+ cell immunity is important in eradicating P. aeruginosa, OprF, OprI, and pilQ antigens were analyzed to determineHelper T cell Lymphocyte (HTL) epitopes by many different immunoinformatics servers. One of the receptor agonists 2 (TLR2), a segment of the Por B protein from Neisseria meningitides was used as an adjuvant in order to stimulate an effective cellular immune response, and suitable linkers were used to connect all the above mentioned parts. In the vaccine construct, linear B cell epitopes were also identified. Results: Conforming the bioinformatics forecasts, the designed vaccine possesses high antigenicity and is not allergen. Conclusion: In this regard, the designed vaccine candidate is strongly believed to possess the potential of inducing cellular and humoral immunity against P. aeruginosa.