Helper T-cell Epitopes Research Articles

Proteomics-based vaccine targets annotation and design of subunit and mRNA-based vaccines for Monkeypox virus (MPXV) against the recent outbreak

Monkeypox Virus (MPXV) is a growing public health threat with increasing cases and fatalities globally. To date, no specific vaccine or small molecule therapeutic choices are available for the treatment of MPXV disease. In this work, we employed proteomics and structural vaccinology approaches to design mRNA and multi-epitopes-based vaccines (MVC) against MPXV. We first identified ten proteins from the whole proteome of MPXV as potential vaccine targets. We then employed structural vaccinology approaches to map potential epitopes of these proteins for B cell, cytotoxic T lymphocytes (CTL), and Helper T lymphocytes (HTL). Finally, 9 CTL, 6 B cell, and 5 HTL epitopes were joined together through suitable linkers to construct MVC (multi-epitope vaccine) and mRNA-based vaccines. Molecular docking, binding free energy calculation, and in silico cloning revealed robust interaction of the designed MVC with toll-like receptor 2 (TLR2) and efficient expression in E. Coli K12 strain. The immune simulation results revealed that the antigen titer after the injection reached to the maximum level on the 5th day and an abrupt decline in the antigen titer was observed upon the production of IgM, IgG and IgM + IgG, dendritic cells, IFN-gamma, and IL (interleukins), which suggested the potential of our designed vaccine candidate for inducing an immune response against MPXV.

Modular adjuvant-free pan-HLA-DR-immunotargeting subunit vaccine against SARS-CoV-2 elicits broad sarbecovirus-neutralizing antibody responses

Subunit vaccines typically require co-administration with an adjuvant to elicit protective immunity, adding development hurdles that can impede rapid pandemic responses. To circumvent the need for adjuvant in a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) subunit vaccine, we engineer a thermostable immunotargeting vaccine (ITV) that leverages the pan-HLA-DR monoclonal antibody 44H10 to deliver the viral spike protein receptor-binding domain (RBD) to antigen-presenting cells. X-ray crystallography shows that 44H10 binds to a conserved epitope on HLA-DR, providing the basis for its broad HLA-DR reactivity. Adjuvant-free ITV immunization in rabbits and ferrets induces robust anti-RBD antibody responses that neutralize SARS-CoV-2 variants of concern and protect recipients from SARS-CoV-2 challenge. We demonstrate that the modular nature of the ITV scaffold with respect to helper Tcell epitopes and diverse RBD antigens facilitates broad sarbecovirus neutralization. Our findings support anti-HLA-DR immunotargeting as an effective means to induce strong antibody responses to subunit antigens without requiring an adjuvant.

Exploring the structural basis to develop efficient multi-epitope vaccines displaying interaction with HLA and TAP and TLR3 molecules to prevent NIPAH infection, a global threat to human health.

Nipah virus (NiV) is an emerging zoonotic virus that caused several serious outbreaks in the south asian region with high mortality rates ranging from 40 to 90% since 2001. NiV infection causes lethal encephalitis and respiratory disease with the symptom of endothelial cell-cell fusion. No specific and effective vaccine has yet been reported against NiV. To address the urgent need for a specific and effective vaccine against NiV infection, in the present study, we have designed two Multi-Epitope Vaccines (MEVs) composed of 33 Cytotoxic T lymphocyte (CTL) epitopes and 38 Helper T lymphocyte (HTL) epitopes. Out of those CTL and HTL combined 71 epitopes, 61 novel epitopes targeting nine different NiV proteins were not used before for vaccine design. Codon optimization for the cDNA of both the designed MEVs might ensure high expression potential in the human cell line as stable proteins. Both MEVs carry potential B cell linear epitope overlapping regions, B cell discontinuous epitopes as well as IFN-γ inducing epitopes. Additional criteria such as sequence consensus amongst CTL, HTL and B Cell epitopes was implemented for the design of final constructs constituting MEVs. Hence, the designed MEVs carry the potential to elicit cell-mediated as well as humoral immune response. Selected overlapping CTL and HTL epitopes were validated for their stable molecular interactions with HLA class I and II alleles and in case of CTL epitopes with human Transporter Associated with antigen Processing (TAP) cavity. The structure based epitope cross validation for interaction with TAP cavity was used as another criteria choosing final epitopes for NiV MEVs. Finally, human Beta-defensin 2 and Beta-defensin 3 were used as adjuvants to enhance the immune response of both the MEVs. Molecular dynamics simulation studies of MEVs-TLR3 ectodomain (Human Toll-Like Receptor 3) complex indicated the stable molecular interaction. We conclude that the MEVs designed and in silico validated here could be highly potential vaccine candidates to combat NiV infections, with great effectiveness, high specificity and large human population coverage worldwide.

Exploring staphylococcal superantigens to design a potential multi-epitope vaccine against Staphylococcus aureus: an in-silico reverse vaccinology approach

Staphylococcus aureus is a horrifying bacteria capable of causing millions of deaths yearly across the globe. A major contribution to the success of S. aureus as an ESKAPE pathogen is the abundance of virulence factors that can manipulate the innate and adaptive immune system of the individual. Currently, no vaccine is available to treat S. aureus-mediated infections. In this study, we present in-silico approaches to design a stable, safe and immunogenic vaccine that could help to control the infections associated with the bacteria. Three vital pathogenic secreted toxins of S. aureus, such as staphylococcal enterotoxin A (SEA), staphylococcal enterotoxin B (SEB), Toxic-shock syndrome toxin (TSST-1), were selected using the reverse vaccinology approach to design the multi-epitope vaccine (MEV). Linear B-lymphocyte, cytotoxic T-lymphocyte (CTL) and helper T-lymphocyte (HTL) epitopes were predicted from these selected proteins. For designing the multi-epitope vaccine (MEV), B-cell epitopes were joined with the KK linker, CTL epitopes were joined with the AAY linker, and HTL epitopes were joined with the GPGPG linker. Finally, to increase the immune response to the vaccine, a human β-defensin-3 (hBD-3) adjuvant was added to the N-terminus of the MEV construct. The final MEV was found to be antigenic and non-allergen in nature. In-silico immune simulation and cloning analysis predicted the immune-stimulating potential of the designed MEV construct along with the cloning feasibility in the pET28a(+) vector with the E. coli expression system. This immunoinformatics study provides a platform for designing a suitable, safe and effective vaccine against S. aureus. Communicated by Ramaswamy H. Sarma

Development of synthetic, self-adjuvanting, and self-assembling anticancer vaccines based on a minimal saponin adjuvant and the tumor-associated MUC1 antigen.

The overexpression of aberrantly glycosylated tumor-associated mucin-1 (TA-MUC1) in human cancers makes it a major target for the development of anticancer vaccines derived from synthetic MUC1-(glyco)peptide antigens. However, glycopeptide-based subunit vaccines are weakly immunogenic, requiring adjuvants and/or additional immunopotentiating approaches to generate optimal immune responses. Among these strategies, unimolecular self-adjuvanting vaccine constructs that do not need coadministration of adjuvants or conjugation to carrier proteins emerge as a promising but still underexploited approach. Herein, we report the design, synthesis, immune-evaluation in mice, and NMR studies of new, self-adjuvanting and self-assembling vaccines based on our QS-21-derived minimal adjuvant platform covalently linked to TA-MUC1-(glyco)peptide antigens and a peptide helper T-cell epitope. We have developed a modular, chemoselective strategy that harnesses two distal attachment points on the saponin adjuvant to conjugate the respective components in unprotected form and high yields via orthogonal ligations. In mice, only tri-component candidates but not unconjugated or di-component combinations induced significant TA-MUC1-specific IgG antibodies able to recognize the TA-MUC1 on cancer cells. NMR studies revealed the formation of self-assembled aggregates, in which the more hydrophilic TA-MUC1 moiety gets exposed to the solvent, favoring B-cell recognition. While dilution of the di-component saponin-(Tn)MUC1 constructs resulted in partial aggregate disruption, this was not observed for the more stably-organized tri-component candidates. This higher structural stability in solution correlates with their increased immunogenicity and suggests a longer half-life of the construct in physiological media, which together with the enhanced antigen multivalent presentation enabled by the particulate self-assembly, points to this self-adjuvanting tri-component vaccine as a promising synthetic candidate for further development.

Immunoinformatics analysis of candidate proteins for controlling bovine paratuberculosis.

Paratuberculosis is debilitating chronic enteritis usually characterized by diarrhea, decreased milk production, and progressive cachexia. Mycobacterium avium subspecies paratuberculosis (MAP) causes significant economic losses by affecting dairy herds globally. Development of protective vaccines is considered as one of the most effective controlling measures for MAP infections. In the current study, hydrophilic parts of MAP2191 and FAP-P proteins as two vaccine candidates were analyzed using immunoinformatics approaches. After selecting the most hydrophilic parts of MAP2191 and FAP-P, helper and cytotoxic T-cell epitopes of ht-MAP2191 and ht-FAP-P were identified. The immunogenic, toxicity and physicochemical properties were assessed. Secondary structures of these proteins were predicted, and their tertiary structures were modeled, refined, and validated. Linear and conformational epitopes of corresponding B-cells were recognized. Then ht-MAP2191 and ht-FAP-P epitopes were employed for molecular docking simulations. The results indicated that ht-MAP2191 and ht-FAP-P were immunogenic, non-allergenic, and non-toxic and possess potent T-cell and B-cell epitopes. Eventually, these protein constructs were docked favorably against TLR4. According to the findings, ht-MAP2191 and ht-FAP-P could be effective protein-based vaccine candidates for paratuberculosis. It should be noted that to examine their efficacy, further in vitro and in vivo experiments are underway.

Immunoinformatics-Aided Design of a Peptide Based Multiepitope Vaccine Targeting Glycoproteins and Membrane Proteins against Monkeypox Virus.

Monkeypox is a self-limiting zoonotic viral disease and causes smallpox-like symptoms. The disease has a case fatality ratio of 3-6% and, recently, a multi-country outbreak of the disease has occurred. The currently available vaccines that have provided immunization against monkeypox are classified as live attenuated vaccinia virus-based vaccines, which pose challenges of safety and efficacy in chronic infections. In this study, we have used an immunoinformatics-aided design of a multi-epitope vaccine (MEV) candidate by targeting monkeypox virus (MPXV) glycoproteins and membrane proteins. From these proteins, seven epitopes (two T-helper cell epitopes, four T-cytotoxic cell epitopes and one linear B cell epitopes) were finally selected and predicted as antigenic, non-allergic, interferon-γ activating and non-toxic. These epitopes were linked to adjuvants to design a non-allergic and antigenic candidate MPXV-MEV. Further, molecular docking and molecular dynamics simulations predicted stable interactions between predicted MEV and human receptor TLR5. Finally, the immune-simulation analysis showed that the candidate MPXV-MEV could elicit a human immune response. The results obtained from these in silico experiments are promising but require further validation through additional in vivo experiments.

Bioinformatics-based SARS-CoV-2 epitopes design and the impact of spike protein mutants on epitope humoral immunities

BackgroundEpitope selection is the key to peptide vaccines development. Bioinformatics tools can efficiently improve the screening of antigenic epitopes and help to choose the right ones. ObjectiveTo predict, synthesize and testify peptide epitopes at spike protein, assess the effect of mutations on epitope humoral immunity, thus provide clues for the design and development of epitope peptide vaccines against SARS-CoV-2. MethodsBioinformatics servers and immunological tools were used to identify the helper T lymphocyte, cytotoxic T lymphocyte, and linear B lymphocyte epitopes on the S protein of SARS-CoV-2. Physicochemical properties of candidate epitopes were analyzed using IEDB, VaxiJen, and AllerTOP online software. Three candidate epitopes were synthesized and their antigenic responses were evaluated by binding antibody detection. ResultsA total of 20 antigenic, non-toxic and non-allergenic candidate epitopes were identified from 1502 epitopes, including 6 helper T-cell epitopes, 13 cytotoxic T-cell epitopes, and 1 linear B cell epitope. After immunization with antigen containing candidate epitopes S206-221, S403-425, and S1157-1170 in rabbits, the binding titers of serum antibody to the corresponding peptide, S protein, receptor-binding domain protein were (415044, 2582, 209.3), (852819, 45238, 457767) and (357897, 10528, 13.79), respectively. The binding titers to Omicron S protein were 642, 12,878 and 7750, respectively, showing that N211L, DEL212 and K417N mutations cause the reduction of the antibody binding activity. ConclusionsBioinformatic methods are effective in peptide epitopes design. Certain mutations of the Omicron would lead to the loss of antibody affinity to Omicron S protein.

A subtractive proteomics and immunoinformatics approach towards designing a potential multi-epitope vaccine against pathogenic Listeriamonocytogenes

Listeria monocytogenes is the causative agent of listeriosis, which is dangerous for pregnant women, the elderly or individuals with a weakened immune system. Individuals with leukaemia, cancer, HIV/AIDS, kidney transplant and steroid therapy suffer from immunological damage are menaced. World Health Organization (WHO) reports that human listeriosis has a high mortality rate of 20–30% every year. To date, no vaccine is available to treat listeriosis. Thereby, it is high time to design novel vaccines against L. monocytogenes. Here, we present computational approaches to design an antigenic, stable and safe vaccine against the L. monocytogenes that could help to control the infections associated with the pathogen. Three vital pathogenic proteins of L. monocytogenes, such as Listeriolysin O (LLO), Phosphatidylinositol-specific phospholipase C (PI-PLC), and Actin polymerization protein (ActA), were selected using a subtractive proteomics approach to design the multi-epitope vaccine (MEV). A total of 5 Cytotoxic T-lymphocyte (CTL) and 9 Helper T-lymphocyte (HTL) epitopes were predicted from these selected proteins. To design the multi-epitope vaccine (MEV) from the selected proteins, CTL epitopes were joined with the AAY linker, and HTL epitopes were joined with the GPGPG linker. Additionally, a human β-defensin-3 (hBD-3) adjuvant was added to the N-terminal side of the final MEV construct to increase the immune response to the vaccine. The final MEV was predicted to be antigenic, non-allergen and non-toxic in nature. Physicochemical property analysis suggested that the MEV construct is stable and could be easily purified through the E. coli expression system. This in-silico study showed that MEV has a robust binding interaction with Toll-like receptor 2 (TLR2), a key player in the innate immune system. Current subtractive proteomics and immunoinformatics study provides a background for designing a suitable, safe and effective vaccine against pathogenic L. monocytogenes.

In silico designing of a novel epitope-based candidate vaccine against Streptococcus pneumoniae with introduction of a new domain of PepO as adjuvant

BackgroundStreptococcus pneumoniae is the leading reason for invasive diseases including pneumonia and meningitis, and also secondary infections following viral respiratory diseases such as flu and COVID-19. Currently, serotype-dependent vaccines, which have several insufficiency and limitations, are the only way to prevent pneumococcal infections. Hence, it is plain to need an alternative effective strategy for prevention of this organism. Protein-based vaccine involving conserved pneumococcal protein antigens with different roles in virulence could provide an eligible alternative to existing vaccines.MethodsIn this study, PspC, PhtD and PsaA antigens from pneumococcus were taken to account to predict B-cell and helper T-cell epitopes, and epitope-rich regions were chosen to build the construct. To enhance the immunogenicity of the epitope-based vaccine, a truncated N-terminal fragment of pneumococcal endopeptidase O (PepO) was used as a potential TLR2/4 agonist which was identified by molecular docking studies. The ultimate construct was consisted of the chosen epitope-rich regions, along with the adjuvant role (truncated N-PepO) and suitable linkers.ResultsThe epitope-based vaccine was assessed as regards physicochemical properties, allergenicity, antigenicity, and toxicity. The 3D structure of the engineered construct was modeled, refined, and validated. Molecular docking and simulation of molecular dynamics (MD) indicated the proper and stable interactions between the vaccine and TLR2/4 throughout the simulation periods.ConclusionsFor the first time this work presents a novel vaccine consisting of epitopes of PspC, PhtD, and PsaA antigens which is adjuvanted with a new truncated domain of PepO. The computational outcomes revealed that the suggested vaccine could be deemed an efficient therapeutic vaccine for S. pneumoniae; nevertheless, in vitro and in vivo examinations should be performed to prove the potency of the candidate vaccine.

Screening of immune epitope in the proteome of the Dabie bandavirus, SFTS, to design a protein-specific and proteome-wide vaccine for immune response instigation using an immunoinformatics approaches

Tick-borne viruses are a major risk from tick bites, which could result in viral infectious diseases among animals and humans. Bunyavirus causes severe fever with thrombocytopenia syndrome (SFTS), with signs and symptoms including high fever, vomiting, diarrhea, thrombocytopenia (low platelet count), leukopenia (low white blood cell count), elevated liver enzyme levels, multiple organ failure, and has a 6%–30% case-fatality rate. To date no effective drug or vaccines are available thus need urgent research for therapeutics formulation. Hence, in this study, the computational meta-analysis approach was implemented that incorporates immunoinformatics to find potential B-cell, HTL (helper T lymphocytes) and T-cell epitopes derived from antigenic SFTS proteins to design multi-epitopes vaccines for the treatment of SFTS. The predicted T cell, B cell and HTL epitopes were shortlisted and checked for antigenic properties and allergenic features. The best epitopes were then joined together to model of multi-epitopes vaccines for specific proteins (replicase and glycoprotein) and proteome wide. The constructed models were validated using in silico molecular docking approach to evaluate binding potential of the designed best constructs with TLR3 (toll like receptor 3). Following the MEVC (multi-epitopes vaccine construct) injection, the response of the immune system was significantly stimulated, and anti-toxicity of induced antibodies was tremendously enhanced. Before being neutralized, the antigen titers remained high 5–10 days after injection of replicase, glycoprotein and proteome wide constructed vaccines. For each antigenic vaccine, a significant antibody response induction was observed. Further, in vivo trials are required to affirm the effectiveness of the constructed vaccine against SFTS.

Potent anti-tumor immune response and tumor growth inhibition induced by HER2 subdomain fusion protein in a mouse tumor model.

Several approaches have so far been employed to establish anti-tumor immunity by targeting HER2 protein. Active immunization with recombinant HER2 subdomains has previously been demonstrated to induce potent immune response and tumor growth inhibition. In the present study, we investigated the immunogenicity and tumor inhibitory effect of a fusion protein consisting of human HER2 extracellular subdomain (ECD-DI + II) together with T-helper cell epitopes of Tetanus toxin (p2 and p30). BALB/c mice were immunized with two recombinant proteins (DI + II and p2p30-DI + II) emulsified in 4 different adjuvants. Anti-DI + II antibody response, cytokine profile, frequency of splenic CD25+FOXP3+ regulatory T cells (Tregs) and CD8+CD107a+ cytotoxic T lymphocytes (CTLs) were assessed in the immunized mice. To assess the anti-tumor effect, the immunized mice were subcutaneously challenged with HER2-overexpressing tumor cells and the tumor growth was determined. Both recombinant proteins were able to induce comparable levels of ECD-DI + II-specific antibodies. Immunization with p2p30-DI + II resulted in a significant increase in the level of Interferon-gamma (IFN-γ) secretion compared to DI + II protein and significantly higher frequency of CTLs and lower frequency of Tregs. The number of mice that remained tumor-free until day 120 was significantly higher in p2p30-DI + II vaccinated groups. Our data suggest that the p2p30-DI + II fusion protein together with CpG adjuvant induces more potent anti-tumor immune responses in a mouse tumor model. Accordingly, this formulation might be considered as a potential immunotherapeutic approach in HER2+ cancers.

Design of a multi-epitope vaccine candidate against Brucella melitensis

Brucella is a typical facultative intracellular bacterium that can cause zoonotic infections. For Brucella, it is difficult to eliminate with current medical treatment. Therefore, a multi-epitope vaccine (MEV) should be designed to prevent Brucella infection. For this purpose, we applied the reverse vaccinology approach from Omp10, Omp25, Omp31 and BtpB. Finally, we obtained 13 cytotoxic T lymphocyte (CTL) epitopes, 17 helper T lymphocyte (HTL) epitopes, 9 linear B cell epitopes, and 2 conformational B cell epitopes for further study. To keep the protein folded normally, we linked AAY, GPGPG, and KK to CTL epitopes, HTL epitopes, and B cell epitopes, respectively. The N-terminal of the vaccine peptide is supplemented with appropriate adjuvants to enhance immunogenicity. To evaluate its immunogenicity, stability, safety, and feasibility, a final MEV containing 806 amino acids was constructed by linking linkers and adjuvants. In addition, molecular docking and molecular dynamics simulations were performed to verify the affinity and stability of the MEV-TLR4. Then, codon adaptation and in silico cloning studies were carried out to identify the possible codons for expressing the MEV. In animal experiments, the results demonstrated that the MEV had high immunogenicity. Collectively, this study provided a theoretical basis for the development of a Brucella vaccine.

Co-Administration of Adjuvanted Recombinant Ov-103 and Ov-RAL-2 Vaccines Confer Protection against Natural Challenge in A Bovine Onchocerca ochengi Infection Model of Human Onchocerciasis.

Onchocerciasis (river blindness), caused by the filarial nematode Onchocerca volvulus, is a neglected tropical disease mainly of sub-Saharan Africa. Worldwide, an estimated 20.9 million individuals live with infection and a further 205 million are at risk of disease. Current control methods rely on mass drug administration of ivermectin to kill microfilariae and inhibit female worm fecundity. The identification and development of efficacious vaccines as complementary preventive tools to support ongoing elimination efforts are therefore an important objective of onchocerciasis research. We evaluated the protective effects of co-administering leading O. volvulus-derived recombinant vaccine candidates (Ov-103 and Ov-RAL-2) with subsequent natural exposure to the closely related cattle parasite Onchocerca ochengi. Over a 24-month exposure period, vaccinated calves (n = 11) were shown to acquire infection and microfilaridermia at a significantly lower rate compared to unvaccinated control animals (n = 10). Furthermore, adult female worm burdens were negatively correlated with anti-Ov-103 and Ov-RAL-2 IgG1 and IgG2 responses. Peptide arrays identified several Ov-103 and Ov-RAL-2-specific epitopes homologous to those identified as human B-cell and helper T-cell epitope candidates and by naturally-infected human subjects in previous studies. Overall, this study demonstrates co-administration of Ov-103 and Ov-RAL-2 with Montanide™ ISA 206 VG is highly immunogenic in cattle, conferring partial protection against natural challenge with O. ochengi. The strong, antigen-specific IgG1 and IgG2 responses associated with vaccine-induced protection are highly suggestive of a mixed Th1/Th2 associated antibody responses. Collectively, this evidence suggests vaccine formulations for human onchocerciasis should aim to elicit similarly balanced Th1/Th2 immune responses.

Design of a Recombinant Multivalent Epitope Vaccine Based on SARS-CoV-2 and Its Variants in Immunoinformatics Approaches

The development of an effective multivalent vaccine against SARS-CoV-2 variants is an important means to improve the global public health situation caused by COVID-19. In this study, we identified the antigen epitopes of the main global epidemic SARS-CoV-2 and mutated virus strains using immunoinformatics approach, and screened out 8 cytotoxic T lymphocyte epitopes (CTLEs), 17 helper T lymphocyte epitopes (HTLEs), 9 linear B-cell epitopes (LBEs) and 4 conformational B-cell epitopes (CBEs). The global population coverage of CTLEs and HTLEs was 93.16% and 99.9% respectively. These epitopes were spliced together by corresponding linkers and recombined into multivalent vaccine. In silico tests, the vaccine protein was a non-allergen and the docking with TLR-3 molecule showed a strong interaction. The results of immune simulation showed that the vaccine may be helpful to initiate both cellular and humoral immunity against all VOC. The optimistic immunogenicity of the vaccine was confirmed in vivo and in vitro finally. Therefore, our vaccine may have potential protection against SARS-CoV-2 and its variants.

Potential Immunogenic Activity of Computationally Designed mRNA- and Peptide-Based Prophylactic Vaccines against MERS, SARS-CoV, and SARS-CoV-2: A Reverse Vaccinology Approach.

The continued emergence of human coronaviruses (hCoVs) in the last few decades has posed an alarming situation and requires advanced cross-protective strategies against these pandemic viruses. Among these, Middle East Respiratory Syndrome coronavirus (MERS-CoV), Severe Acute Respiratory Syndrome coronavirus (SARS-CoV), and Severe Acute Respiratory Syndrome coronavirus-2 (SARS-CoV-2) have been highly associated with lethality in humans. Despite the challenges posed by these viruses, it is imperative to develop effective antiviral therapeutics and vaccines for these human-infecting viruses. The proteomic similarity between the receptor-binding domains (RBDs) among the three viral species offers a potential target for advanced cross-protective vaccine designs. In this study, putative immunogenic epitopes including Cytotoxic T Lymphocytes (CTLs), Helper T Lymphocytes (HTLs), and Beta-cells (B-cells) were predicted for each RBD-containing region of the three highly pathogenic hCoVs. This was followed by the structural organization of peptide- and mRNA-based prophylactic vaccine designs. The validated 3D structures of these epitope-based vaccine designs were subjected to molecular docking with human TLR4. Furthermore, the CTL and HTL epitopes were processed for binding with respective human Lymphocytes Antigens (HLAs). In silico cloning designs were obtained for the prophylactic vaccine designs and may be useful in further experimental designs. Additionally, the epitope-based vaccine designs were evaluated for immunogenic activity through immune simulation. Further studies may clarify the safety and efficacy of these prophylactic vaccine designs through experimental testing against these human-pathogenic coronaviruses.

Development of peptide vaccine candidate using highly antigenic PE-PGRS family proteins to stimulate the host immune response against Mycobacterium tuberculosis H37Rv: an immuno-informatics approach

Tuberculosis (TB) is a fast spreading; transmissible disease caused by the Mycobacterium tuberculosis (M. tuberculosis). M. tuberculosis has a high death rate in its endemic regions due to a lack of appropriate treatment and preventative measures. We have used a vaccinomics strategy to create an effective multi-epitope vaccine against M. tuberculosis. The antigenic proteins with the highest antigenicity were utilised to predict cytotoxic T-lymphocyte (CTL), helper T-lymphocyte (HTL), and linear B-lymphocyte (LBL) epitopes. CTL and HTL epitopes were covered in 99.97% of the population. Seven epitopes each of CTL, HTL, and LBL were ultimately selected and utilised to develop a multi-epitope vaccine. A vaccine design was developed by combining these epitopes with suitable linkers and LprG adjuvant. The vaccine chimera was revealed to be highly immunogenic, non-allergenic, and non-toxic. To ensure a better expression within the Escherichia coli K12 (E. coli K12) host system, codon adaptation and in silico cloning were accomplished. Following that, various validation studies were conducted, including molecular docking, molecular dynamics simulation, and immunological simulation, all of which indicated that the designed vaccine would be stable in the biological environment and effective against M. tuberculosis infection. The immune simulation revealed higher levels of T-cell and B-cell activity, which corresponded to the actual immune response. Exposure simulations were repeated several times, resulting in increased clonal selection and faster antigen clearance. These results suggest that, if proposed vaccine chimera would test both in-vitro and in-vivo, it could be a viable treatment and preventive strategy for TB. Communicated by Ramaswamy H. Sarma

Precise immunological evaluation rationalizes the design of a self-adjuvanting vaccine composed of glycan antigen, TLR1/2 ligand, and T-helper cell epitope.

Sialyl-Tn (STn), overexpressed on various tumors, has been investigated for its application in anti-cancer vaccine therapy. However, Theratope, an STn-based vaccine, failed in the phase III clinical trial due to poor immunogenicity and epitope suppression by the foreign carrier protein. We therefore developed a self-adjuvanting STn based-vaccine, a conjugate of clustered STn (triSTn) antigen, TLR1/2 ligand (Pam3CSK4), and T-helper (Th) cell epitope, and found that this three-component self-adjuvanting vaccine effectively resulted in the production of anti-triSTn IgG antibodies. We herein analyzed immune responses induced by this self-adjuvanting vaccine in detail. We newly synthesized two-component vaccines, i.e., Pam3CSK4- or Th epitope-conjugated triSTn, as references to evaluate the immune-stimulating functions of Pam3CSK4 and Th epitope. Immunological evaluation of the synthesized vaccine candidates revealed that Pam3CSK4 was essential for antibody production, indicating that the uptake of triSTn antigen by antigen-presenting cells (APCs) was promoted by the recognition of Pam3CSK4 by TLR1/2. The function of the Th epitope was also confirmed. Th cell activation was important for boosting antibody production and IgG subclass switching. Furthermore, flow cytometric analyses of immune cells, including T cells, B cells, dendritic cells, and other monocytes, were first employed in the evaluation of self-adjuvanting vaccines and revealed that the three-component vaccine was able to induce antigen-specific immune responses for efficient antibody production without excessive inflammatory responses. Importantly, the co-administration of Freund's adjuvants was suggested to cause excessive myeloid cell accumulation and decreased plasma cell differentiation. These results demonstrate that vaccines can be designed to achieve the desired immune responses via the bottom-up construction of each immune element.

In silico Design of a Crimean-Congo Hemorrhagic Fever Virus Glycoprotein Multi-Epitope Antigen for Vaccine Development.

There is no licensed vaccine available to prevent the severe tick-borne disease Crimean-Congo hemorrhagic fever (CCHF), caused by the CCHF virus (CCHFV). This study sought to show that a combination of computational methods and data from published literature can inform the design of a multi-epitope antigen for CCHFV that has the potential to be immunogenic. Cytotoxic and helper T-cell epitopes were evaluated on the CCHFV GPC using bioinformatic servers, and this data was combined with work from previous studies to identify potentially immunodominant regions of the GPC. Regions of the GPC were selected for generation of a model multi-epitope antigen in silico, and the percent residue identity and similarity of each region was compared across sequences representing the widespread geographical and ecological distribution of CCHFV. Eleven multi-epitope regions were joined together with flexible linkers in silico to generate a model multi-epitope antigen, termed EPIC, which included 812 (75.7%) of all predicted epitopes. EPIC was predicted to be antigenic by two independent bioinformatic servers, suggesting that multi-epitope antigens should be explored further for CCHFV vaccine development. The results presented within this manuscript provide information for potential targets within the CCHFV GPC for guiding future vaccine development.

An immunoinformatics-based designed multi-epitope candidate vaccine (mpme-VAC/STV-1) against Mycoplasma pneumoniae

Pneumonia is a serious global health problem that accounts for over one million deaths annually. Among the main microorganisms causing pneumonia, Mycoplasma pneumoniae is one of the most common ones for which a vaccine is immediately required. In this context, a multi-epitope vaccine against this pathogen could be the best option that can induce effective immune response avoiding any serious adverse reactions. In this study, using an immunoinformatics approach we have designed a multi-epitope vaccine (mpme-VAC/STV-1) against M. pneumoniae. Our designed mpme-VAC/STV-1 is constructed using CTL (cytotoxic T lymphocyte), HTL (Helper T lymphocyte), and B-cell epitopes. These epitopes are selected from the core proteins of 88 M. pneumoniae genomes that were previously identified through reverse vaccinology approaches. The epitopes were filtered according to their immunogenicity, population coverage, and several other criteria. Sixteen CTL/B- and thirteen HTL/B- epitopes that belong to 5 core proteins were combined together through peptide linkers to develop the mpme-VAC/STV-1. The heat-labile enterotoxin from E. coli was used as an adjuvant. The designed mpme-VAC/STV-1 is predicted to be stable, non-toxic, non-allergenic, non-host homologous, and with required antigenic and immunogenic properties. Docking and molecular dynamic simulation of mpme-VAC/STV-1 shows that it can stimulate TLR2 pathway mediated immunogenic reactions. In silico cloning of mpme-VAC/STV-1 in an expression vector also shows positive results. Finally, the mpme-VAC/STV-1 also shows promising efficacy in immune simulation tests. Therefore, our constructed mpme-VAC/STV-1 could be a safe and effective multi-epitope vaccine for immunization against pneumonia. However, it requires further experimental and clinical validations.