Contriving multiepitope subunit vaccine by exploiting structural and nonstructural viral proteins to prevent Epstein-Barr virus-associated malignancy.

Cancer is one of the common lifestyle diseases and is considered to be the leading cause of death worldwide. Epstein-Barr virus (EBV)-infected individuals remain asymptomatic; but under certain stress conditions, EBV may lead to the development of cancers such as Burkitt's and Hodgkin's lymphoma and nasopharyngeal carcinoma. EBV-associated cancers result in a large number of deaths in Asian and African population, and no effective cure has still been developed. We, therefore, tried to devise a subunit vaccine with the help of immunoinformatic approaches that can be used for the prevention of EBV-associated malignancies. The epitopes were predicted through B-cell, cytotoxic T lymphocytes (CTL), and helper T lymphocytes (HTL) from the different oncogenic proteins of EBV. A vaccine was designed by combining the B-cell and T-cell (HTL and CTL) epitopes through linkers, and for the enhancement of immunogenicity, an adjuvant was added at the N-terminal. Further, homology modeling was performed to generate the 3D structure of the designed vaccine. Moreover, molecular docking was performed between the designed vaccine and immune receptor (TLR-3) to determine the interaction between the final vaccine construct and the immune receptor complex. In addition, molecular dynamics was performed to analyze the stable interactions between the ligand final vaccine model and receptor TLR-3 molecule. Lastly, to check the expression of our vaccine construct, we performed in silico cloning. This study needed experimental validation to ensure its effectiveness and potency to control malignancy.

Hydrogen peroxide vapour is an effective replacement for Formaldehyde in a BSL4 Foot and mouth disease vaccine manufacturing facility.

An evaluation of the efficacy of 35% hydrogen peroxide vapour (HPV) against two strains of FMDV was conducted over a period of 6 months. FMDV biological indicators were produced on-site using strains obtained from a commercial FMDV vaccine manufacturing process. FMDV biological indicators were distributed within a BSL4 laboratory and exposed to short duration hydrogen peroxide cycles. Variations in titre, support matrix (soiling), temperature and humidity were evaluated in a series of 16 exposures using over 200 individual FMDV indicators. Additional verification testing was performed in an operational material transfer lock to replicate real-world use. HPV was found to be efficacious in inactivating FMDV strains; the inoculum titre influenced the level of reduction achieved with the specified cycle. SIGNIFICANCE AND IMPACT OF THE STUDY: The classification of formaldehyde as a presumed human carcinogen has presented regulatory challenges for its continued use as a biocidal product. Institutions are actively seeking fumigants to replace formaldehyde and undertaking studies to validate biocidal efficacy, particularly in high-level biosafety facilities where the consequences of pathogen release can be extremely severe. This study builds on the already substantial scientific efficacy base of 35% hydrogen peroxide vapour and provides a comprehensive evaluation of the applicability of hydrogen peroxide vapour as a replacement for formaldehyde within a Foot & Mouth Disease (FMDV) vaccine manufacturing facility.

[SYNTHETIC PEPTIDE VACCINES].

An update on the development and trials of synthetic peptide vaccines is reviewed. The review considers the successful examples of specific protection as a result of immunization with synthetic peptides using various protocols. The importance of conformation for the immunogenicity of the peptide is pointed out. An alternative strategy of the protection of the organism against the infection using synthetic peptides is suggested.

Nanovaccines and their mode of action.

Nanosized particles including nanovaccines are a novel approach to the development of vaccines to combat diseases. Nanovaccines have the promise to utilize the immune system to cure infections and to prevent infections and diseases from spreading. Rational vaccine development requires an understanding of vaccine mediated stimulation of the immune system. We review here immunostimulatory properties of nanovaccines including their immunogenicity, adjuvant properties, inflammatory responses and the mechanisms of uptake and stimulation of immune cells. Examples of various nanoparticles currently being developed as vaccines are also provided.

[Vesicular stomatitis virus (VSV) as a vaccine vector for immunization against viral infections]. / Zastosowanie wirusa pecherzykowatego zapalenia jamy ustnej (VSV) jako wektora szczepionek przeciwwirusowych.

Vesicular stomatitis virus (VSV), a member of the Rhabdoviridae family, is a promising candidate for potential use in construction of antiviral vaccines. In the natural environment VSV is a pathogen of wild ungulates and livestock. Some of the features that make VSV an excellent platform for the development of a range of viral therapeutics includes its immunogenicity and ability to grow to high titers in cell lines approved for vaccine use. Infection in humans is rare and usually asymptomatic, with mild flu-like symptoms. Moreover, due to affinity of VSV envelope glycoprotein to the LDL (low-density lipoprotein) receptor, VSV is effective at targeting a variety of tissues in vivo. A series of research results confirm the possibility of developing VSV-based vaccines against human papilloma viruses (HPV), human immunodeficiency virus (HIV), hepatitis B virus (HBV) and filoviruses (MARV, ZEBOV and SEBOV), as well as the potential use of a successfully developed vaccine against hepatitis C virus (HCV). VSV is neurotropic and infection can cause a viral encephalitis in experimental animals. Therefore, intensive studies are being undertaken to achieve satisfactory expression of the viral antigens while maintaining the safety of the constructed vectors.

In vitro-synthesized infectious RNA as an attenuated live vaccine in a flavivirus model.

Live virus vaccines have in many cases proven to be an extremely effective tool for the prevention of viral diseases. However, the production of conventional live vaccines in eukaryotic cell cultures has many disadvantages, including the potential for contamination with adventitious agents and genetic alterations during propagation, making it necessary to do extensive testing before distribution. Based on results obtained with a flavivirus (tick-borne encephalitis virus) in an experimental animal system, we propose a novel live attenuated virus vaccination strategy consisting of the application of in vitro-synthesized infectious RNA instead of the live virus itself. When administered using the GeneGun, less than 1 ng of RNA was required to initiate replication of virus that was attenuated by a specifically engineered deletion and this induced a protective immunity in laboratory mice. Because this approach uses RNA, it does not have the potential drawbacks of DNA vaccines and thus combines the advantages of conventional live virus vaccines (for example, mimicking natural infection and inducing long-lasting immunity) with those of nucleic acid-based vaccines (for example, ease of production without a requirement for eukaryotic cell culture, stability and purity).

Targeting Fc effector function in vaccine design.

INTRODUCTION: Antibodies mediate pathogen neutralization in addition to several cytotoxic Fc functions through engaging cellular receptors and recruiting effector cells. Fc effector functions have been well described in disease control and protection against infectious diseases including HIV, Ebola, malaria, influenza and tuberculosis, making them attractive targets for vaccine design. AREAS COVERED: We briefly summarize the role of Fc effector functions in disease control and protection in viral, bacterial and parasitic infectious diseases. We review Fc effector function in passive immunization and vaccination, and primarily focus on strategies to elicit and modulate these functions as part of a robust vaccine strategy. EXPERT OPINION: Despite their known correlation with vaccine efficacy for several diseases, only recently have seminal studies addressed how these Fc effector functions can be elicited and modulated in vaccination. However, gaps remain in assay standardization and the precise mechanisms of diverse functional assays. Furthermore, there are inherent difficulties in the translation of findings from animal models to humans, given the difference in sequence, expression and function of Fc receptors and Fc portions of antibodies. However, overall it is clear that vaccine development to elicit Fc effector function is an important goal for optimal prevention against infectious disease.

Prospective vaccination of COVID-19 using shRNA-plasmid-LDH nanoconjugate.

COVID-19 is the pandemic outbreak that is caused by SARS-CoV-2 virus from December, 2019. Human race do not know the curative measure of this devastating disease. In today's era of nanotechnology, it may use its knowledge to develop molecular vaccine to combat this disease. In this article we are intended to propose a hypothesis on the development of a vaccine that is molecular in nature to work against COVID-19. The nanoconjugate may comprise with the inorganic nanoparticle layered double hydroxide intercalated with shRNA-plasmid that have a sequence targeting towards the viral genome or viral mRNA. This nanoconjugate may be used as a nasal spray to deliver the shRNA-plasmid to the target site. The nanoconjugate will have several advantages such as they are biocompatible, they forms as stable knockdown to the target cells and they are stable in the nasal mucosa.

Vaccinomics strategy for developing a unique multi-epitope monovalent vaccine against Marburg marburgvirus.

Marburg virus is known to cause a severe hemorrhagic fever (MHF) in both humans and non-human primates with high degree of infectivity and lethality. To date no approved treatment is available for Marburg virus infection. A study was employed to design a novel chimeric subunit vaccine against Marburg virus by adopting reverse vaccinology approach. The entire viral proteome was retrieved from UniprotKB and assessed to design highly antigenic epitopes by antigenicity screening, transmembrane topology screening, allergenicity and toxicity assessment, population coverage analysis and molecular docking approach. Envelope glycoprotein (GP) and matrix protein (VP40) were identified as most antigenic viral proteins which generated a plethora of epitopes. The final vaccine was constructed by the combination of highly immunogenic epitopes along with suitable adjuvant and linkers. Physicochemical and secondary structure of the designed vaccine was assessed to ensure its thermostability, hydrophilicity, theoretical PI and structural behaviors. Disulfide engineering, molecular dynamic simulation and codon adaptation were further employed to develop a unique multi-epitope monovalent vaccine. Docking analysis of the refined vaccine structure with different MHC molecules and human immune TLR8 receptor present on lymphocyte cells demonstrated higher interaction. Moreover, disulfide engineering served to lessen the high mobility region of the designed vaccine in order to extend its stability. Complexed structure of the modeled vaccine and TLR8 showed minimal deformability at molecular level. Finally, translational potency and microbial expression of the modeled vaccine was analyzed with pET28a(+) vector for E. coli strain K12. However, further in vitro and in vivo investigation could be implemented for the acceptance and validation of the predicted vaccine against Marburg virus.

Virus-epitope vaccine design: informatic matching the HLA-I polymorphism to the virus genome.

Attempts to develop peptide vaccines, based on a limited number of peptides face two problems: HLA polymorphism and the high mutation rate of viral epitopes. We have developed a new genomic method that ensures maximal coverage and thus maximal applicability of the peptide vaccine. The same method also promises a large number of epitopes per HLA to prevent escape via mutations. Our design can be applied swiftly in order to face rapidly emerging viral diseases. We use a genomic scan of all candidate peptides and join them optimally. For a given virus, we use algorithms computing: peptide cleavage probability, transfer through TAP and MHC binding for a large number of HLA alleles. The resulting peptide libraries are pruned for peptides that are not conserved or are too similar to self peptides. We then use a genetic algorithm to produce an optimal protein composed of peptides from this list properly ordered for cleavage. The selected peptides represent an optimal combination to cover all HLA alleles and all viral proteins. We have applied this method to HCV and found that some HCV proteins (mainly envelope proteins) represent much less peptide than expected. A more detailed analysis of the peptide variability shows a balance between the attempts of the immune system to detect less mutating peptides, and the attempts of viruses to mutate peptides and avoid detection by the immune system. In order to show the applicability of our method, we have further used it on HIV-I, Influenza H3N2 and the Avian Flu Viruses.

Expression of HPV-11 L1 protein in transgenic Arabidopsis thaliana and Nicotiana tabacum.

BACKGROUND: We have investigated the possibility and feasibility of producing the HPV-11 L1 major capsid protein in transgenic Arabidopsis thaliana ecotype Columbia and Nicotiana tabacum cv. Xanthi as potential sources for an inexpensive subunit vaccine. RESULTS: Transformation of plants was only achieved with the HPV-11 L1 gene with the C-terminal nuclear localization signal (NLS-) encoding region removed, and not with the full-length gene. The HPV-11 L1 NLS- gene was stably integrated and inherited through several generations of transgenic plants. Plant-derived HPV-11 L1 protein was capable of assembling into virus-like particles (VLPs), although resulting particles displayed a pleomorphic phenotype. Neutralising monoclonal antibodies binding both surface-linear and conformation-specific epitopes bound the A. thaliana-derived particles and - to a lesser degree - the N. tabacum-derived particles, suggesting that plant-derived and insect cell-derived VLPs displayed similar antigenic properties. Yields of up to 12 microg/g of HPV-11 L1 NLS- protein were harvested from transgenic A. thaliana plants, and 2 microg/g from N. tabacum plants - a significant increase over previous efforts. Immunization of New Zealand white rabbits with approximately 50 microg of plant-derived HPV-11 L1 NLS- protein induced an antibody response that predominantly recognized insect cell-produced HPV-11 L1 NLS- and not NLS+ VLPs. Evaluation of the same sera concluded that none of them were able to neutralise pseudovirion in vitro. CONCLUSION: We expressed the wild-type HPV-11 L1 NLS- gene in two different plant species and increased yields of HPV-11 L1 protein by between 500 and 1000-fold compared to previous reports. Inoculation of rabbits with extracts from both plant types resulted in a weak immune response, and antisera neither reacted with native HPV-11 L1 VLPs, nor did they neutralise HPV-11 pseudovirion infectivity. This has important and potentially negative implications for the production of HPV-11 vaccines in plants.

A Bioinformatics approach to designing a Zika virus vaccine.

The Zika virus infections have reached epidemic proportions in the Latin American countries causing severe birth defects and neurological disorders. While several organizations have begun research into design of prophylactic vaccines and therapeutic drugs, computer assisted methods with adequate data resources can be expected to assist in these measures to reduce lead times through bioinformatics approaches. Using 60 sequences of the Zika virus envelope protein available in the GenBank database, our analysis with numerical characterization techniques and several web based bioinformatics servers identified four peptide stretches on the Zika virus envelope protein that are well conserved and surface exposed and are predicted to have reasonable epitope binding efficiency. These peptides can be expected to form the basis for a nascent peptide vaccine which, enhanced by incorporation of suitable adjuvants, can elicit immune response against the Zika virus infections.

Evolutionary implications of Avian Infectious Bronchitis Virus (AIBV) analysis.

For developing efficient vaccines, it is essential to identify which amino acid changes are most important to the survival of the virus. We investigate the amino acid substitution features in the Avian Infectious Bronchitis Virus (AIBV) antigenic domain of a vaccine serotype (DE072) and a virulent viral strain (GA98) to better understand adaptive evolution of AIBV. In addition, the SARS Coronavirus (SARS-CoV) was also analyzed in the same way. It is interesting to find that extreme comparability exists between AIBV and SARS in amino acid substitution pattern. It suggests that amino acid changes that result in overall shift of residue charge and polarity should be paid special attention to during the development of vaccines.

Accumulation of recombinant SARS-CoV spike protein in plant cytosol and chloroplasts indicate potential for development of plant-derived oral vaccines.

Plants are promising candidates as bioreactors for the production of oral recombinant proteins in the biopharmaceutical industry. As an initial step toward provision of an oral vaccine against the severe acute respiratory syndrome coronavirus (SARS-CoV), we have expressed a partial spike (S) protein of SARS-CoV in the cytosol of nuclear-transformed plants and in the chloroplasts of plastid-transformed plants. In the construction of both nuclear and plastid transformation vectors, a 2-kilobase nucleotide sequence encoding amino acids 1-658 of the SARS-CoV spike protein (S1) was modified with nucleotide changes, but not amino acid changes, to optimize codon usage for expression in plants. To investigate the subcellular localization of S1 during transient expression in tobacco leaves, a translational fusion consisting of S1 and the green fluorescent protein (GFP) was generated. Following agroinfiltration of tobacco leaves, analysis by laser confocal scanning microscopy revealed that the S1:GFP fusion protein was localized to the cytosol. In stable transgenic tobacco plants and lettuce plants generated by Agrobacterium-mediated transformation, tobacco and lettuce leaves were observed to express the S1 at high levels from the Cauliflower Mosaic Virus 35S promoter with Northern blot analysis. When the S1 was expressed in transplastomic tobacco, S1 messenger RNA and its corresponding protein were detected on Northern and Western blot analyses, respectively. Our results demonstrate the feasibility of producing S1 in nuclear- and chloroplast-transformed plants, indicating its potential in subsequent development of a plant-derived and safe oral recombinant subunit vaccine against the SARS-CoV in edible plants.

Synthesis, characterization, and immune efficacy of layered double hydroxide@SiO2 nanoparticles with shell-core structure as a delivery carrier for Newcastle disease virus DNA vaccine.

Layered double hydroxide (LDH)@SiO2 nanoparticles were developed as a delivery carrier for the plasmid DNA expressing the Newcastle disease virus F gene. The LDH was hydrotalcite-like materials. The plasmid DNA encapsulated in the LDH@SiO2 nanoparticles (pFDNA-LDH@SiO2-NPs) was prepared by the coprecipitation method, and the properties of pFDNA-LDH@SiO2-NPs were characterized by transmission electron microscopy, zeta potential analyzer, Fourier transform infrared spectroscopy, and X-ray diffraction analysis. The results demonstrated that the pFDNA-LDH@SiO2-NPs had a regular morphology and high stability with a mean diameter of 371.93 nm, loading capacity of 39.66%±0.45%, and a zeta potential of +31.63 mV. A release assay in vitro showed that up to 91.36% of the total plasmid DNA could be sustainably released from the pFDNA-LDH@SiO2-NPs within 288 hours. The LDH@SiO2 nanoparticles had very low toxicity. Additionally, their high transfection efficiency in vitro was detected by fluorescent microscopy. Intranasal immunization of specific pathogen-free chickens with pFDNA-LDH@SiO2-NPs induced stronger cellular, humoral, and mucosal immune responses and achieved a greater sustained release effect than intramuscular naked plasmid DNA, and the protective efficacy after challenge with the strain F48E9 with highly virulent (mean death time of chicken embryos ≤60 hours, intracerebral pathogenicity index in 1 -day-old chickens >1.6) was 100%. Based on the results, LDH@SiO2 nanoparticles can be used as a delivery carrier for mucosal immunity of Newcastle disease DNA vaccine, and have great application potential in the future.

Synthetic RNAs Mimicking Structural Domains in the Foot-and-Mouth Disease Virus Genome Elicit a Broad Innate Immune Response in Porcine Cells Triggered by RIG-I and TLR Activation.

The innate immune system is the first line of defense against viral infections. Exploiting innate responses for antiviral, therapeutic and vaccine adjuvation strategies is being extensively explored. We have previously described, the ability of small in vitro RNA transcripts, mimicking the sequence and structure of different domains in the non-coding regions of the foot-and-mouth disease virus (FMDV) genome (ncRNAs), to trigger a potent and rapid innate immune response. These synthetic non-infectious molecules have proved to have a broad-range antiviral activity and to enhance the immunogenicity of an FMD inactivated vaccine in mice. Here, we have studied the involvement of pattern-recognition receptors (PRRs) in the ncRNA-induced innate response and analyzed the antiviral and cytokine profiles elicited in swine cultured cells, as well as peripheral blood mononuclear cells (PBMCs).

A review of human papillomavirus vaccines: from basic science to clinical trials.

Human papillomavirus (HPV) infection leads to a spectrum of disease from genital warts to precancerous lesions to cervical and anal cancer and is a worldwide public health problem of epidemic proportions. Unique to HPV-related neoplasia, the presence of specific viral antigens such as the L1 capsid structural protein and the oncoproteins E6 and E7 provide opportunities for vaccine therapy. Although difficult to precisely define, the natural immune response to HPV is vitally important and defects in cell mediated immunity correlate with increased risk of disease and cancer. In preclinical animal models, both prophylactic and therapeutic vaccines have effectively induced HPV-specific cell mediated immune responses protecting animals from viral challenge or eliminating established tumors. Most prophylactic vaccines are virus-like particles (VLP) composed of the L1 structural protein. Phase I trials have demonstrated safety and immunogenicity, but limited efficacy data are available. Therapeutic vaccine trials are reviewed including E6 and E7 vaccines comprised of peptides, fusion proteins, encapsulated plasmid DNA, and recombinant vaccinia virus. All of the vaccines appear to be safe, well tolerated, and preliminary data indicates that most are clinically effective. Multiple trials are in progress and more mature data are expected within the next few years.