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In silico study of rotavirus VP7 surface accessible conserved regions for antiviral drug/vaccine design.

ABSTRACT: BACKGROUND: Rotaviral diarrhoea kills about half a million children annually in developing countries and accounts for one third of diarrhea related hospitalizations. Drugs and vaccines against the rotavirus are handicapped, as in all viral diseases, by the rapid mutational changes that take place in the DNA and protein sequences rendering most of these ineffective. As of now only two vaccines are licensed and approved by the WHO (World Health Organization), but display reduced efficiencies in the underdeveloped countries where the disease is more prevalent. We approached this issue by trying to identify regions of surface exposed conserved segments on the surface glycoproteins of the virion, which may then be targeted by specific peptide vaccines. We had developed a bioinformatics protocol for these kinds of problems with reference to the influenza neuraminidase protein, which we have refined and expanded to analyze the rotavirus issue. RESULTS: Our analysis of 433 VP7 (Viral Protein 7 from rotavirus) surface protein sequences across 17 subtypes encompassing mammalian hosts using a 20D Graphical Representation and Numerical Characterization method, identified four possible highly conserved peptide segments. Solvent accessibility prediction servers were used to identify that these are predominantly surface situated. These regions analyzed through selected epitope prediction servers for their epitopic properties towards possible T-cell and B-cell activation showed good results as epitopic candidates (only dry lab confirmation). CONCLUSIONS: The main reasons for the development of alternative vaccine strategies for the rotavirus are the failure of current vaccines and high production costs that inhibit their application in developing countries. We expect that it would be possible to use the protein surface exposed regions identified in our study as targets for peptide vaccines and drug designs for stable immunity against divergent strains of the rotavirus. Though this study is fully dependent on computational prediction algorithms, it provides a platform for wet lab experiments.

SUBMITTER: Ghosh A

PROVIDER: S-EPMC3406019 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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In silico study of rotavirus VP7 surface accessible conserved regions for antiviral drug/vaccine design.

Ghosh Ambarnil A Chattopadhyay Shiladitya S Chawla-Sarkar Mamta M Nandy Papiya P Nandy Ashesh A

PloS one 20120726 7

<h4>Background</h4>Rotaviral diarrhoea kills about half a million children annually in developing countries and accounts for one third of diarrhea related hospitalizations. Drugs and vaccines against the rotavirus are handicapped, as in all viral diseases, by the rapid mutational changes that take place in the DNA and protein sequences rendering most of these ineffective. As of now only two vaccines are licensed and approved by the WHO (World Health Organization), but display reduced efficiencie ...[more]

PMID: 22844409

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Project description:Species A rotaviruses (RVs) are a leading cause of severe acute gastroenteritis in infants and children younger than 5 years. Currently available RV vaccines were adapted from wild-type RV strains by serial passage of cultured cells or by reassortment between human and animal RV strains. These traditional methods require large-scale screening and genotyping to obtain vaccine candidates. Reverse genetics is a tractable, rapid, and reproducible approach to generating recombinant RV vaccine candidates carrying any VP4 and VP7 genes that provide selected antigenicity. Here, we developed a vaccine platform by generating recombinant RVs carrying VP4 (P[4] and P[8]), VP7 (G1, G2, G3, G8, and G9), and/or VP6 genes cloned from human RV clinical samples using the simian RV SA11 strain (G3P[2]) as a backbone. Neutralization assays using monoclonal antibodies and murine antisera revealed that recombinant VP4 and VP7 monoreassortant viruses exhibited altered antigenicity. However, replication of VP4 monoreassortant viruses was severely impaired. Generation of recombinant RVs harboring a chimeric VP4 protein for SA11 and human RV gene components revealed that the VP8* fragment was responsible for efficient infectivity of recombinant RVs. Although this system must be improved because the yield of vaccine viruses directly affects vaccine manufacturing costs, reverse genetics requires less time than traditional methods and enables rapid production of safe and effective vaccine candidates.IMPORTANCE Although vaccines have reduced global RV-associated hospitalization and mortality over the past decade, the multisegmented genome of RVs allows reassortment of VP4 and VP7 genes from different RV species and strains. The evolutionary dynamics of novel RV genotypes and their constellations have led to great genomic and antigenic diversity. The reverse genetics system is a powerful tool for manipulating RV genes, thereby controlling viral antigenicity, growth capacity, and pathogenicity. Here, we generated recombinant simian RVs (strain SA11) carrying heterologous VP4 and VP7 genes cloned from clinical isolates and showed that VP4- or VP7-substituted chimeric viruses can be used for antigenic characterization of RV outer capsid proteins and as improved seed viruses for vaccine production.

Project description:RNA interference (RNAi) is a post-transcriptional gene silencing mechanism that mediates the sequence-specific degradation of targeted RNA and thus provides a tremendous opportunity for development of oligonucleotide-based drugs. Here, we report on the design and validation of small interfering RNAs (siRNAs) targeting highly conserved regions of the hepatitis C virus (HCV) genome. To aim for therapeutic applications by optimizing the RNAi efficacy and reducing potential side effects, we considered different factors such as target RNA variations, thermodynamics and accessibility of the siRNA and target RNA, and off-target effects. This aim was achieved using an in silico design and selection protocol complemented by an automated MysiRNA-Designer pipeline. The protocol included the design and filtration of siRNAs targeting highly conserved and accessible regions within the HCV internal ribosome entry site, and adjacent core sequences of the viral genome with high-ranking efficacy scores. Off-target analysis excluded siRNAs with potential binding to human mRNAs. Under this strict selection process, two siRNAs (HCV353 and HCV258) were selected based on their predicted high specificity and potency. These siRNAs were tested for antiviral efficacy in HCV genotype 1 and 2 replicon cell lines. Both in silico-designed siRNAs efficiently inhibited HCV RNA replication, even at low concentrations and for short exposure times (24h); they also exceeded the antiviral potencies of reference siRNAs targeting HCV. Furthermore, HCV353 and HCV258 siRNAs also inhibited replication of patient-derived HCV genotype 4 isolates in infected Huh-7 cells. Prolonged treatment of HCV replicon cells with HCV353 did not result in the appearance of escape mutant viruses. Taken together, these results reveal the accuracy and strength of our integrated siRNA design and selection protocols. These protocols could be used to design highly potent and specific RNAi-based therapeutic oligonucleotide interventions.

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In silico study of rotavirus VP7 surface accessible conserved regions for antiviral drug/vaccine design.

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In silico study of rotavirus VP7 surface accessible conserved regions for antiviral drug/vaccine design.

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