Pioneering siRNA-Mediated Protection of Mammalian Cells against Zika Virus (MR-766) Infection (original) (raw)
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Scientific Reports
Zika virus (ZIKV) is a mosquito-borne member of the Flaviviridae family that has been known to circulate for decades causing mild febrile illness. The more recent ZIKV outbreaks in the Americas and the Caribbean associated with congenital malformations and Guillain-Barré syndrome in adults have placed public health officials in high alert and highlight the significant impact of ZIKV on human health. New technologies to study the biology of ZIKV and to develop more effective prevention options are highly desired. In this study we demonstrate that direct delivery in mice of an infectious ZIKV cDNA clone allows the rescue of recombinant (r)ZIKV in vivo. A bacterial artificial chromosome containing the sequence of ZIKV strain Paraiba/2015 under the control of the cytomegalovirus promoter was complexed with a commercial transfection reagent and administrated using different routes in type-I interferon receptor deficient A129 mice. Clinical signs and death associated with ZIKV viremia wer...
A review on current status of antiviral siRNA
Reviews in Medical Virology, 2018
Viral diseases like influenza, AIDS, hepatitis, and Ebola cause severe epidemics worldwide. Along with their resistant strains, new pathogenic viruses continue to be discovered so creating an ongoing need for new antiviral treatments. RNA interference is a cellular gene-silencing phenomenon in which sequence-specific degradation of target mRNA is achieved by means of complementary short interfering RNA (siRNA) molecules. Short interfering RNA technology affords a potential tractable strategy to combat viral pathogenesis because siRNAs are specific, easy to design, and can be directed against multiple strains of a virus by targeting their conserved gene regions. In this review, we briefly summarize the current status of siRNA therapy for representative examples from different virus families. In addition, other aspects like their design, delivery, medical significance, bioinformatics resources, and limitations are also discussed.
Microbes and infection, 2018
Vaccines are considered one of the greatest advances in modern medicine. The global burden of numerous infectious diseases has been significantly reduced, and in some cases, effectively eradicated through the deployment of specific vaccines. However, efforts to develop effective new vaccines against infectious pathogens such as influenza, Human immunodeficiency virus (HIV), dengue virus (DENV), chikungunya virus (CHIKV), Ebola virus, and Zika virus (ZIKV) have proven challenging. Zika virus is a mosquito-vectored flavivirus responsible for periodic outbreaks of disease in Africa, Southeast Asia, and the Pacific Islands dating back over 50 years. Over this period, ZIKV infections were subclinical in most infected individuals and resulted in mild cases of fever, arthralgia, and rash in others. Concerns about ZIKV changed over the past two years, however, as outbreaks in Brazil, Central American countries, and Caribbean islands revealed novel aspects of infection including vertical and...
Zika virus is an aedes mosquito borne pathogen belonging to the member of Flaviviridae subgroup causes an emerging disease called Zika fever, known as a benign infection usually presenting as influenza like illness with cutaneous rash. Nowadays epidemic outbreak caused by Zika virus is highly contagious and incurable with present technologies; thus considered as a major health risk which need enhanced surveillance. Genetic studies on Flavivirus have shown that, the 3' untranslated region (UTR) is consists of seven highly conserved stem loop structure and is important for viral replication and pathogenesis. Therefore, 3' UTR of Zika virus can be utilized as suitable target for controlling Zika virus mediated infection. Viral infection can be reduced by RNA interference (RNAi) technology in which double stranded RNA (siRNA and miRNA) molecules mediate the post transcriptional gene silencing (PTGS) of genes in a sequence specific manner. However genetic variability has been determined in different viral isolates; it is a great challenge to design potential siRNA (small interfering RNA) molecules to repress the expression of respective target gene rather than any other viral gene simultaneously. This work is done using various computational tools to design 21 nucleotides long siRNA sequence on the basis of rational siRNA designing method targeting CDS (coding sequence) of 3' UTR of Zika virus. In this study out of one hundred seventy eight computationally identified siRNAs only four most promising siRNA molecules for gene silencing of 3' UTR of Zika virus were verified using other computer aided tools which might lead to suppress the viral activity. Thus, this approach may provide an insight for chemically synthesized RNA molecules as antiviral agent for Zika virus mediated infection and acts as a foundation stone for an efficient therapeutics at genome level.
Viruses
Zika virus (ZIKV) is a type of RNA virus that belongs to the Flaviviridae family. We have reported the construction of a DNA-launched replicon of the Asian-lineage Natal RGN strain and the production of single-round infectious particles (SRIPs) via the combination of prM/E virus-like particles with the replicon. The main objective of the study was to engineer the ZIKV replicon as mammalian expression vectors and evaluate the potential of ZIKV mini-replicon-based SRIPs as delivery vehicles for heterologous gene expression in vitro and in vivo. The mini-replicons contained various genetic elements, including NS4B, an NS5 methyltransferase (MTase) domain, and an NS5 RNA-dependent RNA polymerase (RdRp) domain. Among these mini-replicons, only ZIKV mini-replicons 2 and 3, which contained the full NS5 and NS4B-NS5 genetic elements, respectively, exhibited the expression of reporters (green fluorescent protein (GFP) and cyan fluorescent protein–yellow fluorescent fusion protein (CYP)) and ...
Biochemical and Biophysical Research Communications, 2006
Gene silencing by RNAi and siRNAs has become a well-used tool for researchers. Because of its relatively small size, siRNA was originally thought to avoid activation of anti-viral responses. Recent reports demonstrating so-called ''off-target effects'' are therefore alarming. One issue raised is that siRNA induces interferon-regulated genes at the transcriptional level. We characterize the anti-viral responses of synthetic siRNA and in vitro-transcribed siRNA by measuring the mRNA levels of IFN-b and OAS2 in HeLa cells. Transfections with both traditional and LNA-modified synthetic siRNA cause no anti-viral responses, whereas transfection with either long dsRNA or in vitro-transcribed siRNA leads to greater than 1000-fold induction of these genes. The lack of response was also demonstrated at the level of phosphorylated eIF2a, and measuring of IFN-b by ELISA in cell culture media from the human cell line MCF-7. Altogether, transfection with synthetic siRNA does not induce anti-viral responses in these two cell lines. Our results reinforce the role of siRNA as an effective tool for reverse genetics and strengthen siLNA as a tool for future therapeutic applications.
siRNA could be a potential therapy for COVID-19
EXCLI Journal, 2020
Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded non-coding RNA molecules, which is 20-25 base pairs in length. siRNAs can regulate the expression of genes, by a phenomenon known as RNA interference (RNAi). Based on the phenomenon, the siRNA based therapeutics have been developed and implemented for anticancer, antiviral, and genetic diseases (Liu et al., 2020). In December 2019, WHO reported the outbreak of a novel coronavirus, designated as SARS-CoV-2 or severe acute respiratory syndrome-related coronavirus. This virus has currently spread across 212 countries which resulted in 2,416,135 active cases of infection, and approximately 165,939 mortalities, as per WHO (2020). There are many drugs currently being tested which include antiviral (remdesivir, favipiravir, lopinavir, ritonavir, and arbidol), antimalarial (hydroxychloroquine), and anticancer (interferon-alpha 2b) agents. These drug candidates are undergoing clinical trials, and their efficacy against SARS-CoV-2 has yet to be proven. Under such a situation, siRNA based treatment can provide an effective solution in combating COVID-19 (Liu et al., 2020). Some earlier studies revealed that siRNA candidates were effectively used against the outbreak of SARS and Middle-East Respiratory Syndrome (MERS), recapitulated in Table 1. The siRNAs identified successfully targeted the sequences which coded for the viral RNAdependent RNA polymerase, helicase, proteolytic enzymes, and the nucleoprotein N of earlier SARS virus leading to a 50, 70, 90, and 95 % decrease in viral load, respectively. The viral genome of SARS-CoV-2 is 29 kbp in size and one of the largest genomes among the RNA virus. This genome consists of fourteen open reading frames (ORFs) which coded for twentyseven structural and nonstructural proteins (Wu et al., 2020a). At the 5' end, there are the two largest ORFs, namely ORF1a and ORF1b which are translated into a single large poly-protein by the ribosome through a frame-shift event. The ORF1a comprises of two viral cysteine pro
Human Vaccines & Immunotherapeutics, 2019
Significant concerns have arisen over the past 3 y from the increased global spread of the mosquitoborne flavivirus, Zika. Accompanying this spread has been an increase in cases of the devastating birth defect microcephaly as well as of Guillain-Barré syndrome in adults in many affected countries. Currently there is no vaccine or therapy for this infection; however, we sought to develop a combination approach that provides more rapid and durable protection than traditional vaccination alone. A novel immune-based prophylaxis/therapy strategy entailing the facilitated delivery of a synthetic DNA consensus prME vaccine along with DNA-encoded anti-ZIKV envelope monoclonal antibodies (dMAb) were developed and evaluated for antiviral efficacy. This immediate and persistent protection strategy confers the ability to overcome shortcomings inherent with conventional active vaccination or passive immunotherapy. A collection of novel dMAbs were developed which were potent against ZIKV and could be expressed in serum within 24-48 h of in vivo administration. The DNA vaccine, from a previous development, was potent after adaptive immunity was developed, protecting against infection, brain and testes pathology in relevant mouse challenge models and in an NHP challenge. Delivery of potent dMAbs protected mice from the same murine viral challenge within days of delivery. Combined injection of dMAb and the DNA vaccine afforded rapid and long-lived protection in this challenge model, providing an important demonstration of the advantage of this synergistic approach to pandemic outbreaks.
In Vivo Delivery of a DNA-Encoded Monoclonal Antibody Protects Non-human Primates against Zika Virus
Molecular Therapy, 2019
Zika virus (ZIKV) infection is endemic to several world regions, and many others are at high risk for seasonal outbreaks. Synthetic DNA-encoded monoclonal antibody (DMAb) is an approach that enables in vivo delivery of highly potent mAbs to control infections. We engineered DMAb-ZK190, encoding the mAb ZK190 neutralizing antibody, which targets the ZIKV E protein DIII domain. In vivo-delivered DMAb-ZK190 achieved expression levels persisting >10 weeks in mice and >3 weeks in non-human primate (NHPs), which is protective against ZIKV infectious challenge. This study is the first demonstration of infectious disease control in NHPs following in vivo delivery of a nucleic acid-encoded antibody, supporting the importance of this new platform.
Viral infection resistance conferred on mice by siRNA transgenesis
Transgenic Research, 2013
RNA interference is an attractive strategy to fight against viral diseases by targeting the mRNA of viral genes. Most studies have reported the transient delivery of small interfering RNA or small hairpin Keywords siRNA Á Pseudorabies virus Á Transgene Á Virus challenge Á Aujeszky's desease Á siRNA PCR quantification Nathalie Daniel-Carlier and Ashraf Sawafta contributed equally to this work.