Multiple shRNAs driven by U6 and CMV promoter enhances efficiency of antiviral effects against foot-and-mouth disease virus

Kim, Su-Mi; Lee, Kwang-Nyeong; Lee, Su Jung; Ko, Young-Joon; Lee, Hyang-Sim; Kweon, Chang-Hee; Kim, Hyunsoo; Park, Jong Hyeon

doi:10.1016/j.antiviral.2010.06.004

Cited by 36 publications

(34 citation statements)

References 34 publications

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“…We previously developed two recombinant adenoviruses, namely, Ad-porcine IFN-␣␥, which simultaneously expresses type I and type II interferons, and Ad-3siRNA, which simultaneously expresses 3 siRNAs, to maximize the efficacy of interferon and siRNA (21,22).…”

Section: Discussionmentioning

confidence: 99%

“…All infections using FMDV were conducted in a biosafety level 3 containment facility at the Animal and Plant Quarantine Agency (QIA). The preparation of recombinant adenoviruses expressing 3 different siRNA sequences targeting mRNA encoding the nonstructural proteins 2B and 3C (Ad-3siRNA) or porcine IFN-␣ and IFN-␥ proteins (Ad-porcine IFN-␣␥) has been described previously (21,22). Amplified adenoviruses were purified using a ViraBind adenovirus purification kit (Cell Biolabs, San Diego, CA, USA) or a Vivapure AdenoPack 500 kit (Sartorius, Goettingen, Germany).…”

Section: Cells Viruses and Virus Titrationsmentioning

confidence: 99%

“…Viral RNA was extracted using a MagNA Pure 96 system (Roche, Basel, Switzerland). Real-time RT-PCR was conducted as previously described (21).…”

Section: Cells Viruses and Virus Titrationsmentioning

confidence: 99%

“…To assay the effects on FMDV RNA replication, RNA extraction and quantitative real-time RT-PCR were performed. Viral RNA was extracted using the MagNA Pure 96 system, and real-time RT-PCR was performed as previously described (21).…”

Section: Cells Viruses and Virus Titrationsmentioning

confidence: 99%

“…Viruses can eventually develop resistance mechanisms through point mutations to evade targeted small interfering RNAs (siRNAs), although siRNAs can offer rapid antiviral treatment (19,20). We have developed recombinant adenoviruses that simultaneously express porcine alpha interferon (IFN-␣) and IFN-␥ (referred to here as Ad-porcine IFN-␣␥) and multiple (n ϭ 3) siRNAs targeting nonstructural proteins of FMDV (referred to here as Ad-3siRNA) to enhance the inhibitory effects of these antiviral agents observed in previous studies (21,22). Importantly, both classes of antiviral agents operate through distinct mechanisms and are effective against a broad range of FMDV serotypes.…”

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confidence: 99%

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Robust Protection against Highly Virulent Foot-and-Mouth Disease Virus in Swine by Combination Treatment with Recombinant Adenoviruses Expressing Porcine Alpha and Gamma Interferons and Multiple Small Interfering RNAs

Kim

Park

Lee

et al. 2015

J Virol

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Because the currently available vaccines against foot-and-mouth disease (FMD) provide no protection until 4 to 7 days postvaccination, the only alternative method to halt the spread of the FMD virus (FMDV) during outbreaks is the application of antiviral agents. Combination treatment strategies have been used to enhance the efficacy of antiviral agents, and such strategies may be advantageous in overcoming viral mechanisms of resistance to antiviral treatments. We have developed recombinant adenoviruses (Ads) for the simultaneous expression of porcine alpha and gamma interferons (Ad-porcine IFN-␣␥) as well as 3 small interfering RNAs (Ad-3siRNA) targeting FMDV mRNAs encoding nonstructural proteins. The antiviral effects of Ad-porcine IFN-␣␥ and Ad-3siRNA expression were tested in combination in porcine cells, suckling mice, and swine. We observed enhanced antiviral effects in porcine cells and mice as well as robust protection against the highly pathogenic strain O/Andong/ SKR/2010 and increased expression of cytokines in swine following combination treatment. In addition, we showed that combination treatment was effective against all serotypes of FMDV. Therefore, we suggest that the combined treatment with Adporcine IFN-␣␥ and Ad-3siRNA may offer fast-acting antiviral protection and be used with a vaccine during the period that the vaccine does not provide protection against FMD. IMPORTANCEThe use of current foot-and-mouth disease (FMD) vaccines to induce rapid protection provides limited effectiveness because the protection does not become effective until a minimum of 4 days after vaccination. Therefore, during outbreaks antiviral agents remain the only available treatment to confer rapid protection and reduce the spread of foot-and-mouth disease virus (FMDV) in livestock until vaccine-induced protective immunity can become effective. Interferons (IFNs) and small interfering RNAs (siRNAs) have been reported to be effective antiviral agents against FMDV, although the virus has associated mechanisms of resistance to type I interferons and siRNAs. We have developed recombinant adenoviruses for the simultaneous expression of porcine alpha and gamma interferons (Ad-porcine IFN-␣␥) as well as 3 small interfering RNAs (Ad-3siRNA) to enhance the inhibitory effects of these antiviral agents observed in previous studies. Here, we show enhanced antiviral effects against FMDV by combination treatment with Ad-porcine IFN-␣␥ and Ad-3siRNA to overcome the mechanisms of resistance of FMDV in swine.

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Section: Discussionmentioning

confidence: 99%

Section: Cells Viruses and Virus Titrationsmentioning

confidence: 99%

“…Viral RNA was extracted using a MagNA Pure 96 system (Roche, Basel, Switzerland). Real-time RT-PCR was conducted as previously described (21).…”

Section: Cells Viruses and Virus Titrationsmentioning

confidence: 99%

Section: Cells Viruses and Virus Titrationsmentioning

confidence: 99%

mentioning

confidence: 99%

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Robust Protection against Highly Virulent Foot-and-Mouth Disease Virus in Swine by Combination Treatment with Recombinant Adenoviruses Expressing Porcine Alpha and Gamma Interferons and Multiple Small Interfering RNAs

Kim

Park

Lee

et al. 2015

J Virol

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Inhibition of porcine transmissible gastroenteritis virus infection in porcine kidney cells using short hairpin RNAs targeting the membrane gene

et al. 2016

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The membrane (M) protein is the most abundant component of the porcine transmissible gastroenteritis virus (TGEV) particle. To exploit the possibility of using RNA interference (RNAi) as a strategy against TGEV infection, three plasmids (pRNAT-1, pRNAT-2, and pRNAT-3) expressing short hairpin RNAs were designed to target three different coding regions of the M gene of TGEV. The plasmids were constructed and transiently transfected into a porcine kidney cells, PK-15, to determine whether these constructs inhibited TGEV production. The analysis of cytopathic effects demonstrated that pRNAT-2 and pRNAT-3 could protect PK-15 cells against pathological changes specifically and efficiently. Additionally, indirect immunofluorescence and 50% tissue culture infectious dose (TCID) assays showed that pRNAT-2 and pRNAT-3 inhibited the multiplication of the virus at the protein level effectively. Quantitative real-time PCR further confirmed that the amounts of viral RNAs in cell cultures pre-transfected with the three plasmids were reduced by 13, 68, and 70%, respectively. This is the first report showing that RNAi targeting of the M gene. Our results could promote studies of the specific function of viral genes associated with TGEV infection and might provide a theoretical basis for potential therapeutic applications.

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Down-regulation of viral replication by lentiviral-mediated expression of short-hairpin RNAs against vesicular stomatitis virus ribonuclear complex genes

Ramírez-Carvajal

Long

2012

Antiviral Research

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Multiple shRNAs driven by U6 and CMV promoter enhances efficiency of antiviral effects against foot-and-mouth disease virus

Cited by 36 publications

References 34 publications

Robust Protection against Highly Virulent Foot-and-Mouth Disease Virus in Swine by Combination Treatment with Recombinant Adenoviruses Expressing Porcine Alpha and Gamma Interferons and Multiple Small Interfering RNAs

Robust Protection against Highly Virulent Foot-and-Mouth Disease Virus in Swine by Combination Treatment with Recombinant Adenoviruses Expressing Porcine Alpha and Gamma Interferons and Multiple Small Interfering RNAs

Inhibition of porcine transmissible gastroenteritis virus infection in porcine kidney cells using short hairpin RNAs targeting the membrane gene

Down-regulation of viral replication by lentiviral-mediated expression of short-hairpin RNAs against vesicular stomatitis virus ribonuclear complex genes

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