Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus
RNase H-dependent gapmer antisense oligonucleotides (ASOs) are a promising therapeutic approach via sequence-specific binding to and degrading target RNAs. However, the efficacy and mechanism of antiviral gapmer ASOs have remained unclear. Here, we investigated the inhibitory effects of gapmer ASOs...
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MDPI AG
2023-10-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/19/14846 |
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| author | Shunsuke Okamoto Yusuke Echigoya Ayaka Tago Takao Segawa Yukita Sato Takuya Itou |
| author_facet | Shunsuke Okamoto Yusuke Echigoya Ayaka Tago Takao Segawa Yukita Sato Takuya Itou |
| author_sort | Shunsuke Okamoto |
| collection | DOAJ |
| description | RNase H-dependent gapmer antisense oligonucleotides (ASOs) are a promising therapeutic approach via sequence-specific binding to and degrading target RNAs. However, the efficacy and mechanism of antiviral gapmer ASOs have remained unclear. Here, we investigated the inhibitory effects of gapmer ASOs containing locked nucleic acids (LNA gapmers) on proliferating a mosquito-borne flavivirus, Japanese encephalitis virus (JEV), with high mortality. We designed several LNA gapmers targeting the 3′ untranslated region of JEV genomic RNAs. In vitro screening by plaque assay using Vero cells revealed that LNA gapmers targeting a stem-loop region effectively inhibit JEV proliferation. Cell-based and RNA cleavage assays using mismatched LNA gapmers exhibited an underlying mechanism where the inhibition of viral production results from JEV RNA degradation by LNA gapmers in a sequence- and modification-dependent manner. Encouragingly, LNA gapmers potently inhibited the proliferation of five JEV strains of predominant genotypes I and III in human neuroblastoma cells without apparent cytotoxicity. Database searching showed a low possibility of off-target binding of our LNA gapmers to human RNAs. The target viral RNA sequence conservation observed here highlighted their broad-spectrum antiviral potential against different JEV genotypes/strains. This work will facilitate the development of an antiviral LNA gapmer therapy for JEV and other flavivirus infections. |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T21:42:58Z |
| publishDate | 2023-10-01 |
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| series | International Journal of Molecular Sciences |
| spelling | doaj.art-a841da77908d4b93a2cc028e41a10c142023-11-19T14:31:32ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672023-10-0124191484610.3390/ijms241914846Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis VirusShunsuke Okamoto0Yusuke Echigoya1Ayaka Tago2Takao Segawa3Yukita Sato4Takuya Itou5Laboratory of Preventive Veterinary Medicine and Animal Health, Department of Veterinary Medicine, College of Bioresource Sciences, Nihon University, Fujisawa, Kanagawa 252-0880, JapanNihon University Veterinary Research Center, Fujisawa, Kanagawa 252-0880, JapanNihon University Veterinary Research Center, Fujisawa, Kanagawa 252-0880, JapanLaboratory of Preventive Veterinary Medicine and Animal Health, Department of Veterinary Medicine, College of Bioresource Sciences, Nihon University, Fujisawa, Kanagawa 252-0880, JapanNihon University Veterinary Research Center, Fujisawa, Kanagawa 252-0880, JapanLaboratory of Preventive Veterinary Medicine and Animal Health, Department of Veterinary Medicine, College of Bioresource Sciences, Nihon University, Fujisawa, Kanagawa 252-0880, JapanRNase H-dependent gapmer antisense oligonucleotides (ASOs) are a promising therapeutic approach via sequence-specific binding to and degrading target RNAs. However, the efficacy and mechanism of antiviral gapmer ASOs have remained unclear. Here, we investigated the inhibitory effects of gapmer ASOs containing locked nucleic acids (LNA gapmers) on proliferating a mosquito-borne flavivirus, Japanese encephalitis virus (JEV), with high mortality. We designed several LNA gapmers targeting the 3′ untranslated region of JEV genomic RNAs. In vitro screening by plaque assay using Vero cells revealed that LNA gapmers targeting a stem-loop region effectively inhibit JEV proliferation. Cell-based and RNA cleavage assays using mismatched LNA gapmers exhibited an underlying mechanism where the inhibition of viral production results from JEV RNA degradation by LNA gapmers in a sequence- and modification-dependent manner. Encouragingly, LNA gapmers potently inhibited the proliferation of five JEV strains of predominant genotypes I and III in human neuroblastoma cells without apparent cytotoxicity. Database searching showed a low possibility of off-target binding of our LNA gapmers to human RNAs. The target viral RNA sequence conservation observed here highlighted their broad-spectrum antiviral potential against different JEV genotypes/strains. This work will facilitate the development of an antiviral LNA gapmer therapy for JEV and other flavivirus infections.https://www.mdpi.com/1422-0067/24/19/14846Japanese encephalitis virusgapmer antisense oligonucleotidelocked nucleic acid (LNA)antiviralRNase H3′ UTR stem-loop region |
| spellingShingle | Shunsuke Okamoto Yusuke Echigoya Ayaka Tago Takao Segawa Yukita Sato Takuya Itou Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus International Journal of Molecular Sciences Japanese encephalitis virus gapmer antisense oligonucleotide locked nucleic acid (LNA) antiviral RNase H 3′ UTR stem-loop region |
| title | Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus |
| title_full | Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus |
| title_fullStr | Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus |
| title_full_unstemmed | Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus |
| title_short | Antiviral Efficacy of RNase H-Dependent Gapmer Antisense Oligonucleotides against Japanese Encephalitis Virus |
| title_sort | antiviral efficacy of rnase h dependent gapmer antisense oligonucleotides against japanese encephalitis virus |
| topic | Japanese encephalitis virus gapmer antisense oligonucleotide locked nucleic acid (LNA) antiviral RNase H 3′ UTR stem-loop region |
| url | https://www.mdpi.com/1422-0067/24/19/14846 |
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