Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer
Hepatitis C virus (HCV) infections frequently recur after liver transplantation in patients with HCV-related liver diseases. Approximately 30% of these patients progress to cirrhosis within 5 years after surgery. In this study, we proposed an effective therapeutic strategy to overcome the recurrence...
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| Format: | Article |
| Language: | English |
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Elsevier
2023-12-01
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| Series: | Molecular Therapy: Methods & Clinical Development |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S2329050123001900 |
| _version_ | 1797526193461788672 |
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| author | Tae Hyeong Kim Seong-Wook Lee |
| author_facet | Tae Hyeong Kim Seong-Wook Lee |
| author_sort | Tae Hyeong Kim |
| collection | DOAJ |
| description | Hepatitis C virus (HCV) infections frequently recur after liver transplantation in patients with HCV-related liver diseases. Approximately 30% of these patients progress to cirrhosis within 5 years after surgery. In this study, we proposed an effective therapeutic strategy to overcome the recurrence of HCV. CRISPR-Cas9 was used to insert an expression cassette encoding an RNA aptamer targeting HCV NS5B replicase as an anti-HCV agent into adeno-associated virus integration site 1 (AAVS1), known as a “safe harbor,” in a hepatocellular carcinoma cell line to confer resistance to HCV. The RNA aptamer expression system based on a dihydrofolate reductase minigene was precisely knocked in into AAVS1, leading to the stable expression of aptamer RNA in the developed cell line. HCV replication was effectively inhibited at both the RNA and protein levels in cells transfected with HCV RNA or infected with HCV. RNA immunoprecipitation and competition experiments strongly suggested that this HCV inhibition was due to the RNA aptamer–mediated sequestration of HCV NS5B. No off-target insertion of the RNA aptamer expression construct was observed. The findings suggest that HCV-resistant liver cells produced by genome editing technology could be used as a new alternative in the development of a treatment for HCV-induced liver diseases. |
| first_indexed | 2024-03-10T09:26:42Z |
| format | Article |
| id | doaj.art-83cac53e924c4d3daba501d42e01995a |
| institution | Directory Open Access Journal |
| issn | 2329-0501 |
| language | English |
| last_indexed | 2024-03-10T09:26:42Z |
| publishDate | 2023-12-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Molecular Therapy: Methods & Clinical Development |
| spelling | doaj.art-83cac53e924c4d3daba501d42e01995a2023-11-22T04:47:38ZengElsevierMolecular Therapy: Methods & Clinical Development2329-05012023-12-0131101151Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamerTae Hyeong Kim0Seong-Wook Lee1Department of Molecular Biology, Dankook University, Cheonan 31116, Republic of KoreaDepartment of Bioconvergence Engineering, Research Institute of Advanced Omics, Dankook University, Yongin 16890, Republic of Korea; R&D Center, Rznomics Inc., Seongnam 13486, Republic of Korea; Corresponding author: Seong-Wook Lee, Department of Bioconvergence Engineering, Research Institute of Advanced Omics, Dankook University, Yongin 16890, Republic of Korea.Hepatitis C virus (HCV) infections frequently recur after liver transplantation in patients with HCV-related liver diseases. Approximately 30% of these patients progress to cirrhosis within 5 years after surgery. In this study, we proposed an effective therapeutic strategy to overcome the recurrence of HCV. CRISPR-Cas9 was used to insert an expression cassette encoding an RNA aptamer targeting HCV NS5B replicase as an anti-HCV agent into adeno-associated virus integration site 1 (AAVS1), known as a “safe harbor,” in a hepatocellular carcinoma cell line to confer resistance to HCV. The RNA aptamer expression system based on a dihydrofolate reductase minigene was precisely knocked in into AAVS1, leading to the stable expression of aptamer RNA in the developed cell line. HCV replication was effectively inhibited at both the RNA and protein levels in cells transfected with HCV RNA or infected with HCV. RNA immunoprecipitation and competition experiments strongly suggested that this HCV inhibition was due to the RNA aptamer–mediated sequestration of HCV NS5B. No off-target insertion of the RNA aptamer expression construct was observed. The findings suggest that HCV-resistant liver cells produced by genome editing technology could be used as a new alternative in the development of a treatment for HCV-induced liver diseases.http://www.sciencedirect.com/science/article/pii/S2329050123001900hepatitis C virusRNA aptamerNS5B RNA replicaseCRISPR/Cas9genome editingAAVS1 |
| spellingShingle | Tae Hyeong Kim Seong-Wook Lee Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer Molecular Therapy: Methods & Clinical Development hepatitis C virus RNA aptamer NS5B RNA replicase CRISPR/Cas9 genome editing AAVS1 |
| title | Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer |
| title_full | Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer |
| title_fullStr | Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer |
| title_full_unstemmed | Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer |
| title_short | Generation of hepatitis C virus–resistant liver cells by genome editing–mediated stable expression of RNA aptamer |
| title_sort | generation of hepatitis c virus resistant liver cells by genome editing mediated stable expression of rna aptamer |
| topic | hepatitis C virus RNA aptamer NS5B RNA replicase CRISPR/Cas9 genome editing AAVS1 |
| url | http://www.sciencedirect.com/science/article/pii/S2329050123001900 |
| work_keys_str_mv | AT taehyeongkim generationofhepatitiscvirusresistantlivercellsbygenomeeditingmediatedstableexpressionofrnaaptamer AT seongwooklee generationofhepatitiscvirusresistantlivercellsbygenomeeditingmediatedstableexpressionofrnaaptamer |