Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration
ABSTRACTThe persistence of HBV covalently closed circular DNA (cccDNA) and HBV integration into the host genome in infected hepatocytes pose significant challenges to the cure of chronic HBV infection. Although CRISPR/Cas9-mediated genome editing shows promise for targeted clearance of viral genomes...
| Main Authors: | , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Taylor & Francis Group
2024-12-01
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| Series: | Emerging Microbes and Infections |
| Subjects: | |
| Online Access: | https://www.tandfonline.com/doi/10.1080/22221751.2023.2284286 |
| _version_ | 1797265991620624384 |
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| author | Wanjia Zeng Liwei Zheng Yukun Li Jing Yang Tianhao Mao Jing Zhang Yanna Liu Jing Ning Ting Zhang Hongxin Huang Xiangmei Chen Fengmin Lu |
| author_facet | Wanjia Zeng Liwei Zheng Yukun Li Jing Yang Tianhao Mao Jing Zhang Yanna Liu Jing Ning Ting Zhang Hongxin Huang Xiangmei Chen Fengmin Lu |
| author_sort | Wanjia Zeng |
| collection | DOAJ |
| description | ABSTRACTThe persistence of HBV covalently closed circular DNA (cccDNA) and HBV integration into the host genome in infected hepatocytes pose significant challenges to the cure of chronic HBV infection. Although CRISPR/Cas9-mediated genome editing shows promise for targeted clearance of viral genomes, a safe and efficient delivery method is currently lacking. Here, we developed a novel approach by combining light-induced heterodimerization and protein acylation to enhance the loading efficiency of Cas9 protein into extracellular vesicles (EVs). Moreover, vesicular stomatitis virus-glycoprotein (VSV-G) was incorporated onto the EVs membrane, significantly facilitating the endosomal escape of Cas9 protein and increasing its gene editing activity in recipient cells. Our results demonstrated that engineered EVs containing Cas9/gRNA and VSV-G can effectively reduce viral antigens and cccDNA levels in the HBV-replicating and infected cell models. Notably, we also confirmed the antiviral activity and high safety of the engineered EVs in the HBV-replicating mouse model generated by hydrodynamic injection and the HBV transgenic mouse model. In conclusion, engineered EVs could successfully mediate functional CRISPR/Cas9 delivery both in vitro and in vivo, leading to the clearance of episomal cccDNA and integrated viral DNA fragments, and providing a novel therapeutic approach for curing chronic HBV infection. |
| first_indexed | 2024-03-08T18:26:29Z |
| format | Article |
| id | doaj.art-269676e66cbe405295e73f086e098e4f |
| institution | Directory Open Access Journal |
| issn | 2222-1751 |
| language | English |
| last_indexed | 2024-04-25T00:53:35Z |
| publishDate | 2024-12-01 |
| publisher | Taylor & Francis Group |
| record_format | Article |
| series | Emerging Microbes and Infections |
| spelling | doaj.art-269676e66cbe405295e73f086e098e4f2024-03-11T11:56:50ZengTaylor & Francis GroupEmerging Microbes and Infections2222-17512024-12-0113110.1080/22221751.2023.2284286Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integrationWanjia Zeng0Liwei Zheng1Yukun Li2Jing Yang3Tianhao Mao4Jing Zhang5Yanna Liu6Jing Ning7Ting Zhang8Hongxin Huang9Xiangmei Chen10Fengmin Lu11Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaSchool of Medicine, Shihezi University, Shihezi, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaDepartment of Gastroenterology and Hepatology, Beijing Youan Hospital, Capital Medical University, Beijing, People’s Republic of ChinaDepartment of Gastroenterology, Beijing Key Laboratory for Helicobacter Pylori Infection and Upper Gastrointestinal Diseases, Peking University Third Hospital, Beijing, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaDepartment of Pathogen Biology and Biosecurity, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaDepartment of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University Health Science Center, Beijing, People’s Republic of ChinaABSTRACTThe persistence of HBV covalently closed circular DNA (cccDNA) and HBV integration into the host genome in infected hepatocytes pose significant challenges to the cure of chronic HBV infection. Although CRISPR/Cas9-mediated genome editing shows promise for targeted clearance of viral genomes, a safe and efficient delivery method is currently lacking. Here, we developed a novel approach by combining light-induced heterodimerization and protein acylation to enhance the loading efficiency of Cas9 protein into extracellular vesicles (EVs). Moreover, vesicular stomatitis virus-glycoprotein (VSV-G) was incorporated onto the EVs membrane, significantly facilitating the endosomal escape of Cas9 protein and increasing its gene editing activity in recipient cells. Our results demonstrated that engineered EVs containing Cas9/gRNA and VSV-G can effectively reduce viral antigens and cccDNA levels in the HBV-replicating and infected cell models. Notably, we also confirmed the antiviral activity and high safety of the engineered EVs in the HBV-replicating mouse model generated by hydrodynamic injection and the HBV transgenic mouse model. In conclusion, engineered EVs could successfully mediate functional CRISPR/Cas9 delivery both in vitro and in vivo, leading to the clearance of episomal cccDNA and integrated viral DNA fragments, and providing a novel therapeutic approach for curing chronic HBV infection.https://www.tandfonline.com/doi/10.1080/22221751.2023.2284286Extracellular vesiclesCRISPR/Cas9gene editingantiviral therapyhepatitis B virus |
| spellingShingle | Wanjia Zeng Liwei Zheng Yukun Li Jing Yang Tianhao Mao Jing Zhang Yanna Liu Jing Ning Ting Zhang Hongxin Huang Xiangmei Chen Fengmin Lu Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration Emerging Microbes and Infections Extracellular vesicles CRISPR/Cas9 gene editing antiviral therapy hepatitis B virus |
| title | Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration |
| title_full | Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration |
| title_fullStr | Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration |
| title_full_unstemmed | Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration |
| title_short | Engineered extracellular vesicles for delivering functional Cas9/gRNA to eliminate hepatitis B virus cccDNA and integration |
| title_sort | engineered extracellular vesicles for delivering functional cas9 grna to eliminate hepatitis b virus cccdna and integration |
| topic | Extracellular vesicles CRISPR/Cas9 gene editing antiviral therapy hepatitis B virus |
| url | https://www.tandfonline.com/doi/10.1080/22221751.2023.2284286 |
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