Cargando…

Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9

The presence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) and the permanent integration of HBV DNA into the host genome confers the risk of viral reactivation and hepatocellular carcinoma. Nucleoside/nucleotide analogs alone have little or no capacity to eliminate replicative H...

Descripción completa

Detalles Bibliográficos
Autores principales: Li, Hao, Sheng, Chunyu, Wang, Shan, Yang, Lang, Liang, Yuan, Huang, Yong, Liu, Hongbo, Li, Peng, Yang, Chaojie, Yang, Xiaoxia, Jia, Leili, Xie, Jing, Wang, Ligui, Hao, Rongzhang, Du, Xinying, Xu, Dongping, Zhou, Jianjun, Li, Mingzhen, Sun, Yansong, Tong, Yigang, Li, Qiao, Qiu, Shaofu, Song, Hongbin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5360708/
https://www.ncbi.nlm.nih.gov/pubmed/28382278
http://dx.doi.org/10.3389/fcimb.2017.00091
_version_ 1782516634718568448
author Li, Hao
Sheng, Chunyu
Wang, Shan
Yang, Lang
Liang, Yuan
Huang, Yong
Liu, Hongbo
Li, Peng
Yang, Chaojie
Yang, Xiaoxia
Jia, Leili
Xie, Jing
Wang, Ligui
Hao, Rongzhang
Du, Xinying
Xu, Dongping
Zhou, Jianjun
Li, Mingzhen
Sun, Yansong
Tong, Yigang
Li, Qiao
Qiu, Shaofu
Song, Hongbin
author_facet Li, Hao
Sheng, Chunyu
Wang, Shan
Yang, Lang
Liang, Yuan
Huang, Yong
Liu, Hongbo
Li, Peng
Yang, Chaojie
Yang, Xiaoxia
Jia, Leili
Xie, Jing
Wang, Ligui
Hao, Rongzhang
Du, Xinying
Xu, Dongping
Zhou, Jianjun
Li, Mingzhen
Sun, Yansong
Tong, Yigang
Li, Qiao
Qiu, Shaofu
Song, Hongbin
author_sort Li, Hao
collection PubMed
description The presence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) and the permanent integration of HBV DNA into the host genome confers the risk of viral reactivation and hepatocellular carcinoma. Nucleoside/nucleotide analogs alone have little or no capacity to eliminate replicative HBV templates consisting of cccDNA or integrated HBV DNA. Recently, CRISPR/Cas9 technology has been widely applied as a promising genome-editing tool, and HBV-specific CRISPR-Cas9 systems were shown to effectively mediate HBV cccDNA disruption. However, the integrated HBV DNA fragments are considered as important pro-oncogenic properties and it serves as an important template for viral replication and expression in stable HBV cell line. In this study, we completely excised a full-length 3,175-bp integrated HBV DNA fragment and disrupted HBV cccDNA in a stable HBV cell line. In HBV-excised cell line, the HBV cccDNA inside cells, supernatant HBV DNA, HBsAg, and HBeAg remained below the negative critical values for more than 10 months. Besides, by whole genome sequencing, we analyzed off-target effects and excluded cell contamination. It is the first time that the HBV infection has been fully eradicated in a stable HBV cell line. These findings demonstrate that the CRISPR-Cas9 system is a potentially powerful tool capable of promoting a radical or “sterile” HBV cure.
format Online
Article
Text
id pubmed-5360708
institution National Center for Biotechnology Information
language English
publishDate 2017
publisher Frontiers Media S.A.
record_format MEDLINE/PubMed
spelling pubmed-53607082017-04-05 Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9 Li, Hao Sheng, Chunyu Wang, Shan Yang, Lang Liang, Yuan Huang, Yong Liu, Hongbo Li, Peng Yang, Chaojie Yang, Xiaoxia Jia, Leili Xie, Jing Wang, Ligui Hao, Rongzhang Du, Xinying Xu, Dongping Zhou, Jianjun Li, Mingzhen Sun, Yansong Tong, Yigang Li, Qiao Qiu, Shaofu Song, Hongbin Front Cell Infect Microbiol Microbiology The presence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) and the permanent integration of HBV DNA into the host genome confers the risk of viral reactivation and hepatocellular carcinoma. Nucleoside/nucleotide analogs alone have little or no capacity to eliminate replicative HBV templates consisting of cccDNA or integrated HBV DNA. Recently, CRISPR/Cas9 technology has been widely applied as a promising genome-editing tool, and HBV-specific CRISPR-Cas9 systems were shown to effectively mediate HBV cccDNA disruption. However, the integrated HBV DNA fragments are considered as important pro-oncogenic properties and it serves as an important template for viral replication and expression in stable HBV cell line. In this study, we completely excised a full-length 3,175-bp integrated HBV DNA fragment and disrupted HBV cccDNA in a stable HBV cell line. In HBV-excised cell line, the HBV cccDNA inside cells, supernatant HBV DNA, HBsAg, and HBeAg remained below the negative critical values for more than 10 months. Besides, by whole genome sequencing, we analyzed off-target effects and excluded cell contamination. It is the first time that the HBV infection has been fully eradicated in a stable HBV cell line. These findings demonstrate that the CRISPR-Cas9 system is a potentially powerful tool capable of promoting a radical or “sterile” HBV cure. Frontiers Media S.A. 2017-03-22 /pmc/articles/PMC5360708/ /pubmed/28382278 http://dx.doi.org/10.3389/fcimb.2017.00091 Text en Copyright © 2017 Li, Sheng, Wang, Yang, Liang, Huang, Liu, Li, Yang, Yang, Jia, Xie, Wang, Hao, Du, Xu, Zhou, Li, Sun, Tong, Li, Qiu and Song. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Microbiology
Li, Hao
Sheng, Chunyu
Wang, Shan
Yang, Lang
Liang, Yuan
Huang, Yong
Liu, Hongbo
Li, Peng
Yang, Chaojie
Yang, Xiaoxia
Jia, Leili
Xie, Jing
Wang, Ligui
Hao, Rongzhang
Du, Xinying
Xu, Dongping
Zhou, Jianjun
Li, Mingzhen
Sun, Yansong
Tong, Yigang
Li, Qiao
Qiu, Shaofu
Song, Hongbin
Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9
title Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9
title_full Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9
title_fullStr Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9
title_full_unstemmed Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9
title_short Removal of Integrated Hepatitis B Virus DNA Using CRISPR-Cas9
title_sort removal of integrated hepatitis b virus dna using crispr-cas9
topic Microbiology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5360708/
https://www.ncbi.nlm.nih.gov/pubmed/28382278
http://dx.doi.org/10.3389/fcimb.2017.00091
work_keys_str_mv AT lihao removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT shengchunyu removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT wangshan removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT yanglang removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT liangyuan removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT huangyong removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT liuhongbo removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT lipeng removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT yangchaojie removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT yangxiaoxia removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT jialeili removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT xiejing removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT wangligui removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT haorongzhang removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT duxinying removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT xudongping removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT zhoujianjun removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT limingzhen removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT sunyansong removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT tongyigang removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT liqiao removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT qiushaofu removalofintegratedhepatitisbvirusdnausingcrisprcas9
AT songhongbin removalofintegratedhepatitisbvirusdnausingcrisprcas9