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Gene Therapy (2015) 22, 404–412; doi:10.1038/gt.2015.2; published online 5 February 2015
Harnessing the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated Cas9 system to disrupt the hepatitis B virus
S Zhen1,2,3,5, L Hua1,4,5, Y-H Liu1, L-C Gao1, J Fu1, D-Y Wan1, L-H Dong1, H-F Song1 and X Gao1
1Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing, China
2Xijing Hospital, Fourth Military Medical University, Xi’an, China
3Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto, Japan
4Laboratory of Pharmacology and Toxicology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan, China
Correspondence: Professor H-F Song or Professor X Gao, Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, 27 Taiping Road, Beijing 100850, China. E-mail: [email protected] or [email protected]
5These authors contributed equally to this work.
Received 11 July 2014; Revised 2 December 2014; Accepted 8 December 2014
Advance online publication 5 February 2015
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Abstract
The current therapies to treat hepatitis B virus (HBV) infection are limited. Recently, clustered regularly interspaced short palindromic repeat (CRISPR) systems, originally identified in bacteria and archaea, have been found to consist of an RNA-based adaptive immune system that degrades complimentary sequences of invading plasmids and viruses. Here, we studied the effects of the CRISPR/CRISPR-associated Cas9 system that was targeted to the surface antigen (HBsAg)-encoding region of HBV, both in a cell culture system and in vivo. The HBsAg levels in the media of the cells and in the sera of mice were analyzed by a quantitative enzyme-linked immunosorbent assay. The HBV DNA levels were assessed by quantitative PCR and HBsAg expression in mouse livers was assessed by an immunohistochemical assay. The amount of HBsAg secreted in the cell culture and mouse serum was reduced by CRISPR/Cas9 treatment. Immunohistochemistry analyses showed almost no HBsAg-positive cells in the liver tissue of CRISPR/Cas9-S1+X3-treated mice. The CRISPR/Cas9 system efficiently produced mutations in HBV DNA. Thus, CRISPR/Cas9 inhibits HBV replication and expression in vitro and in vivo and may constitute a new therapeutic strategy for HBV infection.
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发表于 2015-5-7 20:31
