An Effective Molecular Target Site in Hepatitis B Virus S Gene for Cas9 Cleavage and Mutational Inactivation
نویسندگان
چکیده
Chronic hepatitis B infection remains incurable because HBV cccDNA can persist indefinitely in patients recovering from acute HBV infection. Given the incidence of HBV infection and the shortcomings of current therapeutic options, a novel antiviral strategy is urgently needed. To inactivate HBV replication and destroy the HBV genome, we employed genome editing tool CRISPR/Cas9. Specifically, we found a CRISPR/Cas9 system (gRNA-S4) that effectively targeted the HBsAg region and could suppress efficiently viral replication with minimal off-target effects and impact on cell viability. The mutation mediated by CRISPR/Cas9 in HBV DNA both in a stable HBV-producing cell line and in HBV transgenic mice had been confirmed and evaluated using deep sequencing. In addition, we demonstrated the reduction of HBV replication was caused by the mutation of S4 site through three S4 region-mutated monoclonal cells. Besides, the gRNA-S4 system could also reduce serum surface-antigen levels by 99.91 ± 0.05% and lowered serum HBV DNA level below the negative threshold in the HBV hydrodynamics mouse model. Together, these findings indicate that the S4 region may be an ideal target for the development of innovative therapies against HBV infection using CRISPR/Cas9.
منابع مشابه
CRISPR/Cas9 nickase-mediated disruption of hepatitis B virus open reading frame S and X
Current antiviral therapies cannot cure hepatitis B virus (HBV) infection; successful HBV eradication would require inactivation of the viral genome, which primarily persists in host cells as episomal covalently closed circular DNA (cccDNA) and, to a lesser extent, as chromosomally integrated sequences. However, novel designer enzymes, such as the CRISPR/Cas9 RNA-guided nuclease system, provide...
متن کاملکلونینگ ژن کدکننده آنتیژن سطحی ویروس هپاتیت B در اشرشیاکولی
Background & Aim: Hepatitis B virus(HBV) infection is endemic worldwide. It is estimated every year more than 350 million people become infected with HBV(new cases) worldwide. Unfortunately, there are no satisfactory drugs to cure HBV and related diseases and the only way to control it is through vaccination. Measurements of HBV DNA levels are routinely used to identify infectious chronic c...
متن کاملCRISPR/Cas9 cleavage of viral DNA efficiently suppresses hepatitis B virus
Chronic hepatitis B virus (HBV) infection is prevalent, deadly, and seldom cured due to the persistence of viral episomal DNA (cccDNA) in infected cells. Newly developed genome engineering tools may offer the ability to directly cleave viral DNA, thereby promoting viral clearance. Here, we show that the CRISPR/Cas9 system can specifically target and cleave conserved regions in the HBV genome, r...
متن کاملThe CRISPR/Cas9 System Facilitates Clearance of the Intrahepatic HBV Templates In Vivo
Persistence of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) under current antiviral therapy is a major barrier to eradication of chronic hepatitis B (CHB). Curing CHB will require novel strategies for specific disruption of cccDNA. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is a newly developed tool for site-specific cleavage of DNA tar...
متن کاملHepatitis B Virus Surface Antigen Variants Clustered Within Immune Epitopes in Chronic Hepatitis B Carriers from Hormozgan Province, South of Iran
Objective(s) The aim of this study was to characterize the hepatitis B virus surface protein genotypes and sequence variations among hepatitis B virus surface antigen (HBsAg) positive chronic patients in Hormozgan province, south of Iran. Materials and Methods A total of 8 patients enrolled in this study. The surface gene was amplified and directly sequenced. Genotypes and nucleotide/amino a...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
عنوان ژورنال:
دوره 12 شماره
صفحات -
تاریخ انتشار 2016