小环HBV cccDNA与Gaussia荧光素酶报告人调查HBV cccDNA生物学和开

StephenW

Sci Rep. 2016 Nov 7;6:36483. doi: 10.1038/srep36483.
Minicircle HBV cccDNA with a Gaussia luciferase reporter for investigating HBV cccDNA biology and developing cccDNA-targeting drugs.Li F1,2, Cheng L1, Murphy CM1, Reszka-Blanco NJ1, Wu Y1, Chi L1, Hu J3, Su L1,4.
Author information

• 1Lineberger Comprehensive Cancer Center, Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599, USA.
• 2Institute of Infectious Diseases, Guangzhou Eighth People's Hospital, Guangzhou Medical University, Guangzhou, Guangdong Province 510060, China.
• 3Department of Microbiology and Immunology, Penn State College of Medicine, Hershey, PA 17033-0850, USA.
• 4Department of Translational Medicine, Department of Surgery, Department of Medicine, the First Hospital, Jilin University, Changchun, Jilin Province 130061, China.
  

AbstractChronic Hepatitis B Virus (HBV) infection is generally not curable with current anti-viral drugs. Virus rebounds after stopping treatment from the stable HBV covalently-closed-circular DNA (cccDNA). The development of drugs that directly target cccDNA is hampered by the lack of robust HBV cccDNA models. We report here a novel HBV cccDNA technology that will meet the need. We engineered a minicircle HBV cccDNA with a Gaussia Luciferase reporter (mcHBV-GLuc cccDNA), which serves as a surrogate to measure cccDNA activity. The mcHBV-GLuc cccDNA was easily produced in bacteria, and it formed minichromosomes as HBV cccDNA episome DNA does when it was transfected into human hepatocytes. Compared to non-HBV minicircle plasmids, mcHBV-GLuc cccDNA showed persistent HBV-GLuc activity and HBx-dependent gene expression. Importantly, the mcHBV-GLuc cccDNA showed resistance to interferons (IFN) treatment, indicating its unique similarity to HBV cccDNA that is usually resistant to long-term IFN treatment in chronic HBV patients. Most importantly, GLuc illuminates cccDNA as a surrogate of cccDNA activity, providing a very sensitive and quick method to detect trace amount of cccDNA. The mcHBV-GLuc cccDNA model is independent of HBV infection, and will be valuable for investigating HBV cccDNA biology and for developing cccDNA-targeting drugs.


PMID:27819342DOI:10.1038/srep36483

StephenW

Sci Rep。2016 Nov 7; 6：36483。 doi：10.1038 / srep36483。
小环HBV cccDNA与Gaussia荧光素酶报告人调查HBV cccDNA生物学和开发cccDNA靶向药物。
Li F1,2，Cheng L1，Murphy CM1，Reszka-Blanco NJ1，Wu Y1，Chi L1，Hu J3，Su L1,4。
作者信息

    1Lineberger Comprehensive Cancer Center，Department of Microbiology and Immunology，University of North Carolina at Chapel Hill，Chapel Hill，North Carolina 27599，USA。
    广州医学院广州市第八人民医院传染病研究所广东广州510060。
    3Department of Microbiology and Immunology，Penn State College of Medicine，Hershey，PA 17033-0850，USA。
    吉林大学第一医院外科，外科，吉林大学第一医院外科，吉林长春130061摘要：

抽象

慢性乙型肝炎病毒（HBV）感染通常不能用目前的抗病毒药物治愈。病毒在停止从稳定的HBV共价闭合环状DNA（cccDNA）处理后反弹。直接靶向cccDNA的药物的开发受到缺乏强大的HBV cccDNA模型的阻碍。我们在这里报告一种新的HBV cccDNA技术，将满足的需要。我们用Gaussia荧光素酶报告基因（mcHBV-GLuc cccDNA）设计了一个小环HBV cccDNA，其作为测量cccDNA活性的替代物。 mcHBV-GLuc cccDNA在细菌中容易产生，并且当其转染到人肝细胞中时，其形成微染色体作为HBV cccDNA附加体DNA。与非HBV小环质粒相比，mcHBV-GLuc cccDNA表现出持续的HBV-GLuc活性和HBx依赖性基因表达。重要的是，mcHBV-GLuc cccDNA显示对干扰素（IFN）治疗的抗性，表明其与HBV cccDNA的独特相似性，其通常对慢性HBV患者中的长期IFN治疗有抗性。最重要的是，GLuc照亮cccDNA作为cccDNA活性的替代，提供了一种非常灵敏和快速的方法来检测痕量的cccDNA。 mcHBV-GLuc cccDNA模型独立于HBV感染，对于研究HBV cccDNA生物学和开发cccDNA靶向药物将是有价值的。

PMID：
    27819342
DOI：
    10.1038 / srep36483