Institute of Virology, Technische Universität München/Helmholtz Zentrum München, Munich, Germany.
Gastroenterology. 2011 Aug;141(2):696-706, 706.e1-3. doi: 10.1053/j.gastro.2011.05.001. Epub 2011 May 13.
BACKGROUND & AIMS: Approved therapies for chronic hepatitis B include systemic administration of interferon (IFN)-alfa and inhibitors of hepatitis B virus (HBV) reverse-transcription. Systemic application of IFN-alfa is limited by side effects. Reverse-transcriptase inhibitors effectively control HBV replication, but rarely eliminate the virus and can select drug-resistant variants. We aimed to develop an alternative therapeutic approach that combines gene silencing with induction of IFN in the liver.
To stimulate an immune response while inhibiting HBV activity, we designed 3 small interfering (si)RNAs that target highly conserved sequences and multiple HBV transcripts of all genotypes. A 5'-triphosphate (3p) was added to the siRNAs, turning them into a ligand for the cytosolic helicase retinoic acid-inducible protein I, which becomes activated and induces expression of type-I IFNs. Antiviral activity was investigated in cell lines that replicate HBV, in HBV-infected primary human hepatocytes, and in HBV transgenic mice.
3p-double-stranded RNA (3p-RNA) activated retinoic acid-inducible protein I, induced a strong type I IFN response (expression of IFN-β) in liver cells and showed transient but strong antiviral activity. Bifunctional, HBV-specific, 3p-siRNAs controlled replication of HBV more efficiently and for longer periods of time than 3p-RNAs without silencing capacity or siRNAs that targeted identical sequences but did not contain 3p.
HBV-specific 3p-siRNAs are bifunctional antiviral molecules that induce production of type I IFNs in the liver and target HBV RNAs to inhibit viral replication.
慢性乙型肝炎的批准疗法包括干扰素（IFN）-α的全身给药和乙型肝炎病毒（HBV）逆转录抑制剂。IFN-α的全身应用受到副作用的限制。逆转录酶抑制剂能有效控制 HBV 复制，但很少能消除病毒，并且可能选择耐药变异体。我们旨在开发一种联合基因沉默和肝脏中 IFN 诱导的替代治疗方法。
为了在抑制 HBV 活性的同时刺激免疫反应，我们设计了 3 种靶向高度保守序列和所有基因型的多个 HBV 转录本的小干扰（si）RNA。siRNA 加上 5'-三磷酸（3p），使其成为胞质解旋酶维甲酸诱导蛋白 I 的配体，该蛋白被激活并诱导 I 型 IFN 的表达。在复制 HBV 的细胞系、HBV 感染的原代人肝细胞和 HBV 转基因小鼠中研究了抗病毒活性。
3p-双链 RNA（3p-RNA）激活维甲酸诱导蛋白 I，诱导肝细胞中强烈的 I 型 IFN 反应（IFN-β表达），并表现出短暂但强烈的抗病毒活性。具有双重功能的、HBV 特异性的、3p-siRNA 比没有沉默能力的 3p-RNA 或靶向相同序列但不包含 3p 的 siRNA更有效地控制 HBV 的复制更长时间。
HBV 特异性 3p-siRNA 是具有双重抗病毒功能的分子，可在肝脏中诱导 I 型 IFN 的产生，并靶向 HBV RNA 抑制病毒复制。
