Understanding the Mechanism of the Broad-Spectrum Antiviral Activity of Favipiravir (T-705): Key Role of the F1 Motif of the Viral Polymerase
Understanding the Mechanism of the Broad-Spectrum Antiviral Activity of Favipiravir (T-705): Key Role of the F1 Motif of the Viral Polymerase
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DOI：
10.1128/jvi.00487-17
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发表时间：
2017-06-01
影响因子：
5.4
通讯作者：
Delang, Leen
中科院分区：
文献类型：
--
作者：
Abdelnabi, Rana;de Morais, Ana Theresa Silveira;Delang, Leen
法维拉韦(T-705)是一种广谱抗病毒药物，已在日本获得批准用于治疗流感病毒感染。T-705还可以抑制各种RNA病毒的复制，包括基孔肯雅病毒(CHIKV)。我们早些时候证明了CHIKV依赖RNA聚合酶(RdRp)F1基序的K291R突变是导致对T-705低水平耐药的原因。有趣的是，这种赖氨酸在正义单链RNA(+ssRNA)病毒的RdRp中高度保守。为了深入了解T-705独特的广谱抗病毒活性，我们使用另一种+单链RNA病毒，即柯萨奇病毒B3(CVB3)来探索这种赖氨酸的作用。在CVB3RdRp(K159R)中引入相应的K-to-R替换导致病毒死亡。通过自发获得RdRp中的A239G替换，恢复了K159R变体的复制能力。K159位的突变分析表明，K159M变异体是唯一一个也获得了A239G替换的另一个活的变异体。K159取代显著降低了纯化的病毒RdRp的加工性，通过引入A239G突变恢复了加工性。令人惊讶的是，K159R A239G和K159M A239G变异体被证明比野生型病毒对T-705更敏感，并且在聚合酶检测中表现出较低的保真度。此外，K159R A239G变异体在小鼠中被发现高度减弱。因此，我们证明了+ssRNA病毒RdRp的F1基序中的保守赖氨酸参与了T-705的广谱抗病毒活性，并且它是酶正常发挥作用的关键氨基酸。在本研究中，我们报道了病毒聚合酶高度保守的赖氨酸残基在法韦拉韦(T705)抗阳性单链RNA病毒广谱抗病毒活性中的关键作用。这种保守的赖氨酸的取代对RdRp的功能有重大的负面影响。此外，我们发现这种赖氨酸与RdRp的保真度有关，并且RdRp的保真度影响病毒对T-705抗病毒疗效的敏感性。因此，这些结果为深入了解T-705的抗病毒活性的机制提供了依据，并可能为设计具有比T-705更强的广谱抗病毒活性的新型化学支架奠定基础。
Favipiravir (T-705) is a broad-spectrum antiviral agent that has been approved in Japan for the treatment of influenza virus infections. T-705 also inhibits the replication of various RNA viruses, including chikungunya virus (CHIKV). We demonstrated earlier that the K291R mutation in the F1 motif of the RNA-dependent RNA polymerase (RdRp) of CHIKV is responsible for low-level resistance to T-705. Interestingly, this lysine is highly conserved in the RdRp of positive-sense singlestranded RNA (+ ssRNA) viruses. To obtain insights into the unique broad-spectrum antiviral activity of T-705, we explored the role of this lysine using another +ssRNA virus, namely, coxsackievirus B3 (CVB3). Introduction of the corresponding K-to-R substitution in the CVB3 RdRp (K159R) resulted in a nonviable virus. Replication competence of the K159R variant was restored by spontaneous acquisition of an A239G substitution in the RdRp. A mutagenesis analysis at position K159 identified the K159M variant as the only other viable variant which had also acquired the A239G substitution. The K159 substitutions markedly decreased the processivity of the purified viral RdRp, which was restored by the introduction of the A239G mutation. The K159R A239G and K159M A239G variants proved, surprisingly, more susceptible than the wild-type virus to T-705 and exhibited lower fidelity in polymerase assays. Furthermore, the K159R A239G variant was found to be highly attenuated in mice. We thus demonstrate that the conserved lysine in the F1 motif of the RdRp of +ssRNA viruses is involved in the broad-spectrum antiviral activity of T-705 and that it is a key amino acid for the proper functioning of the enzyme.IMPORTANCE In this study, we report the key role of a highly conserved lysine residue of the viral polymerase in the broad-spectrum antiviral activity of favipiravir (T705) against positive-sense single-stranded RNA viruses. Substitutions of this conserved lysine have a major negative impact on the functionality of the RdRp. Furthermore, we show that this lysine is involved in the fidelity of the RdRp and that the RdRp fidelity influences the sensitivity of the virus for the antiviral efficacy of T-705. Consequently, these results provide insights into the mechanism of the antiviral activity of T-705 and may lay the basis for the design of novel chemical scaffolds that may be endowed with a more potent broad-spectrum antiviral activity than that of T-705.