Although several nucleo(s)tide analogs are available for treatment of HBV infection, long‐term treatment with these drugs can lead to the emergence of drug‐resistant viruses. Recent HIV‐1 studies suggest that combination therapies using nucleo(s)tide reverse transcriptase inhibitors (NRTIs) and non‐nucleo(s)tide reverse transcriptase inhibitors (NNRTIs) could drastically inhibit the viral genome replication of NRTI‐resistant viruses. In order to carry out such combinational therapy against HBV, several new NRTIs and NNRTIs should be developed. Here, we aimed to identify novel NNRTIs targeting the HBV polymerase terminal protein (TP)‐reverse transcriptase (RT) (TP‐RT) domain, which is a critical domain for HBV replication. We expressed and purified the HBV TP‐RT with high purity using an Escherichia coli expression system and established an in vitro ε RNA‐binding assay system. Then, we used TP‐RT in cell‐free assays to screen candidate inhibitors from a chemical compound library, and identified two compounds, 6‐hydroxy‐DL‐DOPA and N‐oleoyldopamine, which inhibited the binding of ε RNA with the HBV polymerase. Furthermore, these drugs reduced HBV DNA levels in cell‐based assays as well by inhibiting packaging of pregenome RNA into capsids. The novel screening system developed herein should open a new pathway the discovery of drugs targeting the HBV TP‐RT domain to treat HBV infection.

中文翻译：

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建立用于寻找与HBV聚合酶结合的εRNA抑制剂的系统。

尽管有几种核苷酸类似物可用于治疗HBV感染，但是用这些药物进行长期治疗可导致耐药性病毒的出现。近期的HIV-1研究表明，使用核苷酸逆转录酶抑制剂（NRTIs）和非核苷酸逆转录酶抑制剂（NNRTIs）的联合疗法可以显着抑制NRTI抗性病毒的病毒基因组复制。为了进行这种针对HBV的联合治疗，应开发几种新的NRTI和NNRTI。在这里，我们旨在鉴定针对HBV聚合酶末端蛋白（TP）-逆转录酶（RT）（TP-RT）域的新型NNRTI，这是HBV复制的关键域。我们使用大肠杆菌表达并纯化了高纯度的HBV TP‐RT表达系统并建立了体外εRNA结合测定系统。然后，我们在无细胞分析中使用TP‐RT从化合物库中筛选候选抑制剂，并鉴定出两种化合物，它们可抑制εRNA与HBV聚合酶的结合，即6-羟基-DL-DOPA和N-油基多巴胺。 。此外，这些药物还通过抑制前基因组RNA包装入衣壳，降低了基于细胞的测定中的HBV DNA水平。本文开发的新型筛查系统应为发现针对HBV TP-RT域的药物治疗HBV感染开辟一条新途径。