Hepatitis B virus (HBV) infection is a major public health problem. The high levels of HBV DNA and HBsAg are positively associated with the development of secondary liver diseases, including hepatocellular carcinoma (HCC). Current treatment with nucleos(t)ide analogues mainly reduces viral DNA, but has minimal, if any, inhibitory effect on the viral antigen. Although IFN reduces both HBV DNA and HBsAg, the serious associated side effects limit its use in clinic. Thus, there is an urgent demanding for novel anti‐HBV therapy. In our study, viral parameters were determined in the supernatant of HepG2.2.15 cells, HBV‐expressing Huh7 and HepG2 cells which transfected with HBV plasmids and in the serum of HBV mouse models with hydrodynamic injection of pAAV‐HBV1.2 plasmid. RT‐qPCR and Southern blot were performed to detect 35kb mRNA and cccDNA. RT‐qPCR, Luciferase assay and Western blot were used to determine anti‐HBV effects of MLN4924 and the underlying mechanisms. We found that treatment with MLN4924, the first‐in‐class neddylation inhibitor currently in several phase II clinical trials for anti‐cancer application, effectively suppressed production of HBV DNA, HBsAg, 3.5kb HBV RNA as well as cccDNA. Mechanistically, MLN4924 blocks cullin neddylation and activates ERK to suppress the expression of several transcription factors required for HBV replication, including HNF1α, C/EBPα and HNF4α, leading to an effective blockage in the production of cccDNA and HBV antigen. Our study revealed that neddylation inhibitor MLN4924 has impressive anti‐HBV activity by inhibiting HBV replication, thus providing sound rationale for future MLN4924 clinical trial as a novel anti‐HBV therapy.

中文翻译：

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Neddylation 抑制剂 MLN4924 通过调节 ERK-HNF1α-C/EBPα-HNF4α 轴具有抗 HBV 活性

乙型肝炎病毒（HBV）感染是一个重大的公共卫生问题。高水平的 HBV DNA 和 HBsAg 与继发性肝病的发展呈正相关，包括肝细胞癌 (HCC)。目前使用核苷（酸）类似物的治疗主要减少病毒 DNA，但对病毒抗原的抑制作用很小（如果有的话）。尽管 IFN 可降低 HBV DNA 和 HBsAg，但严重的相关副作用限制了其在临床中的应用。因此，迫切需要新的抗HBV治疗方法。在我们的研究中，测定了转染 HBV 质粒的 HepG2.2.15 细胞、表达 HBV 的 Huh7 和 HepG2 细胞的上清液以及流体动力学注射 pAAV-HBV1.2 质粒的 HBV 小鼠模型的血清中的病毒参数。进行 RT-qPCR 和 Southern blot 以检测 35kb mRNA 和 cccDNA。RT-qPCR, 荧光素酶测定和蛋白质印迹用于确定 MLN4924 的抗 HBV 作用及其潜在机制。我们发现，用 MLN4924（目前在几个用于抗癌应用的 II 期临床试验中的一流的 neddylation 抑制剂）治疗有效地抑制了 HBV DNA、HBsAg、3.5kb HBV RNA 以及 cccDNA 的产生。从机制上讲，MLN4924 阻断 cullin neddylation 并激活 ERK 以抑制 HBV 复制所需的几种转录因子的表达，包括 HNF1α、C/EBPα 和 HNF4α，从而有效阻断 cccDNA 和 HBV 抗原的产生。我们的研究表明，neddylation 抑制剂 MLN4924 通过抑制 HBV 复制具有令人印象深刻的抗 HBV 活性，从而为未来 MLN4924 作为一种新型抗 HBV 疗法的临床试验提供了合理的依据。