核糖核苷酸还原酶在乙型肝炎相关肝癌发病中的作用和机制及其作为药物靶标的意义

Hepatitis B virus (HBV) infections are a major cause of viral hepatitis and a key factor for hepatocellular carcinoma (HCC) worldwide. However, the current approved drugs against HBV infections and HCC are far from being satisfactory. Novel treatments with novel targets and different modes of action are still pressingly demanded. Ribonucleotide reductase (RR), supplying dNDPs, is the rate-limiting enzyme for DNA synthesis in cells. Research reports and our previous studies demonstrate that HBV, which has no RR genes, can activate the expression and activity of RR in the host hepatocytes for the viral DNA replication, and dysregulated RR is involved in malignant transformation and cancer development. We determined the X-ray crystal structures of RRM2 subunit protein and established in vitro and in vivo RR activity assays. Based on the 3D structure of RRM2 and computer-assisted design together with RR activity assay screenings, we obtained several novel potent RR inhibitors. We found that the novel RR inhibitors not only had potent anti-HBV activities in HBV-harbouring HepG2.2.15 cells and HBV-transgenic mice, but also efficiently inhibit the proliferation of HCC cells. Furthermore, the compounds have synergistic effects against the replication of HBV with the clinical anti-HBV drug Lamivudine. The above evidence suggests that dysregulated RR may play a key role in the development of HBV-related HCC, and the enzyme could serve as an important target for developing a new category of anti-HBV-related HCC drugs. This proposed study will focus on two aspects. (1) To confirm the role and related mechanism of RR in promoting HBV replication and HCC proliferation using cellular and animal models, and validate the correlation between RR dysregulation and HBV-related HCC development using clinical specimens from HBV-positive and negative HCC patients; (2) To confirm that our novel potent RR inhibitors can inhibit HBV replication and HCC proliferation using our established in vitro and in vivo experiments, and validate the molecular mechanism of action of the compounds. Our goal is to clarify whether dysregulated RR contributes to HBV-related HCC development and whether RR inhibitors have double functions in the inhibition of HBV replication and HCC proliferation. The finding will provide a basis for developing novel drugs against HBV-related HCC with a different mechanism of action from existing therapeutics, as well as help to establish a new strategy for the prevention and treatment of the diseases in the future.

乙型肝炎病毒（HBV）引起乙型肝炎、是肝细胞癌（HCC）主要病因。核糖核苷酸还原酶（RR）提供DNA合成原料脱氧核糖核苷酸（dNDPs）、是细胞DNA合成限速酶。有关报道和我们研究表明：HBV（无RR基因）激活宿主肝细胞表达RR以获得病毒DNA复制所需原料；RR表达和酶活性增高与肿瘤密切相关；我们研发的新RR酶抑制剂能阻止HBV复制和肝癌细胞增殖。因此我们推测：RR是乙型肝炎相关肝癌的关键发病因子和重要药物靶标。本项目拟进一步：通过HBV和HCC细胞和动物模型以及临床乙型肝炎相关肝癌病例研究，阐明RR与HBV和肝癌关系；并应用我们研发的新RR酶抑制剂，在细胞模型以及HBV转基因小鼠和裸鼠移植瘤模型研究，证明抑制RR酶是否具有抗HBV和肝癌双重作用。研究结果将明确RR促进乙型肝炎相关肝癌发生发展作用和机制及其作为药物靶标意义，为研发RR靶向的乙型肝炎相关肝癌创新药物和防治新策略提供科学依据。

乙型肝炎病毒（HBV）引起乙型肝炎、是肝细胞癌（HCC）主要病因。核糖核苷酸还原酶（RR）提供DNA合成原料脱氧核糖核苷酸（dNDPs）、是细胞DNA合成限速酶。HBV本身无RR基因、可能通过激活宿主肝细胞表达RR以获得病毒DNA复制所需原料。RR在HCC细胞和组织中高表达。本项目（1）通过细胞和小鼠模型分析以及临床病例验证表明，HBV-X蛋白促进RR小亚基RRM2转录和RR酶活性，而RR促进HBV基因组DNA和cccDNA复制；肝癌细胞中RRM2表达和RR酶活性显著增加，促进肝癌细胞增殖。因此明确了RR是乙型肝炎及其相关肝癌发生发展的关键因子之一。（2）我们确定了RRM2蛋白X线衍射晶体结构，基于RRM2三维结构结合计算机辅助分析，设计和合成了系列活性小分子化合物。通过体内外细胞和小鼠模型实验分析表明，筛选到的老药和具有新颖结构的RR酶抑制剂具有抗HBV复制和抗肝癌增殖双重作用，并与现有临床抗HBV药物和抗肝癌药物具有协同作用。从而明确了RR可以作为乙型肝炎相关肝癌的药物靶标。综上，本研究结果为明确RR促进乙型肝炎相关肝癌发生发展作用和机制、以及研发RR靶向的乙型肝炎相关肝癌创新药物和防治新策略提供了重要科学依据。.已在相关研究领域主流学术SCI期刊发表标注论文10篇（其中第一标注通讯作者5篇），授权及公示发明专利2项（各1项）），PDB注册蛋白质X线衍射晶体结构4个。此外，还有后续数据正在整理成稿和申请专利之中。