靶向抗乙型肝炎病毒相关性肝细胞癌NTCP抑制剂的设计、合成与构效关系研究

Hepatitis B virus (HBV) associated hepatocellular carcinoma (HCC) is a serious threat to human health. Sodium taurocholate cotransporting polypeptide (NTCP) transporter is a functional receptor for HBV infection in human live cells and plays an important role in the development of HBV associated HCC. Our previous research indicated that a NTCP-targeted inhibitor PMC-0 can effectively inhibits the growth of xenografts in BALB/c nude mice and HBV infection, suggesting that the lead compound PMC-0 shows a potential potency against HCC. In this research project, we plan to design and synthesize a series of new PMC-0 derivatives based on computer aided drug design (CADD) methods such as molecular docking, drug-like analysis and molecular dynamics simulation. The experimental methods including anti-HBV infection and anti-HCC activity were used to evaluate the activity of compounds and elucidate their structure-activity relationship. Furthermore, in order to explore the biological relationship between NTCP and HBV-associated HCC, we constructed the network of NTCP and HCC related proteins using bioinformatics methods, then the protein content of NTCP and HCC was analyzed by proteomic analysis. All these can provide a basis for the development of targeted innovative drugs for the treatment of HBV-associated HCC.

乙型肝炎病毒（HBV）相关性肝细胞癌（HCC）严重威胁着人类的健康。HBV感染人肝细胞的功能性受体--钠离子-牛磺胆酸共转运蛋白（NTCP）在HBV相关性HCC发展过程中起着重要作用。前期我们发现一个具有较高NTCP抑制活性的先导化合物PMC-0，该化合物能有效抑制HBV侵染和BALB/c裸鼠异种移植瘤的生长，提示其具有抗HCC的有效潜质。本研究拟在此基础上，利用分子对接、分子动力学模拟等药物设计方法，将前期得到的PMC-0进行基于结构的药物设计、类药性分析及合成修饰。采用分子药理学试验评价化合物的酶活、HBV侵染及抗HCC活性，阐明其构效关系。进一步采用生物信息学方法构建NTCP与HCC相关性蛋白网络，结合蛋白质组学分析给药前后蛋白的含量变化，揭示NTCP与HBV相关性HCC之间的生物学关系，为治疗HBV相关性HCC的靶向创新药物的研发提供基础。

我国肝癌（HCC）的病死率高居第二。乙型肝炎病毒（HBV）是HCC的危险因素，它的受体钠离子-牛磺胆酸共转运蛋白（NTCP）是HBV相关性HCC的潜在靶标。运用多种药物设计方法获得先导化合物，在此基础上设计合成了32个化合物，并结合生物活性测试初步探讨了抑制剂的构效关系，化合物B7对HBV具有一定的抑制作用，对正常肝细胞无明显毒性，对HepG2细胞的抑制作用（IC50为7.360 ± 0.3 μM）。CETSA实验显示化合物B7靶向NTCP，用Hoechst 33258染色法在化合物B7处理后的HepG2细胞中观察到具有明亮荧光的浓缩染色质，即化合物B7处理后HepG2细胞凋亡显著增加。此外，Annexin-V/PI双重染色也证实化合物B7诱导了剂量依赖性凋亡。另外，评估JNK调节的经典线粒体凋亡途径，发现化合物B7明显诱导JNK、p-JNK、caspase-3、Bax的上调和Bcl-2的下调。同时，化合物B7可以降低NTCP的表达。这些结果表明化合物B7通过诱导细胞凋亡对HepG2细胞具有抗增殖作用。