新型苯并噻唑类NS5A抑制剂的设计、合成和抗HCV活性研究及其它研究

发布时间：2018-09-07 15:48

【摘要】：本论文由两个部分组成：第一部分是以丙型肝炎病毒(Hepatitis C Virus, HCV)非结构蛋白NS5A抑制剂GL100953为先导化合物，设计合成了新型苯并噻唑类抗HCV化合物，并对其生物活性进行了测试和评价；第二部分是发现了一种新颖的多取代苯类化合物合成方法，即以噻唑烷二酮或罗丹宁构建的螺环化合物为底物，经氢氧化钾处理一步构建得到多取代苯类化合物。一、新型苯并噻唑类HCV NS5A抑制剂的设计、合成和抗HCV活性研究全球目前约有1.7亿人感染了HCV病毒，且感染发病率还在逐年递增，已经成为一种受到高度重视的健康危害。如得不到有效治疗，HCV感染还会导向发展为肝硬化和肝癌。目前主要使用利巴韦林和聚乙二醇干扰素联合治疗HCV感染，但是该治疗方案能发挥效用的基因型有限，效果不佳，并容易引起耐药。非结构蛋白NS5A是HCV RNA复制必不可少的关键病毒蛋白，基于NS5A研发抗HCV新药是目前的热点领域。本研究课题以HCV NS5A抑制剂GL100953为先导化合物，开展了苯并噻唑类新型抑制剂的设计、合成和抗HCV活性研究。首先，对先导化合物GL100953的苯基噻唑母核进行了骨架跃迁研究，发现了苯并噻唑类化合物GL110509的抗HCV抑制活性优于先导化合物GL100953。其次，对化合物GL110509的酰胺部分进行构效关系研究，设计合成了酰胺类(A01-A13)、胺类(B01-B02)、磺酰胺类(C01-C03)和双磺酰胺类(D01)四类化合物。其中，化合物B02、C03和D01的抗HCV抑制活性最为突出，IC50值分别为1.46mol·L-1、5.76mol·L-1和2.99mol·L-1。在此基础上，对双磺酰胺衍生物D01进行结构优化，发现2-氟衍生物D02具有最佳的抗HCV抑制活性和选择性，其抑制活性达到纳摩尔级(IC50值为486.2nmol·L-1)，并且细胞毒性低(CC5050μmol·L-1)。在2-氟衍生物D02的基础上，进一步优化另一侧脯氨酸上取代基。发现去除苄氧羰基后所得化合物D15抑制活性(IC50=0.4mol·L-1)与D02(IC50=0.49mol·L-1)基本相当，说明苄氧羰基对化合物的抑制活性贡献不大。但在脯氨酸上引入3-苯丙酰基后，，化合物D19的活性比D02提高了约2.5倍，IC50值达到0.19μmol·L-1是所有目标化合物中抗HCV抑制活性最强的。此外，D19的细胞毒性非常低，CC50值大于50μmol·L-1，选择性指数SI大于357，是一个比较有前景的先导化合物，值得进一步结构优化。由于目前还没有NS5A相关分子水平筛选模型的报道，本课题组初步建立了基于表面等离子共振(SPR)的检测方法，可用于评价化合物与NS5A的相互作用。二、五取代苯类化合物的合成新方法研究多取代苯类化合物是常见且应用广泛的功能分子，常用作化工产业中重要的中间体和有机材料，并且普遍存在于药物中。但是现有多取代苯类化合物的合成方法存在诸多缺陷，例如反应条件苛刻，需要使用过渡金属或重金属催化剂参与反应，相对合成成本较高，多采用甲苯、乙腈等有毒溶剂等。本课题组前期基于噻唑烷二酮或罗丹宁构建了螺环化合物。进一步研究发现以氢氧化钾为碱，EtOH/H2O混合液(1:1)为溶剂，可经一步反应合成五取代苯类化合物。该方法具有较高的收率(50.26%~73.26%)，并且具有良好的底物适用范围，使用的氢氧化钾廉价易得，使用的溶剂较为绿色环保。该方法不仅提供了一种全新的、经济的、绿色环保的多取代苯类化合物合成方法；而且新合成的多取代苯类化合物同时具有醛基、酚羟基和酰胺基三种活泼性的反应基团，可进行进一步衍生化，因此也可作为重要的医药和染料化工原料进行开发。综上所述，本研究共设计合成得到64个以苯并噻唑为母核的抗HCV新化合物，并完成了一项多取代苯合成新方法学研究。本论文的创新点在于：(1)设计合成得到了新型苯并噻唑类NS5A抑制剂，发现多个化合物的抗HCV活性达到了nM级。尤其是化合物D19具有高效低毒的特点，为开发抗HCV创新药物提供了高质量的先导结构，为NS5A相关的化学生物学研究提供了新颖的分子探针；(2)发展了一项多取代苯合成新方法，具有反应效率高，绿色环保，易衍生化等优点。
[Abstract]:This paper consists of two parts: the first part is a novel benzothiazole anti-HCV compound designed and synthesized with GL100953, a non-structural protein NS5A inhibitor of hepatitis C virus (HCV), as the lead compound, and its biological activity was tested and evaluated; the second part is the discovery of a novel polysubstituted benzene. Polysubstituted benzenes were synthesized by one-step reaction with thiazolidinedione or rhodanine as substrates.
Design, synthesis and anti HCV activity of a new benzothiazole HCV NS5A inhibitor
There are about 170 million people infected with HCV worldwide, and the incidence of infection is increasing year by year, which has become a highly valued health hazard. Without effective treatment, HCV infection will also lead to cirrhosis and liver cancer. Non-structural protein NS5A is a key viral protein essential for replication of HCV RNA. The development of new anti-HCV drugs based on NS5A is a hot area at present.
In this study, a novel benzothiazole inhibitor GL100953 was designed, synthesized and its anti-HCV activity was studied. Firstly, the skeleton transition of the parent nucleus of benzothiazole GL100953 was studied. It was found that the anti-HCV inhibitory activity of benzothiazole GL110509 was better than that of the precursor GL110509. Compound GL100953. Secondly, the amides of GL110509 were studied by structure-activity relationship. Four compounds were designed and synthesized, including amides (A01-A13), amines (B01-B02), sulfonamides (C01-C03) and disulfonamides (D01). On this basis, the structure of disulfonamide derivative D01 was optimized. It was found that the 2-fluoride derivative D02 had the best anti-HCV inhibitory activity and selectivity. Its inhibitory activity reached nanomolar level (IC50 value 486.2 nmol L-1), and its cytotoxicity was low (CC5050_ 50_ micromol L-1). On the basis of the 2-fluoride derivative D02, the other derivative D02 was further optimized. It was found that the inhibitory activity of D15 (IC50 = 0.4mol L 1) was similar to that of D02 (IC50 = 0.49mol L 1), indicating that benzyloxycarbonyl had little contribution to the inhibitory activity of the compounds. However, when 3-phenylpropionyl was added to proline, the inhibitory activity of D19 was about 2.5 times higher than that of D02, and the IC50 value was 0. In addition, the cytotoxicity of D19 is very low, the CC50 value is greater than 50 micromol. L-1 and the selectivity index SI is greater than 357. It is a promising lead compound and deserves further structural optimization. A method based on surface plasmon resonance (SPR) was developed to evaluate the interaction between compounds and NS5A.
A new method for the synthesis of two, five substituted benzene compounds
Polysubstituted benzenes are common and widely used functional molecules, often used as important intermediates and organic materials in the chemical industry, and are ubiquitous in drugs. However, the existing synthesis methods of polysubstituted benzenes have many drawbacks, such as harsh reaction conditions, need to use transition metals or heavy metal catalysts. The reaction costs higher relative synthesis cost, such as toluene, acetonitrile and other toxic solvents.
The spirocyclic compounds were synthesized from thiazolidinedione or rhodanine in the previous stage. Further studies showed that PENTASUBSTITUTED benzenes could be synthesized by one-step reaction using potassium hydroxide as the base and EtOH/H2O mixture (1:1) as the solvent. The method had high yield (50.26%~73.26%) and good substrate application. This method not only provides a new, economical and environmentally friendly synthesis method of polysubstituted benzenes, but also has three active reaction groups, aldehyde group, phenolic hydroxyl group and amide group, which can be further derived. Therefore, it can also be used as an important pharmaceutical and Dyestuff Chemical raw material.
In summary, 64 new anti-HCV compounds with benzothiazole as the parent nucleus were designed and synthesized, and a new method of synthesizing polysubstituted benzenes was completed. Compound D19 has the characteristics of high efficiency and low toxicity, providing a high-quality lead structure for the development of innovative anti-HCV drugs, and providing a novel molecular probe for NS5A-related chemical and biological research; (2) A new synthetic method of multi-substituted benzene has been developed, which has the advantages of high reaction efficiency, green environmental protection and easy derivation.
【学位授予单位】：第二军医大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R91;R914.5

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