【作者】 杜娟；

【导师】 姚小军；

【作者基本信息】 兰州大学 ， 化学信息学， 2011， 博士

【摘要】 随着抗癌药物伊马替尼(商品名Gleevec)的成功开发,发展蛋白激酶抑制剂(Protein Kinase Inhibitor, PKI)成为药物开发的重要领域。目前已经有10个蛋白激酶小分子抑制剂通过认证并用于临床,仍有200多个激酶抑制剂正在临床实验中,为一些重大疾病的治疗带来希望的曙光。计算机辅助药物(Computer-aided Drug Design, CADD)设计作为药物研发中的重要技术和工具,推动了这一研究领域的快速发展。定量构效关系、分子对接、药效团、同源模建和分子动力学模拟等多种CADD方法均被成功地应用于蛋白激酶抑制剂的设计中。本论文从蛋白激酶及其抑制剂的结构出发,采用多种计算机辅助分子设计方法,包括定量构效关系,分子对接,分子动力学模拟,结合自由能计算等研究多种蛋白激酶抑制剂分子结构与其活性之间的关系,抑制剂与蛋白之间的相互作用,抑制剂的选择性机理,期望通过建立定量模型快速准确预测活性未知化合物并获得影响抑制剂活性的重要结构特征,从而为抑制剂的设计、结构优化提供新的思路,辅助设计和合成更高效的靶向激酶抑制剂。论文的第一章对蛋白激酶分类,结构(活性和非活性构象)及其抑制剂结构,分类及研究进展进行概述,对计算机辅助药物设计中常用的方法,包括定量构效关系、分子对接、分子动力学模拟、结合自由能计算方法等进行介绍,并总结了计算机辅助药物设计方法在蛋白激酶抑制剂的分子设计及相关机理研究中的应用。论文的第二章将多种分子模拟方法应用于细胞周期检验点激酶(Checkpoint kinase, Chk)抑制剂的结合亲和力预测,针对两类骨架结构不同的Chkl抑制剂进行研究。在第一个工作中以一类5,10-dihydro-dibenzo[b,e][1,4]diazepin-11-ones Chkl抑制剂为研究对象,考虑了传统QSAR研究中忽略的重要问题—化合物活性构象的选取,采用精确对接软件Glide将该类抑制剂对接到Chk1的活性位点,从而获得可靠的复合物结构。在此基础上计算了ADME性质相关的描述符、基于配体-受体相互作用的描述符等对小分子结构进行表征。通过遗传算法选择出影响化合物分子抑制活性的关键描述符,同时建立了多元线性回归模型,并对这类抑制剂进行了3D-QSAR研究,建立了CoMFA (Comparative Molecular Field Analysis)模型。所建立的模型均具有较好的稳定性、鲁棒性以及预测能力。在第二个工作中,以喹诺酮类Chkl抑制剂为研究对象,充分考虑了传统对接方法中忽略的因素,比如蛋白的柔性,水分子的影响以及极化效应对结果的影响,采用多种分子对接方法研究该类抑制剂和Chkl的结合模式,在对接复合物的基础上,进行Prime/MM-GBSA计算,预测了这类抑制剂的结合自由能。分子对接和Prime/MM-GBSA联用的方法不但可以快速准确的预测这些化合物的结合自由能,也为Chkl先导的发现和结构优化提供了新思路。论文的第三章将计算机辅助分子设计方法应用于c-Jun氨基末端激酶(JNK)抑制剂的分子设计和相关机理研究。在第一个工作中,以一系列异喹啉酮类JNK1抑制剂为研究对象,首先利用分子对接方法考察了这类抑制剂和JNK1的结合模式,利用GA-MLR和CoMFA方法研究了结构和活性之间关系,并找出了影响活性的关键结构因素,在此基础上设计了新的抑制剂,经预测其活性有所提高。在第二个工作中,从两个高选择性的JNK3抑制剂分子结构入手,采用计算模拟的方法在分子水平上研究它们的选择性机理。首先采用分子对接的方法预测这两个化合物和JNK3, p38准确的结合模式。然后利用分子动力学模拟对复合物结构进行长时间尺度的采样,基于分子动力学模拟的轨迹,用分子力学／广义波恩表面积(Molecular Mechanics Generalized Born Surface Area, MM-GBSA)方法进行结合自由能计算,找出了决定其选择性的关键残基,从而为设计新的高选择性的JNK3抑制剂提供理论指导。论文的第四章从一类血管内皮生长因子受体(VEGFR)酪氨酸激酶抑制剂的结构入手,首先通过分子对接方法来考察这类抑制剂和受体的结合模式,并且确定这些抑制剂的生物活性构象。采用CoMFA和CoMSIA (Comparative Molecular Similarity Index Analysis)分析来找到影响抑制活性的结构特征,并采取了两种不同的叠合策略来考察对模型的影响,基于对接构象的叠合方法得到的模型更加稳定,预测能力更好。基于3D-QSAR所找到的提高活性的关键结构因素,进行了新化合物设计,经预测所设计化合物活性明显提高。论文的第五章,以一类具有选择性的p型甲状腺激素受体(TRβ)激动剂作为研究对象,通过分子对接方法研究激动剂和受体相互作用模式,在此基础上建立这些激动剂的CoMFA和CoMSIA模型,三维等值线图结果表明在A环间位引入体积大、给电子取代基有利于活性提高,B环邻位引入体积大的取代基对活性有利,这些结构信息为进一步的结构修饰提供了理论指导。更多还原

【Abstract】 With the successful application of anticancer drug imatinib (trade name Gleevec), developing protein kinase inhibitor has gained heightened concern in the field of drug development. Ten protein kinase inhibitors have been approved and used for clinicals. There are now more than 200 kinase inhibitors in clinical development, which shed light on the treatment of some major diseases. As the important technology and tool for drug design, computer-aided drug design (CADD) has been applied to PKI discovery. Molecular modeling approaches, such as quantitative structure-activity relationship (QSAR), molecular modeling, pharmacophore, molecular dynamic simulation and binding free energy calculation have been applied to PKI design and discovery.Based on the structure of kinase and inhibitors, this dissertation uses several techniques (QSAR, molecular docking, molecular dynamic simulation and binding free energy calculation) to correlate molecular structure features to their bioactivity, to study the interaction mode between the targeted protein and their inhibitors and to investigate the selectivity mechanism of some inhibitors. We aim at gaining insights into the key structural features affecting activity, and the interaction mechanism for inhibitor-protein binding and selectivity, guiding the design, structural modification and activity prediction of PKI and to aid the design and synthesize of highly effective inhibitors targeted PK.In Chapter 1, we present a general introduction of protein kinase and its inhibitors, including protein kinase classification, structure, kinase inhibitor structure and classification etc. A brief introduction of the computer-aided drug design method was also given, such as QSAR, molecular docking, molecular dynamic simulation and binding free energy calculation. We also describe and overview the recent progress in the study of protein and protein inhibitors.In Chapter 2, multiple molecular modeling methods were applied to predict the inhibitory activity and binding free energy of Chkl inhibitors. In the first work, a series of 5,10-dihydro-dibenzo[b,e][1,4]diazepin-11-ones as inhibitor of Chk1, were taken as the research object, we focus on an important issues overlooked in traditional QSAR study, the selection of active conformation. The accurate molecular docking program—Glide was employed to dock all the studied compounds into the active site of Chk1, and then reliable complex structure was obtained. Structural descriptors, descriptors related to ADME and the ligand-receptor binding free energy were calculated to characterize compounds. Genetic algorithm was used to select the important descriptors influencing on inhibitory activity. At the same time, MLR model was constructed. The CoMFA model was constructed to find some key structural feature affecting activity. Both GA-MLR and CoMFA models are reliable, robust and predictive. In the second work, by considering the important issues including protein flexibility, water molecules and the effect of polarization, multiple docking methods were employed to investigate the binding mode between a series of quinolones and Chk1. Prime/MM-GBSA simulation based on the docked complex is employed to predict the binding free energy of these Chk1 inhibitors. The docking combined with Prime/MM-GBSA simulation can not only be employed to rapidly and accurately predict the binding free energy, but also provide a novel strategy for lead discovery and optimization of Chk1.In Chapter 3, we applied the computer-aided drug design methods to design the novel JNK inhibitors and study the related mechanism. In the first work, based on a set of isoquinolones as inhibitors of c-Jun N-terminal kinase 1, molecular docking was employed to explore the binding mode of this inhibitor at the active site of JNK 1. GA-MLR and CoMFA models were constructed and gained insight into the key structural factors affecting the bioactivity of these inhibitors. According to the results of QSAR study, we designed a series of novel compounds with high JNK1 inhibition activity. In the second work, we studied the selectivity mechanism of two aminopyrazole inhibitors (SR3576 and SR3451) for JNK3 over p38 on molecular level using the combined molecular modeling method. Molecular docking was employed to explore the binding mode of these inhibitors at the active site of JNK3 and p38. A long time molecular dynamic simulation was performed for sampling, MM-GBSA method based on the molecular dynamic simulation trajectories was employed to get insight to the binding and selectivity of them. The key residues responsible for the high selectivity were identified and all these findings can be utilized in the design of new potent selective JNK3 inhibitors.In chapter 4, based on a series of VEGFR-2 (KDR) inhibitors, molecular modeling was performed using 3D-QSAR and docking approach. Docking study was performed to explore the binding mode between all of the compounds and the KDR, which produced the bioactive conformer of the whole data set. CoMFA and CoMSIA, were applied to these inhibitors to gain insights into the key structural factors affecting the bioactivity of these inhibitors. To construct more reasonable 3D-QSAR models, we adopted two different conformer-based alignment methods. The results indicated that the models based on the docked conformer performed better than that based on the co-crystallized conformer. According to the results of QSAR study, we designed a series of novel compounds with high VEGFR kinase inhibition activity.In Chapter 5, a series of agonist of thyroid hormone receptorβwas taken as the research object. Molecular docking was employed to explore the binding mode of the compound at the active site of TRβ.CoMFA and CoMSIA models were constructed based on the docking conformation. The obtained 3D contour maps suggest that the bulky and electropositive substituents at the meta position of the A ring are favorable for the inhibitory activity. The bulky substituents the ortho position of the B ring will enhance the activity. This information is very useful for futher structural modification.更多还原