【作者】 刘涛；

【导师】 万升标；

【作者基本信息】 中国海洋大学 ， 药物化学， 2013， 硕士

【摘要】 目前在肿瘤患者治疗过程中，多药耐药(multidrug resistance, MDR)现象现已成为肿瘤治疗的主要障碍之一。肿瘤细胞MDR的形成机制有多种，其中P糖蛋白（P-gp）、多药耐药相关蛋白(MRP)和乳腺癌耐药蛋白(BCRP)的过表达是重要机制之一。P-糖蛋白(P-glycoprotein, P-gp)的过表达是多药耐药相关机制研究最早最深入的机制之一。寻找高效、低毒、特异性强的MDR逆转剂是近年来抗肿瘤研究的热点。研究发现，天然低毒的多酚类化合物表现出显著的抗肿瘤和逆转多药耐药的活性。本课题组前期研究发现全甲基化EGCG和全甲基化槲皮素衍生物具有较高的MDR逆转活性。为完善黄酮类似物逆转肿瘤多药耐药的构效关系并考察C环立体结构和4位羰基对化合物活性的影响，选取二氢杨梅素（Dihydromyricetin,DMY）作为研究对象。二氢杨梅素是一种多酚双氢黄酮醇，其结构不同于EGCG之处在于二氢杨梅素C环4位含有羰基，而与二氢杨梅素相比槲皮素C环含有双键使得其A环与C环在同一平面。本论文结合EGCG和槲皮素类似物的结构特点与活性，设计并合成了14个甲基化二氢杨梅素结构类似物，并对其进行以P-gp为靶点的肿瘤MDR逆转活性测试。活性实验结果表明，1.0μM浓度的化合物28、29、31能够逆转LCC6MDR细胞对紫杉醇(paclitaxel)的耐药，其逆转倍数为9.5、10.5、16.2，高于阳性对照组维拉帕米(RF=3.3)。β-内酰胺类抗生素是目前临床上应用最为广泛的一类抗生素，但由于抗生素的滥用以及细菌中β-内酰胺酶的出现使得细菌对β-内酰胺类抗生素产生耐药性。新德里金属-β-内酰胺酶（NDM-1）是2009年在一名感染肺炎克雷伯氏菌的瑞士病人身上首次发现的。NDM-1阳性菌株对除了替加环素和粘菌素之外的所有抗生素均有较高的耐药性，NDM-1现在成为全球潜在的健康威胁，因此寻找安全、有效的β-内酰胺酶抑制剂已迫在眉睫。NDM-1的活性位点中共含有两个锌离子，其中第二个锌离子在水解β-内酰胺类抗生素的过程中起着非常重要的作用，因此结合NDM-1活性位点中锌离子以抑制其活性是设计β-内酰胺酶抑制剂的思路之一。近十几年来大量实验发现联苯四氮唑类、碳青霉烯类、巯基化合物和琥珀酸衍生物等化合物对金属内酰胺酶具有一定的抑制作用，化合物主要通过巯基、羧基和咪唑等基团与锌离子结合。本课题组以舒巴坦、他唑巴坦和琥珀酸为基本母核，结合NDM-1水解机制设计了66个化合物及43个水解产物，并将所设计化合物与现有的17个药物分子共计126个化合物建立药物分子数据库，利用计算机辅助药物设计软件sybyl-X2.0中的suflex-dock模块对其进行对接并虚拟筛选。结果显示化合物LT-3-SJ与NDM-1蛋白的对接得分（15.38）最高，并且高于蛋白质自身配体水解氨苄青霉素的对接得分（15.10）。选取化合物LT-3-SJ进行结合模式的研究，为进一步设计与开发新型NDM-1抑制剂提供了一定的思路。更多还原

【Abstract】 n the course of treatment of cancer patients, the phenomenon performance ofmultidrug resistance (multidrug resistance, MDR) has now become the main obstacleto cancer treatment. The mechanism of MDR is complicated, but the over-expressionof P-glycoprotein (P-gp), multidrug resistance associated protein (MRP) and breastcancer resistance protein(BCRP)is one of the important mechanisms. Overexpressionof P-glycoprotein is the earliest and most in-depth study mechanism of multidrugresistance. Searching for efficiency, low toxicity, specificity MDR reversal agents hasbecome one of the hotspots in anticancer research field.Many studies have found that the polyphenol compounds of natural low toxicityshowed significant anti-tumor and reversal activity to multidrug resistance.Preliminary study of our group found that the full methylation EGCG and fullmethylation quercetin derivatives have high MDR reversal activity. Aimed to developthe structure-activity relationship of designed flavonoids reversing multi-drug oftumors and investigate the influence of three-dimensional structure and carbonyl of C-ring, we select dihydromyricetin as the research object. Dihydromyricetin (DMY) isa polyphenol flavonols. Compared with structure of EGCG dihydromyricetin containscarbonyl in the C-4bite of C-ring, but the presence of double bond in C-ring ofquercetin makes the A ring and C ring in the same plane. Combined with thestructural characteristics and activity of EGCG and quercetin analogues, we havedesigned and synthetized14methylation dihydromyricetin analogues and test tumorMDR reversal activity. The activity test results show that the1.0μm compound of28,29,31can reverse resistance to Taxol (paclitaxel) of LCC6MDR cells, with the RF being9.5,10.5,16.2respectively, which is significantly higher activity than thepositive control sample verapamil（RF=3.3）.β-lactam antibiotics are the most widely used class of antibiotics clinically, but theemergence of bacterial β-lactamases and the abuse of antibiotics cause the resistanceof bacterium to Beta-lactam antibiotics. The New Delhi Metallo-β-lactamase (NDM-1)was first reported in2009in a Swedish patient, who travelled to New Delhi andacquired a urinary tract infection caused by Klebsiella Pneumonia. A recent studyreported that NDM-1positive strain was highly resistant to all antibiotics testedexcept tigecycline and colistin, NDM-1now becomes potentially a major globalhealth threat. So searching for the safe and effective β-lactam inhibitors is imminent.The NDM-1active site contains two zinc ions, wherein the second zinc ion playsa very important role in the process of hydrolysis of β-lactam antibiotics. Thereforecombining with zinc ion to inhibit the activity is one of the mechanisms of design ofβ-lactamase inhibitors. A large number of experiments have found that a variety ofcompounds has a certain inhibitory effect to the metal lactamase, such as biphenyltetrazole class, carbapenems, sulfhydryl compounds and derivatives of succinic acid,et al, the mercapto group, carboxyl group and phosphoric acid group of compoundshave strong binding force with zinc ions. Based on hydrolytic mechanisms we design66compounds and43hydrolysis on account of the basic nucleus of sulbactam,tazobactam and succinic acid. Subsequently, we establish a database composing of109compounds and17drug molecules and then use sulflex-dock modules ofcomputer-aided drug design software sybyl-X2.0for docking and virtual screening.Results show that the binding activity of the compound LT-3-SJ is the highest (15.38),which is higher than the binding activity of the ligand hydrolysis of ampicillin (15.10).We study the binding mode of LT-3-SJ and3Q6X, providing certain idea for furtherdesign and development of the new NDM-1inhibitors.更多还原