新型尿苷肽类化合物Sansanmycin P的提取分离、结构鉴定及活性研究

目的从放线菌 Streptomyces sp SS 发酵液中分离新型活性尿苷肽类似物。方法发酵液经 D4006大孔吸附树脂、DEAE-葡聚糖凝胶及制备液相分离纯化,获得单一化合物;根据 UV、MS、MS/MS、1D 和2D NMR 确定化合物结构,并进行抗菌活性研究。结果分离纯化得到1个 N-末端含有1-甲基-6-羟基四氢异喹啉的尿苷肽类化合物 Sansanmycin P。初步体外抗菌活性,抗铜绿假单胞菌 MIC 〉32μg/ml,抗结核杆菌 MIC 〉32μg/ml。结论 Sansanmycin P 为全新结构 Sansanmycins 类似物,具有一定抗结核杆菌和铜绿假单胞菌活性。     

核糖体蛋白L12-L10相互作用抑制剂的筛选及其抗结核活性的研究

目的以L12-L10相互作用为靶点筛选具有抗结核活性的先导化合物。方法应用酵母双杂交模型AH109(pAD-L12+pBD-L10)通过生长抑制方法筛选阳性化合物,以AH109(pAD-T+pBD-53)作为对照;通过96孔板法检测阳性化合物对耻垢分枝杆菌的抑制活性;应用定量微孔板快速显色法(MABA)检测抗结核杆菌活性;通过β-半乳糖苷酶活性定量检测判断阳性化合物在模型上对L12-L10相互作用的阻断活性;应用体外蛋白表达系统检测阳性化合物对蛋白表达的抑制作用;应用平皿二倍稀释法检测阳性化合物的药敏作用。结果筛选到4个在模型上具有活性的阳性化合物,其对耻垢分枝杆菌具有比较好的抑制活性,其最小抑制浓度(MIC)在3.125~12.5μg/ml之间;其中IBM-T275对结核分枝杆菌标准株和临床分离株均具有比较好的抑制活性,MIC在5~10μg/ml之间,而对细菌抑制活性较低,其MIC均在64μg/ml以上;IBM-T275能够抑制酵母模型内β-半乳糖苷酶的表达,并且能够体外抑制蛋白表达,其IC50为12.57μg/ml。结论筛选到1个具有抗结核杆菌活性的阳性化合物,其抗结核活性可能与阻断L12-L10蛋白相互作用相关。     

以次黄嘌呤单核苷酸脱氢酶为靶点的新型抗结核药物高通量筛选模型的建立及应用

目的 建立以结核分枝杆菌次黄嘌呤单核苷酸脱氢酶为靶点的新型抗结核药物高通量筛选模型.方法 以结核分枝杆菌H37Rv基因组为模板,pBEV表达质粒为载体,将guaB2基因克隆至pBEV以构建pBEV::guaB2重组表达质粒,表达并纯化重组的结核分枝杆菌次黄嘌呤单核苷酸脱氢酶;建立以测定反应体系340 nm吸光值变化速率...     

以丙氨酸消旋酶为靶点的高通量抗结核药物筛选模型的建立及应用

目的建立以结核分枝杆菌丙氨酸消旋酶为靶点的新型高通量抗结核药物筛选模型,筛选丙氨酸消旋酶的抑制剂,获得以丙氨酸消旋酶为靶点的新型抗结核药物先导物。方法以结核分枝杆菌H37Rv基因组为模板,pET28a表达质粒为载体,将alr基因克隆至pET28a,构建pET28a::alr重组表达质粒,表达并纯化得到重组结核分枝杆菌丙氨酸消旋酶;通过测定反应产物NADH在340 nm处光密度变化速率,检测酶反应活性,构建并优化该酶抑制剂的高通量筛选模型;应用该模型对化合物库进行筛选;测定活性化合物IC50以及对结核分枝杆菌的MIC。结果成功构建了结核分枝杆菌alr基因的表达载体;得到了纯度较高的重组丙氨酸消旋酶,测得该酶的比活力为13.53 kU/mg;所建立的丙氨酸消旋酶高通量筛选模型稳定性高,符合高通量筛选的要求;通过对70 000个化合物进行筛选,得到了5个活性较高的化合物,其中,IMB-XZ5对结核分枝杆菌的MIC为4~8μg/ml,且对结核分枝杆菌的作用具有较高的特异性。结论建立了稳定性好、灵敏度较高的结核分枝杆菌丙氨酸消旋酶抑制剂高通量筛选模型,应用该模型筛选得到了具有较好抗结核活性的丙氨酸消旋酶抑制剂。     

基于TNF功能表位设计新型人源单链抗体

目的开发基于TNF功能表位的新型人源单链抗体。方法利用自主研制的鼠抗TNF-α中和单抗Z12能特异性识别TNF-α的141~146位功能表位特性,通过理论模拟构建TNF/抗体Z12相互作用的复合物模型设计获得功能性拮抗肽(PT2、PT3、PT4、PT7)以及单域抗体PTVH5(以人抗体可变区重链框架VH5为支架合理展示PT2、PT3、PT4),利用计算机辅助分子设计以及同源模建、分子对接方法,进一步合理选择人抗体可变区轻链框架(Vκ1)作为展示支架,通过构象判别、作用能比较以及识别区域确认并选择合适的连接肽设计新型单链分子ScFv_AB1。结果理论分析发现,ScFv_AB1稳定性较好,识别TNF-α的141~146功能位点(即Z12识别的位点)。生物学实验证明,ScFv_AB1能与TNF-α结合、抑制TNF-α与TNFR的结合、抑制TNF-α介导的细胞毒作用。结论初步验证了"借助计算机模建,基于人抗体可变区的框架结构和拮抗肽设计单链抗体分子"的策略是可行的,从而为人源小分子抗体的制备提供了一条可供选择的途径。     

胆酸二聚体化合物的设计、合成及抑制肝细胞凋亡研究

目的通过设计合成胆酸二聚体化合物及评价其体外抑制肝细胞凋亡活性,发现新型治疗慢性肝病的候选化合物。方法以熊去氧胆酸、鹅去氧胆酸和奥贝胆酸为起始原料,经亲电取代、脱水缩合、氢化等反应制备目标化合物。通过测定半胱氨酸天冬氨酸蛋白酶3、7的活性水平评价目标化合物体外抑制肝细胞凋亡的活性。结果合成胆酸二聚体化合物24个,体外活性结果表明部分目标化合物具有较好的抑制肝细胞凋亡活性,12个化合物(1a、1e、1h、2a、2c、2e、2f、2g、3a、3c、3d、5a)在20μmol/L浓度下抑制肝细胞凋亡的活性优于阳性对照物牛磺熊去氧胆酸(40.94%)。化合物1a、1e、2a和5a的抑制率分别为75.60%、88.56%、83.25%和105.24%,是阳性对照的2倍。结论熊去氧胆酸和鹅去氧胆酸二聚体具有抑制肝细胞凋亡的活性,奥贝胆酸二聚体无抑制肝细胞凋亡的活性;连接胆酸骨架的连接链长度对活性有明显的影响,其中具有较短连接链(N,N'-二甲基乙二胺、四甲基乙二胺、乙二醇二甲醚)的化合物(1a、1e、2a)的活性优于具有较长连接链(N,N'-二甲基-1,8-辛二胺、2,5,8,11,14,17-六氧十八烷)的化合物(1g、3e)的活性。     

双环醇预防抗结核药物肝功能损害的临床疗效及对T淋巴细胞亚群的影响

目的:探讨双环醇预防抗结核药物所致肝功能损害的临床疗效及对T淋巴细胞亚群影响,观察药物性肝损害的发生率及研究其是否具有免疫调节作用.方法:采用前瞻性研究,对66例初治菌阳肺结核患者随机分为治疗组和对照组,治疗组强化抗痨加双环醇片,对照组强化抗痨加肝泰乐,疗程6个月.并用用流式细胞术(FCM)测定外周血中的T淋巴细胞亚群水平.结果:治疗组的患者肝损害发生率5.6%,而对照组发生率16.7%,两组比较差异显著(P<0.01);两组在治疗前及治疗后2个月、6个月比较,其外周血CD4+、CD8+及CD4+/CD8+也存在明显的差异(P<0.01).治疗组双环醇使用6个月后与对照组比较,其CD4+明显升高,CD8+降低,两组比较差异有显著性(P<0.05).结论:双环醇能有效预防抗结核药物所致肝功能损害,安全性好,无明显副作用,有利于患者顺利完成抗结核疗程,其亦有助于CD4+细胞水平增加及CD8+细胞水平降低.     

抑制RORγt活性及Th17分化的新型化合物研究

目的我们在实验中偶然发现了一种能抑制RORγt活性的新型化合物,拟通过鉴定该化合物对Th17体外分化及其细胞因子分泌的影响对其进行生物学评价,以确定该化合物是否可作为一种新型的自身免疫病候选药物。方法构建能在Jurkat细胞中稳定表达Gal4-RORγt的荧光素酶报告系统,通过该体系发现了一种对RORγt有显著抑制活性的新型化合物,并在小鼠Th17细胞分化实验中对该化合物的作用效果进行体外验证。在此基础上,对该化合物进行生物功能分析,包括EC50、CC50的测定以及T细胞特异性分析。结果通过体外验证实验,我们确定该化合物(编号为compound 2)能显著的抑制Th17分化及其细胞因子IL-17A、IL-17F的表达和分泌。在后续的生物学评价中,我们还发现该化合物对RORγt有较高的抑制效率,较低的细胞毒性和较强的T细胞特异性。结论本研究中得到的化合物compound 2可作为一种潜在的新型前导化合物应用于治疗自身免疫性疾病和一些炎症性疾病。     

基于有限元分析的3D打印脊柱侧弯矫形器局部优化设计

目的 对计算机辅助设计的3D打印脊柱侧弯矫形器进行优化设计,从而使其实现具有良好力学强度和模型轻量化等优点。方法 利用手持式三维扫描仪对志愿者躯干体廓进行扫描,构建志愿者体表模型;根据三点力原理、牵引及免荷原理,对志愿者体表模型进行修型,设计出压力区和释放区,初步设计出脊柱侧弯矫形器模型;采用正交试验方法 对32种不同尺寸的镂空组合进行局部优化对比研究,根据优化结果 ,对脊柱侧弯矫形器模型进行镂空处理和生物力学分析,对比镂空设计的3D打印脊柱侧弯矫形器的应力分布,验证矫形器模型的优化效果。结果 采用半径为9 mm的圆孔和间距23 mm的局部镂空优化设计（局部减重约40%）,可得到质量更轻、透气性更好和足够强度的3D打印脊柱侧弯矫形器。结论 基于有限元生物力学分析,采用非压力区域的局部圆孔镂空优化设计,可以实现3D打印脊柱侧弯矫形器的打印材料减少、透气性增加等优点,最终可提高患者的穿戴舒适度和依从性。     

新型N1-取代-1,8-萘啶-3-甲酰胺类ASBT抑制剂的设计、合成及活性研究

目的设计、合成新型N1-取代-7-N,N-二甲氨基-4-氧代-1,4-二氢-1,8-萘啶-3-N-(3,5-二氟苯基)甲酰胺类化合物,测定其对顶端钠依赖性胆酸转运体(ASBT)的抑制活性,考察N1位取代基对化合物活性的影响。方法以2,6-二氯烟酰乙酸乙酯为起始原料,经缩合、亲和取代、环合、水解等9步反应制备目标化合物,其结构均经质谱和核磁氢谱分析确证。以S-1647为阳性对照,利用放射性结合试验(RBA)法测定目标化合物的ASBT抑制活性。结果合成了N1-取代-7-N,N-二甲氨基-4-氧代-1,4-二氢-1,8-萘啶-3-N-(3,5-二氟苯基)甲酰胺类化合物48个,活性结果表明目标化合物均具有较好的ASBT抑制活性,其中7个化合物10b_2、10c_3、10c_6、10d_2、10d_3、10d_6和10d_7在10μmol/L浓度下对ASBT的抑制率均大于90%,抑制活性明显高于阳性对照物S-1647(76.1%)。结论目标化合物中N1位连接链的长度对活性有较大的影响,当烷基链为5～7个碳原子时化合物的ASBT抑制活性较好,其中烷基链的碳原子数为7的化合物10d的ASBT抑制活性最好;连接链末端为季铵盐取代的化合物ASBT抑制活性明显优于叔胺取代的化合物。     

Design, synthesis of novel quinazolone alkaloids derivatives as potential antitumor agents

Several novel quinazolone alkaloids derivatives were synthesized. Some of the target compounds were determined against human prostate cancer DU145 and pancreatic cancer Miacapa2 cells in vitro. The entire compounds had been identified by 1HNMR, 13CNMR, IR, MS and EA.     

Microtubule depolymerizing vascular disrupting agents: novel therapeutic agents for oncology and other pathologies

Vascular disrupting agents (VDAs) are a relatively new group of 'vascular targeting' agents that exhibit selective activity against established tumour vascular networks, causing severe interruption of tumour blood flow and necrosis to the tumour mass. Microtubule depolymerizing agents form by far the largest group of small molecular weight VDAs many of which, including lead compound disodium combretastatin A-4 3-O-phosphate (CA-4-P), are under clinical development for cancer. Although distinct from the angiogenesis inhibitors, VDAs can also interfere with angiogenesis and therefore constitute a potential group of novel drugs for the treatment of pathological conditions characterized by excessive angiogenesis, in addition to cancer. The endothelial cytoskeleton is the primary cellular target of this family of drugs, and some progress in understanding the molecular and signalling mechanisms associated with their endothelial disrupting activity has been made in the last few years. Susceptibility of tumour vessels to VDA damage is ascribed to their immature pericyte-defective nature, although the exact molecular mechanisms involved have not been clearly defined. Despite causing profound damage to tumours, VDAs fail to halt tumour growth unless used together with conventional treatments. This failure is attributed to resistance mechanisms, primarily associated with cells that remain viable within the tumour rim, and enhanced angiogenesis. The focus is now to understand mechanisms of susceptibility and resistance to identify novel molecular targets and develop strategies that are more effective.     

Inhibition of Ca2+-activated tension and myosin light chain phosphorylation in skinned smooth muscle strips by the phenothiazines

The antipsychotic phenothiazine drugs trifluoperazine, chlorpromazine, and promethazine inhibited Ca²⁺-activated tension in functionally skinned rabbit ileum and rabbit pulmonary artery strips. Exogenous calmodulin rapidly reversed the inhibition of tension. Inhibition of the endogenous myosin light chain kinase by these drugs resulted with ATP or its analog, ATPS, as a substrate. The evidence presented suggests the inhibition of Ca²⁺ activation of tension in skinned smooth muscle preparations by phenothiazines is due to the inhibition of Ca²⁺-activated phosphorylation of the 20,000 dalton myosin light chain by the endogenous myosin light chain kinase.     

Current landscape in the discovery of novel antibacterial agents

BackgroundStandard treatments against bacterial infections are becoming ineffective due to the rise of antibacterial resistance worldwide. Classical approaches to develop new antibacterial agents are not sufficient to fulfil the current pipeline, therefore new strategies are currently being devised in the field of antibacterial discovery.ObjectivesThe objective of this narrative review is to compile the most successful strategies for drug discovery within the antibacterial context that are currently being pursued.SourcesPeer-reviewed publications from the MEDLINE database with robust data addressing the discovery of new antibacterial agents in the current pipeline have been selected.ContentSeveral strategies to discover new antibacterials are described in this review: (i) derivatives of known antibacterial agents; the activity of a known antimicrobial agent can be improved through two strategies: (a) the modification of the original chemical structure of an antimicrobial agent to circumvent antibacterial resistance mechanisms and (b) the development of a compound that inhibits the mechanisms of resistance to an antibacterial agent; (ii) new antibacterial agents targeting new proteins; (iii) inhibitors of virulence factors; (iv) nanoparticles; (v) antimicrobial peptides and peptidomimetics; (vi) phage therapy and enzybiotics; and (vii) antisense oligonucleotides.ImplicationsThis review intends to provide a positive message affirming that several different strategies to design new antibacterial agents are currently being developed, and we are therefore confident that in the near future some of the most promising approaches will come to fruition.     

Schizophrenia and Evoked Potentials: Maximum Amplitude, Frequency of Peaks, Variability, and Phenothiazine Effects

The purpose of this study was to compare averaged visual evoked potentials AVEPs) in normal subjects and in schizophrenics off and on phenothiazine medication. Flashes of 4 intensities were used. Ss were tested 3 times within a 1 month period. Measures of maximum amplitude (Am), frequency of peaks I FOP), and variability (V) were obtained- With increases in stimulus intensity all Ss showed increases in Ams and decreases in FOPs and V. Schizophrenics had smaller Ams. greater FOPs, and grear V than normals. Schizoprenies on phenothiazines generally had less FOPs initially and after 1 wk mi medication but not after about I month on medication. There was no consistently significant effect of phenothiazines on maximum amplitude. NO drug effect on variability was observed. Schizophrenics showed a decrease in FOPs over time while normals showed an increase. A relationship was found between variability and overall thought disturbance. Changes in clinical condition over a month were not associated with discernible AVEP changes. Relationships between Am, FOP, unit V are presented.     

Computer-aided analysis of the influence of digitizing and surfacing on the accuracy in dental CAD/CAM technology

In dentistry, ceramic materials with high fracture resistance are needed for all-ceramic fixed partial dentures (FPDs). The sophisticated processing of advanced ceramics that can be used for such dental restorations demands the application of CAD/CAM technologies. These techniques necessitate digitizing of the prepared teeth or the planned restoration itself and surfacing of the acquired digital data before milling paths can be generated. As precision in fit is crucial for dental restorations, a computer-aided method for the quantitative and qualitative 3D analysis has been developed and applied. Factors influencing the obtainable precision in the application of CAD/CAM techniques were taken into consideration.