Enhanced hit-to-lead process using bioanalogous lead evolution and chemogenomics: application in designing selective matrix metalloprotease inhibitors

The authors describe an innovative approach for designing novel inhibitors. This approach effectively integrates the emerging chemogenomics concept of target-family-based drug discovery with bioanalogous design strategies, including privileged structures, molecular frameworks as well as bioisosteric and bioanalogous/isofunctional modifications.

The authors applied this method in the design of selective inhibitors of matrix metalloproteases (MMPs), also referred to as matrixins, on the basis of a unique analysis of the ligand–target knowledge base, the ‘matrixinome’. For this analysis, the authors created an annotated MMP database containing ～ 300 inhibitors with their published activity profile. The ligand space was then arranged into a lead evolution tree, where the substructural transformations in each virtual step led to marked changes in the activity pattern. This allowed subtype-specific privileged fragments to be extracted as well as modifications, which improve activity and/or selectivity. Furthermore, the compounds with the preferred activity profile were correlated with sequence homology as well as binding site similarity within the target family, thereby leading to the identification of substructural modifications that turn non-selective, biohomologous structures into selective inhibitors. The matrixinomic application of the authors’ approach, therefore, provides an example of how the combination of ligand space knowledge with sequence-related data can radically improve the outcome of the lead optimisation process to achieve higher selectivity within a given target family.     

Improving the hit-to-lead process: data-driven assessment of drug-like and lead-like screening hits

Drug-like and lead-like hits derived from HTS campaigns provide good starting points for lead optimization. However, too strong emphasis on potency as hit-selection parameter might hamper the success of such projects. A detailed absorption, distribution, metabolism, excretion and toxicology (ADME-Tox) profiling is needed to help identify hits with a minimum number of (known) liabilities. This is particularly true for drug-like hits. Herein, we describe how to break down large numbers of screening hits and we provide a comprehensive overview of the strengths and weaknesses for each structural class. The overall profile (e.g. ligand efficiency, selectivity and ADME-Tox) is the distinctive feature that will define the priority for follow-up.     

正交实验法优选五味子中木脂素的提取工艺

目的 考察五味子中木脂素成分的提取工艺.方法 以五味子醇甲、五味子乙素提取效率为指标,采用正交试验优选提取方法和工艺参数.结果 五味子中木脂素的最佳提取工艺条件为:8倍量70％乙醇回流提取(1 h×2).结论 正交试验确定的五味子木脂素提取工艺条件合理可行,可为工业化大生产提供参考.     

优化枳椇子的提取工艺

目的　确定枳椇子干浸膏的最佳提取工艺?椒ā∫愿山嗟寐屎透山嘀凶芑仆课副?,对索式提取器提取的方法、渗滤法和浸渍法进行筛选。结果　渗滤法出膏率和干浸膏中黄酮含量最高,索式提取次之,浸渍法出膏率和干浸膏中黄酮含量最低。结论　渗滤法提取干浸膏简单易行,成本低廉,适合批量生产。     

均匀设计优化三七总皂苷提取工艺

目的优选三七总皂苷提取工艺的最佳条件。方法通过均匀设计试验。结果提取溶剂用量为药材重量的 1 3倍 ,乙醇浓度为 30 % ,提取温度为 70℃ ,加热时间 4 0min,同法提取 2次 ,三七总皂苷平均提取率为 94 92 %。结论三七总皂苷提取率高、生产成本低 ,适合工业生产。     

正交试验优选芩莲口服液中黄芩苷的提取工艺

目的优选芩莲口服液的最佳提取工艺。方法以芩莲口服液中有效成分黄芩苷的含量为指标,采用正交试验法优选芩莲口服液提取工艺。结果芩莲口服液的最佳提取工艺：料液比为1∶8,浸泡2 h,煎煮2次,每次2 h。结论优选出的提取工艺合理、稳定、经济。     

正交试验法优选功血饮颗粒的醇提工艺

目的优选功血饮颗粒的最佳醇提工艺。方法采用正交试验法,以芍药苷和人参皂苷Rg1及浸出物为指标,对功血饮颗粒的醇提工艺进行优选。结果最佳醇提工艺为：加饮片总量为18倍量的70％乙醇分3次回流提取,每次2h。结论该工艺稳定,可行。     

多指标综合评价结合层次分析法优化黄芩微波酒制工艺

目的 采用正交设计综合加权评分和层次分析法相结合的方法,优化黄芩微波酒制工艺。方法 采用单因素试验优选加酒量、闷润时间、微波功率、微波时间,然后以微波酒制工艺中的加酒量、闷润时间、微波功率、微波时间为考察因素,采用层次分析法确定各指标权重系数,以黄芩苷、汉黄芩苷、黄芩素、汉黄芩素含量进行综合加权评分为评价指标,优化黄芩微波酒制工艺。结果 取生黄芩饮片适量,加入10%辅料酒,置密闭容器内闷润60 min,在300 W功率下微波5 min,经3次验证试验,各指标成分的平均含量分别为15.65%、5.94%、3.00%、0.33%,平均评分98.24,RSD为0.99%(n=3)。结论 优化所得微波炮制酒工艺稳定,重复性良好,可应用于微波酒黄芩的炮制。     

正交试验优选重连口服液的提取工艺

目的优选重连口服液最佳提取工艺。方法水提取工艺以浸膏得率、重楼皂苷、Ⅰ和连翘苷的含量为考察指标,醇沉工艺以重楼皂苷Ⅰ和连翘苷的含量为考察指标,分别采用正交试验法对其提取工艺条件进行优选。结果重连口服液水提最佳工艺条件为加水煎煮3次,每次10倍量水,煎煮1h,醇沉最佳工艺条件为浓缩至相对密度1．10（80℃热测）,加乙醇使药液乙醇浓度达到80％,静置24h。结论优选的提取工艺合理,稳定性好,可为工业生产提供理论依据。     

硝苯地平海藻酸钙凝胶缓释微丸处方及工艺的优化

目的制备硝苯地平 海藻酸钙凝胶缓释微丸。方法采用滴制法。用单因素考察及均匀设计实验优化处方及工艺。结果优化得到的微丸在体外累计释放硝苯地平百分率 ,2h为 2 0 %～ 3 0 % ,6h为 60 %～ 80 % ,1 2h>85 %。结论在体外释放度实验中 ,硝苯地平海藻酸钙凝胶微丸具有良好的缓释作用     

均匀设计优化竹节参总皂苷提取工艺

目的优化超声波提取竹节参总皂苷的工艺。方法以竹节参总皂苷得率为评价指标,以乙醇浓度、溶剂倍数、提取时间、提取温度为考察因素,采用均匀设计实验优化竹节参总皂苷的提取工艺。结果优化的提取工艺为乙醇浓度为50％、乙醇用量为12倍、提取时间为40min,提取温度为30％。结论优化的提取工艺提取率高,操作简便,可为竹节参总皂苷工业化提取提供重要的参考。     

正交实验优选丹葛保心颗粒提取工艺

目的优选丹葛保心颗粒提取工艺。方法根据丹参所含成分和药物有效成分,以丹参酮IIA的含量和干膏得率为指标,以乙醇浓度、用量、提取次数、提取时间为考察因素,筛选丹葛保心颗粒最佳醇提取工艺。以葛根素的含量和干膏得率为指标,以加水量、煎煮时间和提取次数为考察因素,筛选丹葛保心颗粒最佳水提取工艺条件。结果按处方量药物,丹参用10倍量75%乙醇回流提取二次,每次1.5 h。丹参药渣与其余二味药材加10倍量水煎煮2次,每次2 h。结论优选丹葛保心颗粒的提取工艺较稳定,重现性好,可为工业生产提供理论依据。     

正交试验法优选新藤黄酸的醇提工艺

目的优选藤黄中新藤黄酸的提取工艺。方法以新藤黄酸为考察指标,用HPLC法进行含量测定,采用正交试验法优化提取条件。结果最佳醇提条件为先以甲醇（pH=7）常温浸提,时间为20min,再回流提取5h,共2次。结论醇提的次数对新藤黄酸的提取影响最大,该研究确定的新藤黄酸的提取工艺合理、可行。     

Evolution of the thienopyridine class of inhibitors of IkappaB kinase-beta: part I: hit-to-lead strategies

High-throughput screening is routinely employed as a method for the identification of novel hit structures. Large numbers of active compounds are typically procured in this way and must undergo a rigorous validation process. This process is described in detail for a collection of screening hits identified as inhibitors of IkappaB kinase-beta (IKKbeta), a key regulatory enzyme in the nuclear factor-kappaB (NF-kappaB) pathway. From these studies, a promising hit series was selected. Subsequent lead generation activities included the development of a pharmacophore hypothesis and structure-activity relationship (SAR) for the hit series. This led to the exploration of related scaffolds offering additional opportunities, and the various structural classes were comparatively evaluated for enzyme inhibition, selectivity, and drug-like properties. A novel lead series of thienopyridines was thereby established, and this series advanced into lead optimization for further development.     

优化RCA分析法改良肿瘤患者给药

目的针对肿瘤患者用药错误,按照用药规律分组,优化原有RCA分析法,扩大受众范围,降低用药错误发生率,促进肿瘤患者的给药安全。方法回顾性分析某三级甲等肿瘤专科医院2014年1-12月护理管理系统主动上报的用药错误事件101起,进行优化RCA分析方法分析,以制定整改措施,并与整改措施实施后的2015年1-12月进行比较。结果优化方法实施前、后,肿瘤患者用药错误事件发生率比较差异有统计学意义(P<0.01)。结论实施优化RCA分析方法可以增强护士对肿瘤患者用药错误事件风险意识,扩大分享面,促进制度流程进一步规范,促进肿瘤患者给药安全。     

Optimizing delivery of flurbiprofen to the colon using a targeted prodrug approach

The carboxylic group responsible for the gastric side-effects of the propionic acid derivative, flurbiprofen, was masked temporarily to overcome these side-effects and to accomplish colon-specific delivery of the drug. An amide prodrug (FLU-GLY) was synthesized by coupling flurbiprofen with L-glycine. Confirmation and characterization of the structure of the synthesized prodrug included elemental analysis, Fourier transform (FT)-IR, FT-NMR, mass (FAB) spectroscopy, and determinations of R(f), R(t) and R(M) values, respectively. Aqueous solubility and lipophilicity (logP) value were determined at pH 1.2, 4.0, 6.8 and 7.4. In-vitro reversion of FLU-GLY to flurbiprofen was measured at different pHs and in a simulated colonic environment. Acute toxicity and ulceration potential were evaluated in-vivo in albino rats. Pre-formulation studies showed increased hydrophilicity but a non-significant increase in lipophilicity of the prodrug. In-vitro reversion studies suggested that the prodrug remained intact until colonic pH was attained, when the colonic microfloral enzymes (amidase) hydrolysed the FLU-GLY amide linkage, releasing the free drug. In-vivo evaluation indicated that the prodrug was much less toxic and had less ulcerogenic activity than the parent drug. Selective delivery of drugs to the colon can be useful in terms of reducing the dose administered and reducing undesirable side-effects.     

Biosensor analysis of the molecular interactions of pentosan polysulfate and of sulfated glycosaminoglycans with immobilized elastase,hyaluronidase and lysozyme using surface plasmon resonance (SPR) technology

Pentosan polysulfate (NaPPS) and chondroitin sulfates (ChSs) have recently been shown to exhibit both symptom and disease modifying activities in osteoarthritis (OA), but their respective mechanisms of action are still the subject of conjecture. Excessive catabolism of joint articular cartilage is considered to be responsible for the initiation and progression of OA but the abilities of these drugs to mitigate this process has received only limited attention. Human neutrophil elastase (HNE) is a proteinase, which can degrade the collagens and proteoglycans (PGs) of the cartilage directly or indirectly by activating latent matrix metalloproteinases. Hyaluronidase (HAase) is an endoglycosidase, which degrades glycosaminoglycans including hyaluronan, which provides the aggregating component of the PG aggrecan complex. In the present study the molecular interactions between the NaPPS, ChSs and some other sulfated polysaccharides with immobilized HNE, HAase or lysozyme (a cationic protein implicated in PG metabolism) were studied using a SPR biosensor device-BIAcore2000. The above three enzymes were covalently immobilized to a biosensor chip CM5 separately using amine coupling. The binding affinity of each sulfated polysaccharide and the kinetics of NaPPS over the concentration range of 0.3-5.0 microg/ml were determined. The inhibition of HNE by the sulfated polysaccharides as determined using the synthetic substrate succinyl-Ala-Ala-Val-nitroanilide (SAAVNA) in a functional assay was compared with their respective binding affinities for this proteinase using the BIAcore system. The results obtained with the two independent techniques showed good correlation and indicated that the degree and ring positions of oligosaccharide sulfation were major determinants of enzyme inhibitory activity. The observed difference in order of binding affinities of the drugs to the immobilized HNE, HAase and lysozyme suggests a conformational relationship, in addition to the charge interactions between the sulfate esters of the polysaccharides and the cationic amino acids of the enzymes. Significantly, the SPR biosensor technology demonstrated that small differences among sulfated polysaccharides, even subtle variations among different NaPPS batches, could be readily detected. The SPR technology therefore offers not only a sensitive and reproducible method for ranking noncompetitive enzyme inhibitors for drug discovery but a rapid and quantitative bioassay for monitoring batch consistency of manufacture.     

Optimizing double emulsion process to decrease the burst release of protein from biodegradable polymer microspheres

The process parameters such as the compositions of inner and outer aqueous phase and emulsification technique of the primary emulsion were optimized to decrease the burst release of BSA from biodegradable polymer microspheres in double emulsion method. It was found that diminished burst release of -14% was achieved for the microspheres produced by formulations, where no phosphate was present in the inner water phase (non-buffered system). Primary emulsion made by probe sonication rather than homogenization or mechanical stirring led to microspheres with insignificant burst effect. Microspheres obtained using 0.1% aqueous Tween 80 solution as the outer aqueous phase, frequently exhibit reduced burst effect of 2.7%. Low microsphere yield (52.1%), however, was observed. Microsphere yield was, therefore, enhanced by addition of additive such as sodium chloride, glucose or mannitol into the outer aqueous phase. Decrease in BSA entrapment was observed in the presence of sodium chloride, but reduction in entrapment efficiency was observed in the case of glucose. Burst release increased from 2.7% to 9.5% or 3.4% as 2.5% sodium chloride or 7.5% glucose was added into the outer aqueous phase respectively. Marked burst release (>20%) was observed in the presence of additive of higher concentration independent of sodium chloride or glucose. As far as surfactant type was concerned, diminished burst was found when PVP or Tween 80 rather than PVA was utilized as the surfactant during microsphere preparation. In addition to PLGA, the copolymers of L-lactide (LLA) and dimethyl trimethylene carbonate (DTC) or trimethylene carbonate (TMC) were also evaluated. Insignificant burst effect was found for the microspheres composed of DTC or TMC copolymers.     

响应面法优化凤丹内生真菌发酵工艺探究

目的 研究响应面法优化一株凤丹优势内生真菌发酵工艺。 方法 在单因素试验结果的基础上,运用响应面法设计,对凤丹优势内生真菌发酵工艺进行优化。 结果 最优发酵工艺为蔗糖含量20 g·L-1,酵母浸膏含量5 g·L-1,KH₂PO₃含量1.00 g·L-1,K₂HPO₃含量1.00 g·L-1。在该条件下,发酵所得的菌丝体实际生物量为1.647 g。 结论 实验所得数据的多元回归模型与实际情况较好拟合,表明该优化工艺稳定可靠,通过响应面法最终获得了凤丹内生真菌最优发酵工艺。