常见非发酵菌的临床分布及耐药性分析

目的了解医院常见非发酵菌的临床分布及耐药特征,为临床合理使用抗菌药物提供依据。方法对2012-2014年医院临床送检各类标本检出的病原菌构成比、标本来源、科室分布、MDR、XDR、PDR菌株分离率及药敏结果进行统计分析。结果 2012-2014年共分离出非发酵菌5 270株,占全部分离菌株的27.48%。分布构成比:主要为鲍曼不动杆菌(占50.70%),铜绿假单胞菌占38.35%,嗜麦芽寡养单胞菌占4.95%。5 270株非发酵菌主要分离自痰液,占66.26%,其他标本类型均在7%以下。科室分布主要以第一重症监护病房(占21.14%)和第一呼吸内科病房(含内科重症监护病房,占12.33%)为主;其余科室分离率均在10%以下。MDR、XDR、PDR鲍曼不动杆菌和铜绿假单胞菌除痰标本外的标本类型分离率远低于总分离率。3年医院鲍曼不动杆菌对常用抗菌药物均产生了较严重的耐药性,大多数耐药率>80%。铜绿假单胞菌对多种抗菌药物的耐药率均>90%,对亚胺培南、美罗培南耐药率呈逐年上升趋势,已超过40%。嗜麦芽寡养单胞菌的耐药率总体上升比较缓慢。除痰标本外,其他标本的鲍曼不动杆菌和铜绿假单胞菌对绝大部分抗菌药物的耐药率低于总耐药率(P<0.05),而嗜麦芽寡养单胞菌对抗菌药物的耐药率与总耐药率比较差异无统计学意义(P>0.05)。结论非发酵菌对常用抗菌药物的耐药率较高,耐药机制多样复杂,临床在治疗非发酵菌感染时,应结合实验室药敏结果,合理使用抗菌药物,有效地控制医院感染。     

药物临床试验伦理审查质量风险的管理研究

目的:为提高我国药物临床试验伦理审查质量、保障受试者权益提供参考。方法:以风险管理理论为指导,运用文献研究法、专家意见法和层次分析法对药物临床试验伦理审查流程进行梳理,提取并确定影响伦理审查质量的风险因素及其权重,并对我国药物临床试验伦理审查工作提出改进建议。结果与结论:构建的药物临床试验伦理审查风险指标体系包括5个方面共31个影响因素,其中影响药物临床试验伦理审查质量的5个方面的重要性(权重)排序依次为医学伦理委员会的自身建设(0.2635)、审查会议的管理(0.2514)、跟踪审查(0.1945)、审查申请的受理与处理(0.1892)、文件档案的管理(0.1014);权重较高的影响因素包括"讨论投票过程利益冲突人员回避(0.0787)""对方案修改、知情同意修改、严重不良事件等情形及时审查或召开会议讨论(0.0705)""明确伦理审查工作流程及时间并对外公示(0.0598)""统一、规范的审查标准与批准标准(0.0521)"等。建议可通过讨论投票环节利益冲突人员予以回避,对方案修改、知情同意修改、严重不良事件等情形及时审查或召开伦理会议讨论,明确伦理审查工作流程及时间,建立统一、规范的审查标准和批准标准等方式提高药物临床试验伦理审查的质量。     

A highly tumor-targeted nanoparticle of podophyllotoxin penetrated tumor core and regressed multidrug resistant tumors

Podophyllotoxin (PPT) exhibited significant activity against P-glycoprotein mediated multidrug resistant (MDR) tumor cell lines; however, due to its poor solubility and high toxicity, PPT cannot be dosed systemically, preventing its clinical use for MDR cancer. We developed a nanoparticle dosage form of PPT by covalently conjugating PPT and polyethylene glycol (PEG) with acetylated carboxymethyl cellulose (CMC-Ac) using one-pot esterification chemistry. The polymer conjugates self-assembled into nanoparticles (NPs) of variable sizes (20–120 nm) depending on the PPT-to-PEG molar ratio (2–20). The conjugate with a low PPT/PEG molar ratio of 2 yielded NPs with a mean diameter of 20 nm and released PPT at ∼5%/day in serum, while conjugates with increased PPT/PEG ratios (5 and 20) produced bigger particles (30 nm and 120 nm respectively) that displayed slower drug release (∼2.5%/day and ∼1%/day respectively). The 20 nm particles exhibited 2- to 5-fold enhanced cell killing potency and 5- to 20-fold increased tumor delivery compared to the larger NPs. The biodistribution of the 20 nm PPT-NPs was highly selective to the tumor with 8-fold higher accumulation than all other examined tissues, while the larger PPT-NPs (30 and 120 nm) exhibited increased liver uptake. Within the tumor, >90% of the 20 nm PPT-NPs penetrated to the hypovascular core, while the larger particles were largely restricted in the hypervascular periphery. The 20 nm PPT-NPs displayed significantly improved efficacy against MDR tumors in mice compared to the larger PPT-NPs, native PPT and the standard taxane chemotherapies, with minimal toxicity.     

Plasma hydrogenated cationic detonation nanodiamonds efficiently deliver to human cells in culture functional siRNA targeting the Ewing sarcoma junction oncogene

The expression of a defective gene can lead to major cell dysfunctions among which cell proliferation and tumor formation. One promising therapeutic strategy consists in silencing the defective gene using small interfering RNA (siRNA). In previous publications we showed that diamond nanocrystals (ND) of primary size 35 nm, rendered cationic by polyethyleneimine-coating, can efficiently deliver siRNA into cell, which further block the expression of EWS/FLI-1 oncogene in a Ewing sarcoma disease model. However, a therapeutic application of such nanodiamonds requires their elimination by the organism, particularly in urine, which is impossible for 35 nm particles. Here, we report that hydrogenated cationic nanodiamonds of primary size 7 nm (ND-H) have also a high affinity for siRNA and are capable of delivering them in cells. With siRNA/ND-H complexes, we measured a high inhibition efficacy of EWS/FLI-1 gene expression in Ewing sarcoma cell line. Electron microscopy investigations showed ND-H in endocytosis compartments, and especially in macropinosomes from which they can escape before siRNA degradation occurred. In addition, the association of EWS/FLI-1 silencing by the siRNA/ND-H complex with a vincristine treatment yielded a potentiation of the toxic effect of this chemotherapeutic drug. Therefore ND-H appears as a promising delivery agent in anti-tumoral gene therapy.     

Novel nanostructured supramolecular hydrogels for the topical delivery of anionic drugs

A bis-imidazolium-based amphiphilic molecule was used to form novel supramolecular gels in ethanol–water mixtures. The proportion of solvents, the concentration of gellant and the temperature are factors that strongly influence the gelling process. The physical gels that are formed comprise entangled fibers of around 100 nm in diameter, able to incorporate anionic drugs, whose morphology varies depending on the drug they incorporate. These hydrogels are soft and therefore optimum for skin application. They show good stability when compared to previously reported gels. Suitable drug release and skin permeation profiles were obtained, and, moreover, they seem to promote the retention of the drug inside the skin. Finally, effective in vivo anti-inflammatory activity was observed, especially with the indomethacin-incorporated gel, which indicates that these supramolecular hydrogels are a good option for the delivery of poor water soluble drugs for the treatment of acute inflammation or other skin diseases.     

The tyrosine kinase inhibitor nilotinib has antineoplastic activity in prostate cancer cells but up-regulates the ERK survival signal—Implications for targeted therapies

BackgroundNovel therapeutic options beyond hormone ablation and chemotherapy are urgently needed for patients with advanced prostate cancer. Tyrosine kinase inhibitors (TKIs) are an attractive option as advanced prostate cancers show a highly altered phosphotyrosine proteome. However, despite favorable initial clinical results, the combination of the TKI dasatinib with docetaxel did not result in improved patient survival for reasons that are not known in detail.MethodsThe National Cancer Institute–Approved Oncology Drug Set II was used in a phenotypic drug screen to identify novel compounds with antineoplastic activity in prostate cancer cells. Validation experiments were carried out in vitro and in vivo.ResultsWe identified the TKI nilotinib as a novel compound with antineoplastic activity in hormone-refractory prostate cancer cells. However, further analyses revealed that treatment with nilotinib was associated with a significant up-regulation of the phospho-extracellular-signal-regulated kinases (ERK) survival signal. ERK blockade alone led to a significant antitumoral effect and enhanced the cytotoxicity of nilotinib when used in combination.ConclusionsOur findings underscore that TKIs, such as nilotinib, have antitumoral activity in prostate cancer cells but that survival signals, such as ERK up-regulation, may mitigate their effectiveness. ERK blockade alone or in combination with TKIs may represent a promising therapeutic strategy in advanced prostate cancer.     

依维莫司联合全反式维甲酸逆转急性早幼粒细胞白血病细胞耐药的研究

目的探讨依维莫司联合全反式维甲酸(简称维甲酸)逆转急性早幼粒细胞白血病(APL)细胞株NB4-R1耐药的作用。方法应用CD11b染色流式细胞术及硝基四唑氮蓝(NBT)还原实验检测两药联合应用对细胞分化的影响, 流式细胞术检测细胞周期情况, Annexin V/PI双染色检测细胞凋亡情况, 蛋白质印迹法检测自噬相关蛋白微管结合蛋白轻链3(LC3)、Beclin 1及早幼粒白血病-维甲酸受体融合蛋白(PML-RARα)、磷酸化核糖体S6激酶(P-P70S6K)、磷酸化4E结合蛋白1(P-4E-BP1) 等表达水平。结果与维甲酸组比较, 联用组能诱导耐药细胞株NB4-R1细胞的分化, 并将细胞增殖阻止在G ₁期而对细胞凋亡无明显影响。100 nmol/L依维莫司组、1μmol/L维甲酸组、联用组、对照组NB4-R1细胞培养48 h后分化百分率分别为(2.29±0.57)%、(17.06±2.65)%、(54.47±4.91)%、(2.54±0.53)%; 处于G ₁期的细胞百分率分别为(35.20±11.97)%、(33.54±6.25)%、(53.70±8.73)%、(27.40±6.01)%; 四组细胞凋亡细胞百分率分别为(2.30±0.14)%、(2.25±0.21)%、(2.40±0.28)%、(1.95±0.07)%。与维甲酸组比较, 联用组mTOR信号通路下游的P70S6K、4E-BP1分子磷酸化水平下降, LC3-II和Beclin 1的表达上调, 且能部分降解融合蛋白PML-RARα。 结论依维莫司联合维甲酸能诱导NB4-R1细胞分化, 且能阻滞细胞周期而不致细胞凋亡, 其机制可能与依维莫司联合维甲酸抑制mTOR信号通路激活自噬作用从而降解PML-RARα蛋白有关。     

A new self-emulsifying formulation of mefenamic acid with enhanced drug dissolution

To enhance the dissolution of poorly soluble mefenamic acid, self-emulsifying formulation (SEF), composing of oil, surfactant and co-surfactant, was formulated. Among the oils and surfactants studied, Imwitor^® 742, Tween^® 60, Cremophore^® EL and Transcutol^® HP were selected as they showed maximal solubility to mefenamic acid. The ternary phase diagram was constructed to find optimal concentration that provided the highest drug loading. The droplet size after dispersion and drug dissolution of selected formulations were investigated. The results showed that the formulation containing Imwitor^® 742, Tween^® 60 and Transcutol^® HP (10:30:60) can encapsulate high amount of mefenamic acid. The dissolution study demonstrated that, in the medium containing surfactant, nearly 100% of mefenamic acid were dissolved from SEF within 5 min while 80% of drugs were dissolved from the commercial product in 45 min. In phosphate buffer (without surfactant), 80% of drug were dissolved from the developed SEF within 5 min while only about 13% of drug were dissolved in 45 min, from the commercial product. The results suggested that the SEF can enhance the dissolution of poorly soluble drug and has a potential to enhance drug absorption and improve bioavailability of drug.