重组水蛭素-2鼻腔黏附纳米粒的体外评价

目的:对制备的重组水蛭素-2(rHV2)壳聚糖鼻腔黏附纳米粒进行体外评价。方法:测定纳米粒的形态、粒径大小分布及表面电位;以超速离心测定纳米粒在不同介质中的释放度;采用在体蟾蜍上腭纤毛运动试验法考察纳米粒的纤毛毒性。结果:制备的rHV2壳聚糖纳米粒呈较为均匀分散的颗粒状,平均粒径为213.2 nm,纳米粒带正电荷,Zeta电位值为+30.61 mV,体系相对稳定性较高;rHV2纳米粒在乙酸缓冲液中的累积释放百分数明显高于在磷酸缓冲液中的累积释放百分数;rHV2纳米粒溶液的纤毛毒性较小(与生理盐水组相比,P>0.05)。结论:壳聚糖鼻腔黏附纳米粒有望成为rHV2鼻腔给药的递释系统。     

重组水蛭素-2鼻腔喷雾剂的药代动力学和药效学

目的研究重组水蛭素-2(rHV2)喷雾剂鼻腔给药后在大鼠体内的药代动力学过程及其抗凝血作用。方法采用呈色肽法测定大鼠鼻腔给予rHV2喷雾剂后血浆中rHV2的浓度，并计算其药代动力学参数；考察rHV2鼻腔喷雾剂对正常大鼠和弥散性血管内凝血模型(DIC)家兔凝血时间的影响。结果rHV2喷雾剂鼻腔给药后，rHV2的体内过程符合一室模型，其相对生物利用度为28.53%；rHV2鼻腔喷雾剂可使正常大鼠活化的部分凝血活酶时间(APTT)和凝血酶时间(TT)明显延长，使DIC模型家兔的APTT凝血时间明显缩短，接近正常值。结论药代动力学和药效学结果证明，rHV2鼻腔喷雾剂有望成为rHV2鼻腔给药的有效制剂。     

水飞蓟素固体脂质纳米粒在大鼠体内药代动力学研究

目的:研究尾静脉注射水飞蓟素固体脂质纳米粒(SM-SLN)在大鼠体内的药代动力学。方法:HPLC法测定大鼠尾静脉注射SM-SLN及水飞蓟素溶液后,不同时间点眼眶取血,测定血浆中的药物浓度,采用3P97软件,对平均血药浓度-时间曲线进行拟合,并计算主要药动参数。结果:通过拟合后,平均血药浓度-时间曲线在大鼠体内符合二室模型,水飞蓟素溶液及SM-SLN药时曲线下面积面积分别为4.322μg·mL-1·h-1和12.421μg·mL-1·h-1,消除半衰期(t1/2β)分别为4.247h和8.298h。结论:与水飞蓟素溶液组相比,SM-SLN延长了在体内的驻留时间,具有了一定的缓释效果。     

建立用生色底物法研究重组水蛭素在大鼠血浆中的药动学

目的:建立用生色底物法(chromogenic substrate assay,CSA)测定大鼠血浆中重组水蛭素(re-combinant hirudin,rH)含量的方法,并用该法研究rH在大鼠血浆中的药动学。方法:以TGPApNA(tosyl-gly-cyl-polyl-arginine-para-nitraniline)为底物,凝血酶(thrombin,Th)能水解TGPApNA释放出对-硝基苯胺(para-nitraniline,pNA),pNA在405nm处有特异性吸收,通过测定反应体系中未被rH中和的Th量,间接测定rH的活性。结果:CSA法能有效地测定大鼠血浆中rH的含量;rH的浓度在3.125~40 ng.mL-1范围内呈线性关系;日间和日内精密度相对标准偏差均符合要求。应用上述方法测定大鼠iv 2.0 mg.kg-1的rH后不同时间的血药浓度,结果表明该药符合二室模型一级动力学过程。结论:用生色底物法测定大鼠血浆中rH具有准确、可靠、重现性好的特点,可用于rH药动学的研究。     

蛇床子素-pH敏感性纳米粒的制备及其在大鼠体内的药代动力学

目的:制备蛇床子素-Eudragit S100纳米粒(Ost-S100-NP)及其冻干制剂(Fd-Ost-S100-NP),并考察相关特性;考察蛇床子素、纳米粒制剂和冻干制剂在大鼠体内的药代动力学及相对生物利用度。方法:以乳化-溶剂扩散法联合冷冻干燥技术制备Ost-S100-NP胶体溶液及其冻干粉末;动态透析法研究其体外释药动力学;以HPLC测定大鼠灌胃后血浆中的Ost浓度,3P97程序计算药代动力学参数。结果:Ost-S100-NP和Fd-Ost-S100-NP的平均粒径分别为(55.4±2.1)和(87.7±2.3)nm;包封率>95%;电镜下呈均匀规则的圆球形;且纳米粒制剂在pH>6.0的释放介质中释药速率明显增大,具有显著pH敏感性;药物在大鼠体内药代动力学均符合二室模型,Ost-S100-NP和Fd-Ost-S100-NP的相对生物利用度分别为原料药的161.1%和118.4%。结论:以pH敏感性材料Eudragit S100为载体有望开发成一种高效、低毒的蛇床子素纳米制剂。     

PEG-PLA-α-细辛脑纳米粒经鼻腔、静脉给药后的药动学研究

目的 采用与静脉注射对比的方式,研究聚乙二醇-聚乳酸-α-细辛脑纳米粒（PEG-PLA-α-细辛脑纳米粒）鼻腔给药后在大鼠体内的药物动力学。方法 以大鼠为动物模型,采用血药动力学、脑药动力学及荧光标记法对比研究PEG-PLA-α-细辛脑纳米粒经鼻腔给药与静脉注射后药物/纳米粒在大鼠体内的分布情况。结果 PEG-PLA-α-细辛脑纳米粒静脉注射及鼻腔给药后血浆中的AUC_（0-∞）分别为（11032.4±1 827.1）ng·mL^-1·min及（5 992.9±717.5）ng·mL^-1·min,C_max分别为（421.9±100.2）ng·mL^-1及（171.7±26.3）ng·mL^-1,PEG-PLA-α-细辛脑纳米粒鼻腔给药后的绝对生物利用度F为54.3%。PEG-PLA-α-细辛脑纳米粒静脉注射后α-细辛脑在脑组织中的C_max与鼻腔给药后α-细辛脑在脑组织中的浓度C_max分别为（217.9±29.9）ng·mL^-1及（334.2±62.7）ng·mL^-1,PEG-PLA-α-细辛脑纳米粒静脉注射与鼻腔给药后的AUC_brain/AUC_plasma值分别为1.37和2.85,且两者具有统计学意义。PEG-PLA-α-细辛脑纳米粒鼻腔给药后的药物脑靶向效率及鼻-脑传递百分比分别为208.03%及52.01%。荧光标记法结果显示,PEG-PLA-α-细辛脑纳米粒鼻腔给药后脑靶向性比静脉注射后更强。结论 PEG-PLA-α-细辛脑纳米粒适合于鼻腔给药治疗脑部疾病。     

姜黄素-PLGA纳米粒提高口服给药生物利用度的研究

目的 研究乳酸/羟基乙酸共聚物（PLGA）纳米粒子提高姜黄素口服生物利用度。方法 采用乳液挥发法制备姜黄素-PLGA纳米粒;通过透射电镜（transmission electron microscope,TEM）观察纳米粒形态;采用动态光散射法（dynamic light scattering,DLS）测定纳米粒大小、表面电位（Zeta电位）;考察药物的体外稳定性以及药物释放行为;以大鼠口服灌胃给药方式考察姜黄素和姜黄素-PLGA纳米粒的体内药物生物利用度。结果 姜黄素-PLGA纳米粒粒度分布均匀,平均粒径大小约200 nm;姜黄素-PLGA纳米粒具有较高的载药量和包封率以及稳定性,体外药物释放实验结果显示具有一定的缓释效果;口服灌胃100 mg·kg^-1姜黄素和姜黄素-PLGA纳米粒,给药30 min之后,姜黄素-PLGA纳米粒给药组的血药浓度水平显著高于姜黄素组（P〈0.05）,药物生物利用度提高到原来的5.2倍。结论 姜黄素-PLGA纳米粒可以有效的提高姜黄素稳定性和口服给药生物利用度。     

重组人血管内皮抑制素壳聚糖纳米粒的制备

目的:通过星点设计-效应面法优化重组人血管内皮抑制素(Endostar)壳聚糖纳米粒的制备工艺。方法:以壳聚糖浓度、三聚磷酸钠(TPP)浓度及壳聚糖溶液与TPP溶液体积比为考察因素,以载药量、包封率和粒径为指标,采用多元线性回归和多元非线性回归拟合选择合适模型,并根据最佳模型绘制效应面图,选择最佳处方,并进行预测分析,同时考察最佳处方的体外释放。结果:用多元非线性回归对实验中各因素和指标进行拟合明显优于线性拟合。最佳处方制备的Endostar载药壳聚糖纳米粒,载药量、包封率及粒径的实测值与预测值的偏差均在±7%以内,体外释放1周累积释放80%。结论:通过星点设计-效应面法可以准确快速优化处方工艺。优化后的工艺制备的纳米粒可以延缓Endostar的释放,达到了缓释效果,符合预期的试验目标。     

盐酸阿霉素壳聚糖纳米粒大鼠鼻腔给药的脑内药动学研究

目的研究盐酸阿霉素壳聚糖纳米粒(adriamycinhydrochloride chitosan nanoparticles,ADM-CS-NPs)大鼠鼻腔给药后的脑内药动学特征。方法以壳聚糖为载体材料,采用离子交联法制备ADM-CS-NPs。以清醒自由活动大鼠为实验动物模型,采用脑微透析取样技术,连续收集ADM-CS-NPs及阿霉素溶液经鼻腔给药与静脉注射给药后大鼠海马部位透析液,HPLC法测定药物浓度,分析数据计算药动学参数。结果 ADM-CS-NPs经大鼠鼻腔和尾静脉注射给药(给药剂量为1.45 mg.kg-1ADM)后,Tmax分别为(300.00±26.12)和(180.00±19.11)min,Cmax分别为(93.00±8.53)和(19.11±1.91)mg·L-1,AUC0→11h分别为(17809.05±650.24)和(5159.97±120.59)mg·h·L-1,MRT分别为(390.49±6.87)和(281.53±4.99)min;ADM-Sol经大鼠鼻腔和尾静脉注射给药后,Tmax分别为(20.00±2.91)和(20.00±2.00)min,Cmax分别为(90.00±7.31)和(10.70±0.96)mg·L-1,AUC0→11h分别为(4736.70±53.40)和(312.68±4.99)mg·h·L-1,MRT分别为(73.43±2.37)和(23.39±1.32)min。结论 ADM-CS-NPs鼻腔给药可增加药物脑内浓度,有效实现脑内递药,同时由于纳米粒的缓释作用,可延长脑内有效药物浓度的持续时间,发挥长效作用。     

盐酸小檗碱脂质纳米粒的药代动力学及生物等效性研究

目的 研究盐酸小檗碱（BH）和盐酸小檗碱脂质纳米粒（BHLN）在大鼠体内的药代动力学及其生物等效性。方法 采用溶剂蒸发法制备盐酸小檗碱脂质纳米粒,并测定其粒径与电位。将大鼠随机分组给予相同药量,于不同的时间点采集血样,用HPLC法测定血药浓度,用DAS（2.1.1版）软件分析主要的药代动力学参数及计算生物等效性。结果 BHLN的平均粒径和电位分别为（128.32±2.55） nm和（-30.34±0.83） mV。BHLN和盐酸小檗碱的T_max分别为0.75和1.50 h, AUC_{0-72 h}分别为1 192.35和353.47μg·h·L^-1,C_max分别是537.21和157.92μg·L^-1。BHLN的C_max较盐酸小檗碱增大了3.40倍,AUC_{0-72 h}是盐酸小檗碱的3.37倍,AUC_{0-72 h},C_max和T_max差异均有统计学意义（均P<0.05）。...     

Influence of chitosan coating on the oral bioavailability of gold nanoparticles in rats

Gold nanoparticles are one of the most extensively investigated metallic nanoparticles for several applications. It is less toxic than other metallic nanolattices. The exceptional electrical and thermal conductivity of gold make it possible to be administered as non-invasive radiofrequency irradiation therapy that produces sufficient heat to kill tumor cells. Nanoparticles are generally administered intravenously instead of orally due to negligible oral absorption and cellular uptake. This study evaluated the oral bioavailability of gold nanoparticles coated with chitosan (C-AuNPs), a natural mucoadhesive polymer. We employed traditional method of evaluating bioavailability that involve estimation of maximum concentrations and area under the curve of 3?nm chitosan coated gold nanoparticles (C-AuNPs) in the rat plasma following intravenous and oral administrations (0.8?mg and 8?mg/kg body weight respectively). The oral bioavailability of C-AuNPs was found to be 2.46% (approximately 25 folds higher than polyethylene glycol (PEG) coated gold nanoparticles, reported earlier). These findings suggest that chitosan coating could be better than PEG coating for the enhancement of oral bioavailability of nanoparticles.     

Effects of Li(+) transport and Li(+) immobilization on Li(+)/Mg(2+) competition in cells: implications for bipolar disorder

Li(+)/Mg(2+) competition has been implicated in the therapeutic action of Li(+) treatment in bipolar illness. We hypothesized that this competition depended on cell-specific properties. To test this hypothesis, we determined the degree of Li(+) transport, immobilization, and Li(+)/Mg(2+) competition in lymphoblastomas, neuroblastomas, and erythrocytes. During a 50 mM/L Li(+)-loading incubation, Li(+) accumulation at 30 min (mmoles Li(+)/L cells) was the greatest in lymphoblastomas (11.1+/-0.3), followed by neuroblastomas (9.3+/-0.5), and then erythrocytes (4.0+/-0.5). Li(+) binding affinities to the plasma membrane in all three cell types were of the same order of magnitude; however, Li(+) immobilization in intact cells was greatest in neuroblastomas and least in erythrocytes. When cells were loaded for 30 min in a 50 mM/L Li(+)-containing medium, the percentage increase in free intracellular [Mg(2+)] in neuroblastoma and lymphoblastoma cells ( approximately 55 and approximately 52%, respectively) was similar, but erythrocytes did not exhibit any substantial increase ( approximately 6%). With the intracellular [Li(+)] at 15 mM/L, the free intracellular [Mg(2+)] increased by the greatest amount in neuroblastomas ( approximately 158%), followed by lymphoblastomas ( approximately 75%), and then erythrocytes ( approximately 50%). We conclude that Li(+) immobilization and transport are related to free intracellular [Mg(2+)] and to the extent of Li(+)/Mg(2+) competition in a cell-specific manner.     

Characterization of organic anion transporting polypeptide 1b2 knockout rats generated by CRISPR/Cas9: a novel model for drug transport and hyperbilirubinemia disease

Organic anion transporting polypeptide 1B1 and 1B3 (OATP1B1/3) as important uptake transporters play a fundamental role in the transportation of exogenous drugs and endogenous substances into cells. Rat OATP1B2, encoded by the Slco1b2 gene, is homologous to human OATP1B1/3. Although OATP1B1/3 is very important, few animal models can be used to study its properties. In this report, we successfully constructed the Slco1b2 knockout (KO) rat model via using the CRISPR/Cas9 technology for the first time. The novel rat model showed the absence of OATP1B2 protein expression, with no off-target effects as well as compensatory regulation of other transporters. Further pharmacokinetic study of pitavastatin, a typical substrate of OATP1B2, confirmed the OATP1B2 function was absent. Since bilirubin and bile acids are the substrates of OATP1B2, the contents of total bilirubin, direct bilirubin, indirect bilirubin, and total bile acids in serum are significantly higher in Slco1b2 KO rats than the data of wild-type rats. These results are consistent with the symptoms caused by the absence of OATP1B1/3 in Rotor syndrome. Therefore, this rat model is not only a powerful tool for the study of OATP1B2-mediated drug transportation, but also a good disease model to study hyperbilirubinemia-related diseases.     

Delivery strategies of amphotericin B for invasive fungal infections

Invasive fungal infections (IFIs) represent a growing public concern for clinicians to manage in many medical settings, with substantial associated morbidities and mortalities. Among many current therapeutic options for the treatment of IFIs, amphotericin B (AmB) is the most frequently used drug. AmB is considered as a first-line drug in the clinic that has strong antifungal activity and less resistance. In this review, we summarized the most promising research efforts on nanocarriers for AmB delivery and highlighted their efficacy and safety for treating IFIs. We have also discussed the mechanism of actions of AmB, rationale for treating IFIs, and recent advances in formulating AmB for clinical use. Finally, this review discusses some practical considerations and provides recommendations for future studies in applying AmB for combating IFIs.     

The Impact of Product and Process Related Critical Quality Attributes on Immunogenicity and Adverse Immunological Effects of Biotherapeutics

The pharmaceutical industry has experienced great successes with protein therapeutics in the last two decades and with novel modalities, including cell therapies and gene therapies, more recently. Biotherapeutics are complex in structure and present challenges for discovery, development, regulatory, and life cycle management. Biotherapeutics can interact with the immune system that may lead to undesired immunological responses, including immunogenicity, hypersensitivity reactions (HSR), injection site reactions (ISR), and others. Many product and process related critical quality attributes (CQAs) have the potential to trigger or augment such immunological responses to the product. Tremendous efforts, both clinically and preclinically, have been invested to understand the impact of product and process related CQAs on adverse immunological effects. The information and knowledge are critical for the implementation of Quality by Design (QbD), which requires risk assessment and establishment of specifications and control strategies for CQAs. A quality target product profile (QTPP) that identifies the key CQAs through process development can help assign severity scores based on safety, immunogenicity, pharmacokinetics (PK) and pharmacodynamics (PD) of the molecule. Gaps and future directions related to biotherapeutics and emerging novel modalities are presented.