载药聚氰基丙烯酸丁酯纳米粒治疗脑胶质瘤的研究
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摘要
第一部分阿霉素聚氰基丙烯酸丁酯纳米粒的制备及载药研究
目的:制备阿霉素(doxorubicin,DOX)聚氰基丙烯酸丁酯( polybutylcyanoacrylate, PBCA)纳米粒(DOX-PBCA-NP),并对其进行测定。
方法:应用乳化聚合法制备DOX-PBCA-NP,用正交设计优选DOX-PBCA-NP的制备条件。纳米粒的粒径及分布用马尔文激光粒度分析仪测试,透射电镜(TEM)观察纳米粒形态、大小,紫外分光光度法在波长481nm处测定纳米粒中DOX的载药量和包封率。
结果:激光粒度分析仪测定纳米粒的平均粒径为120nm(载药)、100(空白),多分散系数(Polydispersity Index)为0.12。TEM下,纳米粒呈圆形,无粘连,纳米粒的载药量与包封率分别为10.58%与87.43%。
结论:应用乳化聚合的方法制备的纳米粒大小形态一致,性能稳定,可为进一步的体内和体外实验提供基础。
第二部分阿霉素聚氰基丙烯酸丁酯纳米粒抑制脑胶质瘤SHG44细胞的研究
目的:制备阿霉素(doxorubicin)聚氰基丙烯酸丁酯(polybutylcyanoacrylate)纳米粒(DOX-PBCA-NP),并研究其体外对脑胶质瘤SHG44的抑制作用。
方法:乳化聚合法制备DOX-PBCA-NP;MTT法测定细胞抑制率;流式细胞仪(flow cytometry,FCM)测定细胞凋亡和细胞周期的改变;倒置显微镜观察细胞生长情况;免疫组织化学法检测SHG44细胞端粒酶蛋白的表达。
结果:1、用药24h后,DOX的三个不同浓度组(按DOX浓度分别为2μg/ml,5μg/ml,10μg/ml)对SHG44细胞的抑制率分别为:(32.3±0.2)%、(48.7±0.7)%、(67.1±0.4)%;DOX-PBCA-NP组分别为:(45.7±0.2)%、(61.6±0.4)%、(76.2±0.8)%;PBCA-NP组为(1.6±0.7)%。2、在DOX,DOX-PBCA-NP三个不同浓度组(按DOX浓度分别为2μg/ml,5μg/ml,10μg/ml),DOX组细胞端粒酶蛋白阳性率分别为(86.4±3.7)%、75.6±2.1)%、(69.7±1.3)%,DOX-PBCA-NP组分别为(72.3±1.6)%、63.2±3.1)%、(55.2±0.8)%,各组端粒酶蛋白的表达阳性率均与对照组有显著性差异(P<0.01)。3、抑制效应表现为细胞周期改变明显,G1/Go期细胞增多,S期细胞明显减少,细胞凋亡增多,端粒酶蛋白表达明显下降。DOX组、DOX-PBCA-NP组与对照组比较均有显著性差异(P<0.01),DOX-PBCA-NP组较DOX组对SHG44细胞的抑制效应更明显,两者比较差异有统计学意义(P<0.05)。。
结论:阿霉素聚氰基丙烯酸丁酯纳米粒与阿霉素比较,对脑胶质瘤SHG44细胞的抑制作用明显增强。
第三部分端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒的制备及载药研究
目的:制备端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒(hTERT ASODN- PBCA-NP),并测定其粒径大小、形态、Zeta电位等。
方法:应用乳化聚合法制备hTERT ASODN-PBCA-NP,正交设计优选hTERT ASODN-PBCA-NP的制备条件。纳米粒的粒径及分布用马尔文激光粒度分析仪测试,透射电镜(TEM)观察纳米粒大小、形态,高效液相色谱法测定纳米粒中ASODN的载药量和包封率。
结果:激光粒度分析仪测定纳米粒的平均粒径为120nm,多分散系数(Polydispersity Index)为0.11。TEM下,纳米粒呈圆形,颗粒间无粘连,纳米粒的载药量与包封率分别为71.17%与95.20 %,Zeta为+41.3mV。。
结论:应用乳化聚合的方法制备的纳米粒大小形态一致,表面带正电荷,性能稳定,具有较高的载药量和包封率。
第四部分端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒抑制脑胶质瘤SHG44细胞的研究
目的:以聚氰基丙烯酸丁酯纳米粒(polybutylcyanoacrylate nanoparticles, PBCA-NP)为载体,以端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)反义寡核苷酸(antisense oligodeoxynucletide,ASODN)为目的基因体外转染脑胶质瘤SHG44细胞,研究其体外对SHG44细胞生长的抑制作用。
方法:应用乳化聚合法制备端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒(hTERT ASODN-PBCA-NP),琼脂糖凝胶电泳法分析其对抗核酸酶的降解作用,MTT法检测细胞的生长抑制率;倒置显微镜观察细胞的生长状况;流式细胞仪(flow cytometry,FCM)测定细胞周期的改变以及5'-FITC标记的ASODN(FASODN-NP)转染后的细胞内荧光强度;免疫细胞化学法测定SHG44细胞端粒酶蛋白的表达,RT-PCR检测hTERT mRNA的表达。
结果: 1、琼脂糖凝胶电泳结果表明ODN-PBCA-NP具有不被核酸酶降解的作用;转染后48h空白对照组、NP组、FASODN组和FASODN-NP组的胞内荧光道数分别为138.20,140.40,142.06和490.00,FASODN-NP组细胞内荧光强度增加,与其它各组相比有显著性差异(P<0.01);2、hTERT mRNA表达的相对值在ASODN1-3-NP各组分别为1.45±0.11、1.39±0.22、1.43±0.14,空白对照组为(2.23±0.12);3、端粒酶蛋白表达的阳性率在ASODN1-3各组分别为(53.1±1.8)%、(56.7±1.5)%和(59.6±4.3)%,空白对照组为(94.6±1.3)%;4、ASODN-NP组与空白对照组、SODN-NP组、ASODN组、NP组相比,SHG44细胞的抑制率、细胞周期的改变以及hTERT mRNA、端粒酶蛋白的表达均有显著性差异(P<0.05),ASODN-NP组细胞在G1/G0期增多,S期细胞减少,hTERT mRNA、端粒酶蛋白的表达下降。
结论: PBCA-NP为载体介导的hTERT-ASOND可以有效地透过细胞膜到达SHG44细胞内部,阻断hTERT mRNA的表达,抑制肿瘤细胞的生长。
第五部分端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒体内治疗裸鼠脑胶质瘤动物模型的研究
目的:以聚氰基丙烯酸丁酯纳米粒(polybutylcyanoacrylate nanoparticles, PBCA-NP)为载体,以端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)反义寡核苷酸(antisense oligodeoxynucletide,ASODN)为目的基因转染裸鼠脑胶质瘤动物模型,研究其体内对裸鼠脑胶质瘤动物模型的治疗作用。
方法:制备裸鼠脑胶质瘤动物模型,尾静脉注射端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒,测定裸鼠颅内肿瘤的大小;免疫组织化学法检测肿瘤组织端粒酶蛋白的表达、病理改变,RT-PCR检测hTERT mRNA的表达。
结果:1、肿瘤组织端粒酶蛋白的阳性表达率在空白对照组为(92.6±2.7)%,NP组为(91.2±3.4)%,SODN-NP组为(94.7±1.3)%,ASODN组为(90.5±2.6)%,ASODN-NP组为(65.2±2.1)%;2、空白细胞对照组、NP组、SODN-NP组、ASODN组肿瘤组织hTERT mRNA表达的相对值分别为2.34±0.15、2.46±0.22、2.41±0.35、2.39±0.24,ASODN-NP组为1.56±0.39;3、ASODN-NP组与对照组、SODN-NP组、ASODN组、NP组相比,肿瘤体积、端粒酶蛋白的表达以及hTERT mRNA表达均有显著性差异(P<0.05),ASODN-NP组肿瘤体积小,端粒酶蛋白、hTERT mRNA的表达下降。
结论: PBCA-NP为载体介导的hTERT-ASOND在体内具有抑制裸鼠脑胶质瘤生长的作用。
PartⅠPreparation and drug loading of doxorubicin–polybutylcyano -acrylate nanoparticles
Objective To prepare the doxorubicin–polybutylcyanoacrylate nanoparticles. Methods The DOX-PBCA-NP was prepared using the emulsion polymerization method under the optimal conditions of orthogonal design.The size and size distribution of nanoparticles were measured using Malven laser mastersizer 3000HS, nanoparticle morphology observed by transmission electron microsope.The Drud loading and entrapment efficiency of doxorubicin in the nanoparticles were measured by means of UV spectra at weave length of 481 nm.
Results The nanoparticles were discrete and uniform spheres with average diameter of 120nm and 100nm for bare and drug loaded nanoparticles respectively.
Conclusion Nanoparticles obtained by emulsion polymerization method were steady and the same size and morphology, which can be provided for the further in vitro and in vivo tests.
PartⅡInhibition of SHG44 cells by polybutylcyanoacrylate nanoparticles loaded with Doxorubicin
Objective To investigate the effect of polybutylcyanoacrylate nanoparticles loaded with Doxorubicin on the SHG44 cells.
Methods The polybutylcyanoacrylate nanoparticles loaded with Doxorubicin were prepared by emulsion polymerization process .The inhibitory rate of SHG44 cells were detected by MTT assay, cell modalities were observed using inverse microscope and the cell cycles determined by flow cytometry. The expression of telomerase protein was measured by immunocytochemical method.
Results 1. At three concentration levels of DOX, DOX-PBCA-NP, the inhibitory rate of SHG44 cells were (32.3±0.2)%、(48.7±0.7)%、(67.1±0.4)%% for DOX groups and(45.7±0.2)%、(61.6±0.4)%、(76.2±0.8)% for DOX-PBCA-NP groups, when the concentration of DOX were 2μg/ml, 5μg/ml and 10μg/ml for each group,with PBCA-NP group for (1.6±0.7)%.2. The telomerase protein expression were (86.4±3.7)%,75.6±2.1)% and(69.7±1.3)% for DOX groups, (72.3±1.6)%、63.2±3.1)% and(55.2±0.8)% for DOX-PBCA-NP groups , with the concentration of DOX were 2μg/ml, 5μg/ml and 10μg/ml for each group. 3. Compared with other groups, DOX-PBCA-NP groups displayed higher inhibitory rate, obviously cell modality change and cell cycle variety, reduced telomerase protein expression. Their differences are evidently remarkable(P<0.05).
Conclusion Polybutylcyanoacrylate nanoparticles loaded with doxorubicin were more effective on the SHG44 cells than doxorubicin as to cell inhibition .
PartⅢPreparation and drug loading of polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA
Objective To prepare the hTERT-ASODN-polybutylcyanoacrylate nanoparticles. Methods The hTERT ASODN-PBCA-NP was prepared using the emulsion polymerization method under the optimal conditions of orthogonal design.The size , size distribution and Zeta potential of nanoparticles were measured using Malven laser Mastersizer 3000HS, nanoparticle morphology observed by transmission electron microsope.The drud loading and entrapment efficiency of ASODN in the nanoparticles were measured by means of HPLC method at weave length of 260 nm.
Results The nanoparticles were discrete and uniform spheres with average diameter of 120nm PBCA nanoparticles, with drud loading and entrapment efficiency for 71.17% and 95.20%, and +41.3mV for Zeta potential.
Conclusion Nanoparticles obtained by emulsion polymerization method were steady and the same size and morphology,with higher drud loading and entrapment efficiency of oligodeoxynucletide in the nanoparticles, and a high positive Zeta potential.
PartⅣInhibition of SHG44 cells by polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA
Objective: To investigate the effect of polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA on SHG44 cells.
Methods: The polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA were prepared using the emulsion polymerization process .The inhibition rate of SHG44 cells were detected by MTT assay, cell modalities were observed using inverse microscope and the cell cycles and intracellular fluorescence intensity determined by flow cytometry after transfection . The expression of hTERT mRNA was measured using RT-PCR method, and expression telomerase protein was measured by immunocytochemical method. Results: Nanoparticles obtained by emulsion polymerizations process were the same size and morphology. Compared with FASODN group ( NP-free ) , the intracellular fluorescence intensity of FASODN-NP group was obviously stronger after transfection of 48h(P<0.01). The telomerase protein expression were (53.1±1.8)%, (56.7±1.5)% and(59.6±4.3)% for ASODN1-NP group, ASODN2-NP group and ASODN3-NP group, with control group for(94.6±1.3)%. The hTERT mRNA expression were 2.23±0.12、2.31±0.14、2.26±0.19 for ASODN1-NP group, ASODN2-NP group and ASODN3-NP group, with control group for 1.45±0.11.The ASODN-NP groups displayed higher inhibitory rate, obviously cell modality change and cell cycle variety, reduced hTERT mRNA and telomerase protein expression than that of ASODN groups(NP-free), SODN group , NP group and control group, their differences are evidently remarkable.
Conclusion: Polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA can enter the inside of SHG44 cells, preventing the expression of hTERT mRNA, and inhibit cell proliferation .
PartⅤChemotheraphy of glioma nude mice using polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA
Objective: To investigate the chemotheraphic effect of polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA on SHG44 cell glioma nude mice.
Methods: The polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA were prepared by emulsion polymerization process . The glioma nude mice models were established by means of tumor tissue pieces of SHG44 cells inoculated into BALB/c-nu nude mice. The preparation of drugs were injected i.v. into the tail vein.The expression of hTERT mRNA was measured using RT-PCR method,the telomerase protein and the pathology of the tumor tissue were measured by immunohistochemical method.
Results: The glioma models in nude mice were successfully established by means of tumor tissue pieces inoculation.The telomerase protein expression were (92.6±2.7)%,(91.2±3.4)%,(94.7±1.3)%,(90.5±2.6)%,(65.2±2.1)% for control group, NP group, SODN-NP group, ASODN group and ASODN-NP group, and the hTERT mRNA expression were 2.34±0.15, 2.46±0.22, 2.41±0.35, 2.39±0.24, 1.56±0.39 respectively . Compared with other groups , the ASODN-NP group displayed reduced hTERT mRNA and telomerase protein expression,smaller tumor size, their differences are evidently remarkable(P<0.05). The above differences between control group, NP group, SODN-NP group and ASODN group were no statistical meaning.
Conclusion: Polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA were chemotheraphic effective to SHG44 glioma nude mice .
目的:制备阿霉素(doxorubicin,DOX)聚氰基丙烯酸丁酯( polybutylcyanoacrylate, PBCA)纳米粒(DOX-PBCA-NP),并对其进行测定。
方法:应用乳化聚合法制备DOX-PBCA-NP,用正交设计优选DOX-PBCA-NP的制备条件。纳米粒的粒径及分布用马尔文激光粒度分析仪测试,透射电镜(TEM)观察纳米粒形态、大小,紫外分光光度法在波长481nm处测定纳米粒中DOX的载药量和包封率。
结果:激光粒度分析仪测定纳米粒的平均粒径为120nm(载药)、100(空白),多分散系数(Polydispersity Index)为0.12。TEM下,纳米粒呈圆形,无粘连,纳米粒的载药量与包封率分别为10.58%与87.43%。
结论:应用乳化聚合的方法制备的纳米粒大小形态一致,性能稳定,可为进一步的体内和体外实验提供基础。
第二部分阿霉素聚氰基丙烯酸丁酯纳米粒抑制脑胶质瘤SHG44细胞的研究
目的:制备阿霉素(doxorubicin)聚氰基丙烯酸丁酯(polybutylcyanoacrylate)纳米粒(DOX-PBCA-NP),并研究其体外对脑胶质瘤SHG44的抑制作用。
方法:乳化聚合法制备DOX-PBCA-NP;MTT法测定细胞抑制率;流式细胞仪(flow cytometry,FCM)测定细胞凋亡和细胞周期的改变;倒置显微镜观察细胞生长情况;免疫组织化学法检测SHG44细胞端粒酶蛋白的表达。
结果:1、用药24h后,DOX的三个不同浓度组(按DOX浓度分别为2μg/ml,5μg/ml,10μg/ml)对SHG44细胞的抑制率分别为:(32.3±0.2)%、(48.7±0.7)%、(67.1±0.4)%;DOX-PBCA-NP组分别为:(45.7±0.2)%、(61.6±0.4)%、(76.2±0.8)%;PBCA-NP组为(1.6±0.7)%。2、在DOX,DOX-PBCA-NP三个不同浓度组(按DOX浓度分别为2μg/ml,5μg/ml,10μg/ml),DOX组细胞端粒酶蛋白阳性率分别为(86.4±3.7)%、75.6±2.1)%、(69.7±1.3)%,DOX-PBCA-NP组分别为(72.3±1.6)%、63.2±3.1)%、(55.2±0.8)%,各组端粒酶蛋白的表达阳性率均与对照组有显著性差异(P<0.01)。3、抑制效应表现为细胞周期改变明显,G1/Go期细胞增多,S期细胞明显减少,细胞凋亡增多,端粒酶蛋白表达明显下降。DOX组、DOX-PBCA-NP组与对照组比较均有显著性差异(P<0.01),DOX-PBCA-NP组较DOX组对SHG44细胞的抑制效应更明显,两者比较差异有统计学意义(P<0.05)。。
结论:阿霉素聚氰基丙烯酸丁酯纳米粒与阿霉素比较,对脑胶质瘤SHG44细胞的抑制作用明显增强。
第三部分端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒的制备及载药研究
目的:制备端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒(hTERT ASODN- PBCA-NP),并测定其粒径大小、形态、Zeta电位等。
方法:应用乳化聚合法制备hTERT ASODN-PBCA-NP,正交设计优选hTERT ASODN-PBCA-NP的制备条件。纳米粒的粒径及分布用马尔文激光粒度分析仪测试,透射电镜(TEM)观察纳米粒大小、形态,高效液相色谱法测定纳米粒中ASODN的载药量和包封率。
结果:激光粒度分析仪测定纳米粒的平均粒径为120nm,多分散系数(Polydispersity Index)为0.11。TEM下,纳米粒呈圆形,颗粒间无粘连,纳米粒的载药量与包封率分别为71.17%与95.20 %,Zeta为+41.3mV。。
结论:应用乳化聚合的方法制备的纳米粒大小形态一致,表面带正电荷,性能稳定,具有较高的载药量和包封率。
第四部分端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒抑制脑胶质瘤SHG44细胞的研究
目的:以聚氰基丙烯酸丁酯纳米粒(polybutylcyanoacrylate nanoparticles, PBCA-NP)为载体,以端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)反义寡核苷酸(antisense oligodeoxynucletide,ASODN)为目的基因体外转染脑胶质瘤SHG44细胞,研究其体外对SHG44细胞生长的抑制作用。
方法:应用乳化聚合法制备端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒(hTERT ASODN-PBCA-NP),琼脂糖凝胶电泳法分析其对抗核酸酶的降解作用,MTT法检测细胞的生长抑制率;倒置显微镜观察细胞的生长状况;流式细胞仪(flow cytometry,FCM)测定细胞周期的改变以及5'-FITC标记的ASODN(FASODN-NP)转染后的细胞内荧光强度;免疫细胞化学法测定SHG44细胞端粒酶蛋白的表达,RT-PCR检测hTERT mRNA的表达。
结果: 1、琼脂糖凝胶电泳结果表明ODN-PBCA-NP具有不被核酸酶降解的作用;转染后48h空白对照组、NP组、FASODN组和FASODN-NP组的胞内荧光道数分别为138.20,140.40,142.06和490.00,FASODN-NP组细胞内荧光强度增加,与其它各组相比有显著性差异(P<0.01);2、hTERT mRNA表达的相对值在ASODN1-3-NP各组分别为1.45±0.11、1.39±0.22、1.43±0.14,空白对照组为(2.23±0.12);3、端粒酶蛋白表达的阳性率在ASODN1-3各组分别为(53.1±1.8)%、(56.7±1.5)%和(59.6±4.3)%,空白对照组为(94.6±1.3)%;4、ASODN-NP组与空白对照组、SODN-NP组、ASODN组、NP组相比,SHG44细胞的抑制率、细胞周期的改变以及hTERT mRNA、端粒酶蛋白的表达均有显著性差异(P<0.05),ASODN-NP组细胞在G1/G0期增多,S期细胞减少,hTERT mRNA、端粒酶蛋白的表达下降。
结论: PBCA-NP为载体介导的hTERT-ASOND可以有效地透过细胞膜到达SHG44细胞内部,阻断hTERT mRNA的表达,抑制肿瘤细胞的生长。
第五部分端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒体内治疗裸鼠脑胶质瘤动物模型的研究
目的:以聚氰基丙烯酸丁酯纳米粒(polybutylcyanoacrylate nanoparticles, PBCA-NP)为载体,以端粒酶逆转录酶(human telomerase reverse transcriptase,hTERT)反义寡核苷酸(antisense oligodeoxynucletide,ASODN)为目的基因转染裸鼠脑胶质瘤动物模型,研究其体内对裸鼠脑胶质瘤动物模型的治疗作用。
方法:制备裸鼠脑胶质瘤动物模型,尾静脉注射端粒酶反义寡核苷酸聚氰基丙烯酸丁酯纳米粒,测定裸鼠颅内肿瘤的大小;免疫组织化学法检测肿瘤组织端粒酶蛋白的表达、病理改变,RT-PCR检测hTERT mRNA的表达。
结果:1、肿瘤组织端粒酶蛋白的阳性表达率在空白对照组为(92.6±2.7)%,NP组为(91.2±3.4)%,SODN-NP组为(94.7±1.3)%,ASODN组为(90.5±2.6)%,ASODN-NP组为(65.2±2.1)%;2、空白细胞对照组、NP组、SODN-NP组、ASODN组肿瘤组织hTERT mRNA表达的相对值分别为2.34±0.15、2.46±0.22、2.41±0.35、2.39±0.24,ASODN-NP组为1.56±0.39;3、ASODN-NP组与对照组、SODN-NP组、ASODN组、NP组相比,肿瘤体积、端粒酶蛋白的表达以及hTERT mRNA表达均有显著性差异(P<0.05),ASODN-NP组肿瘤体积小,端粒酶蛋白、hTERT mRNA的表达下降。
结论: PBCA-NP为载体介导的hTERT-ASOND在体内具有抑制裸鼠脑胶质瘤生长的作用。
PartⅠPreparation and drug loading of doxorubicin–polybutylcyano -acrylate nanoparticles
Objective To prepare the doxorubicin–polybutylcyanoacrylate nanoparticles. Methods The DOX-PBCA-NP was prepared using the emulsion polymerization method under the optimal conditions of orthogonal design.The size and size distribution of nanoparticles were measured using Malven laser mastersizer 3000HS, nanoparticle morphology observed by transmission electron microsope.The Drud loading and entrapment efficiency of doxorubicin in the nanoparticles were measured by means of UV spectra at weave length of 481 nm.
Results The nanoparticles were discrete and uniform spheres with average diameter of 120nm and 100nm for bare and drug loaded nanoparticles respectively.
Conclusion Nanoparticles obtained by emulsion polymerization method were steady and the same size and morphology, which can be provided for the further in vitro and in vivo tests.
PartⅡInhibition of SHG44 cells by polybutylcyanoacrylate nanoparticles loaded with Doxorubicin
Objective To investigate the effect of polybutylcyanoacrylate nanoparticles loaded with Doxorubicin on the SHG44 cells.
Methods The polybutylcyanoacrylate nanoparticles loaded with Doxorubicin were prepared by emulsion polymerization process .The inhibitory rate of SHG44 cells were detected by MTT assay, cell modalities were observed using inverse microscope and the cell cycles determined by flow cytometry. The expression of telomerase protein was measured by immunocytochemical method.
Results 1. At three concentration levels of DOX, DOX-PBCA-NP, the inhibitory rate of SHG44 cells were (32.3±0.2)%、(48.7±0.7)%、(67.1±0.4)%% for DOX groups and(45.7±0.2)%、(61.6±0.4)%、(76.2±0.8)% for DOX-PBCA-NP groups, when the concentration of DOX were 2μg/ml, 5μg/ml and 10μg/ml for each group,with PBCA-NP group for (1.6±0.7)%.2. The telomerase protein expression were (86.4±3.7)%,75.6±2.1)% and(69.7±1.3)% for DOX groups, (72.3±1.6)%、63.2±3.1)% and(55.2±0.8)% for DOX-PBCA-NP groups , with the concentration of DOX were 2μg/ml, 5μg/ml and 10μg/ml for each group. 3. Compared with other groups, DOX-PBCA-NP groups displayed higher inhibitory rate, obviously cell modality change and cell cycle variety, reduced telomerase protein expression. Their differences are evidently remarkable(P<0.05).
Conclusion Polybutylcyanoacrylate nanoparticles loaded with doxorubicin were more effective on the SHG44 cells than doxorubicin as to cell inhibition .
PartⅢPreparation and drug loading of polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA
Objective To prepare the hTERT-ASODN-polybutylcyanoacrylate nanoparticles. Methods The hTERT ASODN-PBCA-NP was prepared using the emulsion polymerization method under the optimal conditions of orthogonal design.The size , size distribution and Zeta potential of nanoparticles were measured using Malven laser Mastersizer 3000HS, nanoparticle morphology observed by transmission electron microsope.The drud loading and entrapment efficiency of ASODN in the nanoparticles were measured by means of HPLC method at weave length of 260 nm.
Results The nanoparticles were discrete and uniform spheres with average diameter of 120nm PBCA nanoparticles, with drud loading and entrapment efficiency for 71.17% and 95.20%, and +41.3mV for Zeta potential.
Conclusion Nanoparticles obtained by emulsion polymerization method were steady and the same size and morphology,with higher drud loading and entrapment efficiency of oligodeoxynucletide in the nanoparticles, and a high positive Zeta potential.
PartⅣInhibition of SHG44 cells by polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA
Objective: To investigate the effect of polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA on SHG44 cells.
Methods: The polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA were prepared using the emulsion polymerization process .The inhibition rate of SHG44 cells were detected by MTT assay, cell modalities were observed using inverse microscope and the cell cycles and intracellular fluorescence intensity determined by flow cytometry after transfection . The expression of hTERT mRNA was measured using RT-PCR method, and expression telomerase protein was measured by immunocytochemical method. Results: Nanoparticles obtained by emulsion polymerizations process were the same size and morphology. Compared with FASODN group ( NP-free ) , the intracellular fluorescence intensity of FASODN-NP group was obviously stronger after transfection of 48h(P<0.01). The telomerase protein expression were (53.1±1.8)%, (56.7±1.5)% and(59.6±4.3)% for ASODN1-NP group, ASODN2-NP group and ASODN3-NP group, with control group for(94.6±1.3)%. The hTERT mRNA expression were 2.23±0.12、2.31±0.14、2.26±0.19 for ASODN1-NP group, ASODN2-NP group and ASODN3-NP group, with control group for 1.45±0.11.The ASODN-NP groups displayed higher inhibitory rate, obviously cell modality change and cell cycle variety, reduced hTERT mRNA and telomerase protein expression than that of ASODN groups(NP-free), SODN group , NP group and control group, their differences are evidently remarkable.
Conclusion: Polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA can enter the inside of SHG44 cells, preventing the expression of hTERT mRNA, and inhibit cell proliferation .
PartⅤChemotheraphy of glioma nude mice using polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA
Objective: To investigate the chemotheraphic effect of polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA on SHG44 cell glioma nude mice.
Methods: The polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA were prepared by emulsion polymerization process . The glioma nude mice models were established by means of tumor tissue pieces of SHG44 cells inoculated into BALB/c-nu nude mice. The preparation of drugs were injected i.v. into the tail vein.The expression of hTERT mRNA was measured using RT-PCR method,the telomerase protein and the pathology of the tumor tissue were measured by immunohistochemical method.
Results: The glioma models in nude mice were successfully established by means of tumor tissue pieces inoculation.The telomerase protein expression were (92.6±2.7)%,(91.2±3.4)%,(94.7±1.3)%,(90.5±2.6)%,(65.2±2.1)% for control group, NP group, SODN-NP group, ASODN group and ASODN-NP group, and the hTERT mRNA expression were 2.34±0.15, 2.46±0.22, 2.41±0.35, 2.39±0.24, 1.56±0.39 respectively . Compared with other groups , the ASODN-NP group displayed reduced hTERT mRNA and telomerase protein expression,smaller tumor size, their differences are evidently remarkable(P<0.05). The above differences between control group, NP group, SODN-NP group and ASODN group were no statistical meaning.
Conclusion: Polybutylcyanoacrylate nanoparticles loaded with antisense oligodeoxynucleotide of hTERT mRNA were chemotheraphic effective to SHG44 glioma nude mice .
引文
[1] Colin W,Pouton,Leonard W Seymour.Key issues in non-viral gene delivery [J].Adv Drug Delve Rev.1998,34(1):3-19.
[2] Cristiano,Richard J.Targeted,non-viral gene delivery for cancer gene therapy [J].Front Biosci.1998,3:D1161-D1170.
[3] SONG CX,LABHASETWAR V,MURPHY H,etal.Formulation and characterization of biodegradable nanoparticales for intravascular local drug delivery[J]. J Contr Rel.1997,43(3):197-201.
[4] Cleland L. Solvent evaporation processes for the production of controlled release biodegradable microsphere formulations for therapeutics and vaccines[J]. Biotechnology Progerss.1998,14:102-107.
[5] Song C, Labhasetwar V,Levy J.Controlled release of U-86983 from double-layer biodegradable matrices: effect of additives on release mechanism and kinetics[J].J Controlled Release.1997,45:177-182.
[6] LUBBE AS, BERGEMANN C, RIESS H,etal.Clincal experiecses with magnetic drug targeting: a phase I study with 4'-epidoxorubicin in 14 patients with advanced aolid tumors [J].Cancer Res,1996,56(20):4686.
[7] GREIDER CW. Telomerase activity,cell proliferation and cancer[J]. Proc Natl Acad Sci USA,1998,95:90.
[8] OLIVIER Z, FRANCIS C, SZOKA J. Intracellular distribution and mechanism of delivery of oligonucleotides mediated by cationic lipid [J]. PharmRes. 1996,13(9): 1367-1372.
[9] Lambert G,Fattal E,Couvreur P,et al.Nanoparticulate systems for the delivery of Antisense oligonucleotides[J].Adv Drug Deliv Rev.2001,47(1):99-112.
[10] Cohen H,Levy RJ, Gao J, et al. Sustained delivery and expression of DNA encapsulated in polymeric nanoparticles [J].Gene Therapy.2000,7:1896-1905.
[11] Labhasetwar V et al. A DNA controlled-release coating for gene transfer: transfection in skeletal and cardiac mulscle[J].J PHarm Sci.1998,87:1347-1350.
[12] Leong KW, et a1.DNA-polycation nanospheres as non-virus gene delivery vehicles[J]. J Control Release. 1998,53:183-193.
[13]De Verdiere AC, Dubernet C, Nemati F, et al. Revertion of muitidrug resistance with polyalkylcyanoacrylate nanoparticles: towards a mechanism of action[J]. British J Cancer,1997,76(2):198-205.
[14]Gonzalez-Martin G, Isabel Merino M, Rodriguez-Cabezas N,et al.Characterization and trypanocidal activity of Nifurtimox-containing and empty nanoparticles of polyethylcyanoacrylate[J].J PHarm PHarmacol,1998,50(1):29-35.
[1] Stan AC, Casares S, Radu D, Walter GF, Brumeanu TD. Doxorubicin induced cell death in highly invasive human gliomas. Anticancer Res. 1999;19:941-50.
[2] Walter KA,Tamargo RJ, Qlivi A, Bueger PC,Brem H. Intratumoral chemotheraphy.Neurosurgery .1995,37:1128-45.
[3] von Holst H, Knochenhauer E, Blomgren H, Collins VP, Ehn L, Lindquist M, Noren G, Peterson C. Uptake of adriamycin in tumour and surrounding brain tissue in patients with malignant gliomas. Acta Neurochir 1990;104:13-16.
[4] Steiniger SC, Kreuter J, Khalansky AS, Skidan IN, Bobruskin AI, etal. Chemotherapy of glioblastoma in rats using doxorubicin-loaded nanoparticles. Int J Cancer. 2004,109:759-767.
[5]M.Simeonova,R.Velichkova,G.Ivanova,etal.Poly(butyalcyanoacrylate)nanoparticles for topical delivery of 5-fluorouacil.Int.J.Pharm. 2003,263:133-140.
[6]Alyaudtin.R,Reichel.A,L?benberg.R,etal.Internation of poly(butyalcyanoacrylate) nanoparticles with the Blood-Brain Barrier in vivo and in vitro.Journal of Drug Targeting.No.3,Vol.9:209-221.
[7]杨西晓,陈建海,郭丹.聚氰基丙烯酸正丁酯纳米粒的生物相容性研究[J].第一军医大学学报,2005,25(10):1261-1262.
[8] S. I. Park, J. H. Lim, Y. H. Hwang etal. In vivo and in vitro antitumor activity of doxorubicin-loaded magnetic fluids. phys. stat. sol. (c) 4, No. 12, 4345-4351 (2007).
[9] J?rg Kreuter,Peter Ramge,Valery Petrovetal. Direct Evidence that Polysorbate-80- Coated Poly(Butylcyanoacrylate) Nanoparticles Deliver Drugs to the CNS via Specific Mechanisms Requiring Prior Binding of Drug to the Nanoparticles. Pharmaceutical Research,2003 , No. 3,Vol. 20:409-416.
[10]Dunn SE, Coombes AGA, Garnett MC, Davis SS, Davies MC, Illum L. In vitro cell interaction and in vivo biodistribution of poly(lactide-co-glycolide) nanospheres surface modified by poloxamer and poloxamine copolymers. J Controlled Release 1997;44:65–76.
[11]Lourenco C, Teixeira M, Simoes S, Gaspar R. Steric stabilization of nanoparticles: Size and surface properties. Int J Pharm1996;138:1–12.
[12] Coombes AGA, Yeh MK, Lavelle EC, Davis SS. The control of protein release from poly(DL-lactide-co-glycolide) microparticles by variation of the external aqueousphase surfactant in the water-in oil-in-water method. J Controlled Release. 1998;52:311–320.
[13] Quintanar–Guerrero D, Ganem–Quintanar A, Allemann E, etal. Influence of the stabilizer coating layer on the purification and freeze-drying of poly(D,L-lactic acid) nanoparticles prepared by an emulsion-diffusion technique. J Microencapsulation 1998;15:107-119.
[14]Bouillot P, Babak V, Dellacherie E. Novel bioresorbable and bioeliminable surfactants for microsphere preparation. Pharm Res, 1999,16:148–154.
[1] Stan AC, Casares S, Radu D,Walter GF, Brumeanu TD. Doxorubicin induced cell death in highly invasive human gliomas. Anticancer Res ,1999,19:941-950.
[2] Brigger I,Dubernet C,Courveur P. Nanoparticles in cancer therapy and diagnosis. Advdrug Del Rev, 2002,54:631-651.
[3] Ferrari M.Cancer nanotechnology.Opportunities and challenges. Nat Rev Cancer, 2005, 5:161-171..
[4] Hillyer J,Albrecht R.Gastrointestinal persorption and tissue distribution of differently sized colloidal gold nanoparticles.J Pharm Sci, 2001,90:1927-1936.
[5] Kayser O,Lemke A,Hernandez-Trejo N.The impact of nanobiotechnology on the development of new drug delivery systems.Curr Pharm Biotechnol, 2005,6:3-5..
[6] Kreuter J.Nanoparticulate systems for the brain delivery of drugs. Advdrug Deliv Rev, 2001, 47:65-81.
[7] Kreuter J,Ramge P,Petrov V,etal.Direct evidence that polysorbate-80-coated (Polybutylcyanoacrylate) nanoparticles deliver drugs to the CNS via specific mechanisms requiring prior binding of drugs to the nanoparticles. Pharm Res, 2003,20 :409-416.
[8] Vauthier C, Dubernet C, Fattal E,etal. Poly(butylcyanoacrylate ) as biodegradable materials for biomedical applications. Advdrug Deliv Rev, 2003,55:519-548.
[9]杨西晓,陈建海,郭丹.聚氰基丙烯酸正丁酯纳米粒的生物相容性研究[J].第一军医大学学报,2005,25(10):1261-1262.
[10] 1. R. Alyautdin, D. Gothier, V. Petrov, D. Kharkevich, and J.Kreuter. Analgesic activity of the hexapeptide dalargin adsorbed on the surface of polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles. Eur. J. Pharm. Biopharm, 1995 ,41:44-48.
[11] R. N. Alyautdin, V. E. Petrov, K. Langer, etal. Kreuter. Delivery of loperamide across the blood–brain barrier with poly-sorbate 80-coated polybutylcyanoacrylate nanoparticles. Pharm. Res, 1997,14:325-328.
[12] R. N. Alyautdin, E. B. Tezikov, P. Ramge, etal. Significant entry of tubocurarine into the brain of rats by absorption to polysorbate 80-coated polybutyl-cyanoacrylate nanoparticles: an in situ brain perfusion study.J. Microencapsul, 1998, 15:67-74 .
[13] A. E. Gulyaev, S. E. Gelperina, I. N. Skidan,etal. Kreuter. Significant transport of doxorubicin into the brain with polysorbate 80-coated nanoparticles. Pharm.Res, 1999,16:1564-1569.
[14] A. Friese, E. Seiler, G. Quack, B. Lorenz, and J. Kreuter. Enhancement of the duration of the anticonvulsive activity of a novel NMDA receptor antagonist using poly(butylcyanoacylate) nanoparticles as a parenteral controlled release delivery system.Eur. J. Pharm. Biopharm, 2000, 49:103-109.
[15] Sebastian C.J. Steiniger, Jorg Kreuter, Alexander S,etal. Chemotherphy of glioblastoma in rats using doxorubicin-loaded nanoparticles. Cancer, 2004,109: 759-767.
[16] Mahaley MS Jr. Immunological considerations and the malignant gliomas problem. Clin Neurosurg, 1968,15:175-189.
[17] Giometto B, Bozza F, Faresin F, etal.Immune infiltrates and cytokines in gliomas.Acta Neurochir, 1996,138:50-56.
[18] S, Mehraein P, Graeber MB. Differential expression of MHC class II molecules by microglia and neoplastic astroglia: relevance for the escape of astrocytoma cells from immune surveillance. Neuropathol Appl Neurobiol,1998,24:293-230.
[19] Aldskogius H, Liu L, Svensson M. Glial responses to synaptic damage and plasticity. J Neurosci Res,1999,58:33-41.
[20] Badie B, Schartner J. Role of microglia in glioma biology. Microsc Res Tech, 2001,54:106 -113.
[21] Boiardi A, Pozzi A, Salmaggi A, etal.Safety and potential effectiveness of daunorubicin-containing liposomesin patients with advanced recurrent malignant CNS tumors.Cancer Chemother Pharmacol, 1999,43:178-179.
[22] Lippens RJ. Liposomal daunorubicin (DaunoXome) in children with recurrent orprogressive brain tumors. Pediatr Hematol Oncol, 1999,16:131-139.
[23] Koukourakis MI, Koukouraki S, Fezoulidis I, etal. High intratumoural accumulation of stealth liposomal doxorubicin (Caelyx) in glioblastomas and in metastatic brain tumours. Br J Cancer, 2000,83:1281-1286.
[24] Gelperina SE, Khalansky AS, Skidan IN, etal.Toxicological studies of doxorubicin bound to polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles in healthy rats and rats with intracranial glioblastoma.Toxicol Lett,2002,126:131-141.
[25] Neuwelt EA, Pagel M, Barnett P, etal. Pharmacology and toxicity of intracarotid adriamycin administration following osmotic blood-brain barrier modification. Cancer Res,1981,41:4466-4470.
[1] HANEY C B,FUTCHER C W,GREIDER C W.Telomeres shorten during of human fibroblasts[J].Natttre,1990,245:458-460.
[2] Makarov VL, Hirose Y, Langmore JP: Long G tails at both ends of human chromosomes suggest a C strand degradationmechanism for telomere shortening. Cell 1997, 88:657-666.
[3] Wright WE, Tesmer VM, Huffman KE, Levene SD, Shay JW: Normal human chromosomes have long G-rich telomeric overhangs at one end. Genes Dev, 1997, 11:2801-2809.
[4] Colgin LM, Reddel RR: Telomere maintenance mechanisms and cellular immortalization. Curr Opin Genet Dev,1999, 9:97-103.
[5] Fattal E, Vauthier C, Ayine I, et al. Biadegadable polyalkylcyanoacrylate nano- particles for the delivery of oligonucleotides[J]. J Controlled Release,1998,53:137.
[1] Bennett WP, Hollstein MC, Hsu IC,etal.Mutational spectra and immunohistochemical analysis of p53 in human cancer [J]. Chest,1992,101:19-20.
[2]杜子威,徐庚达,王尧等.人脑恶性胶质瘤体外细胞系的建立及其特征.中华肿瘤杂志.1984;6(4):241-245.
[3] W. J. Bowers, D. F. Howard, and H. J. Federoff. Gene therapeutic strategies for neuroprotection: implications for Parkinson,s disease. Exp. Neurol, 1997,144:58–68.
[4] B. Blits, and M. B. Bunge. Direct gene therapy for repair of the spinal cord. J. Neurotrauma, 2006,23:508-520.
[5] J. M. Alisky, and B. L. Davidson. Gene therapy for amyotrophic lateral sclerosis and other motor neuron diseases. Hum.Gene. Ther, 2000, 11:2315-2329.
[6] R. Tinsley, and P. Eriksson. Use of gene therapy in central nervous system repair. Acta. Neurol. Scand, 2004,109:1-8.
[7] T. Federici, and N. M. Boulis. Gene-based treatment of motor neuron diseases. Muscle Nerve, 2006,33:302-323.
[8] B. K. Kaspar, J. Llado, N. Sherkat, etal. Retrograde viral delivery of IGF-1 prolongs survival in amouse ALS model. Science, 2003, 301:839-842.
[9] A. P. Kells, D. M. Fong, M. Dragunow,etal. AAV-mediated gene delivery of BDNF or GDNF is neuroprotective in a model of Huntington disease. Mol. Ther, 2004,9:682-688.
[10] R. J. Mandel, S. K. Spratt, R. O. Snyder,etal.Midbrain injection of recombinant adeno-associated virus encoding rat glial cell line-derived neurotrophic factor protects nigral neurons in a progressive 6-hydroxydopamine-induced degeneration model of Parkinson,s disease in rats. Proc. Natl.Acad. Sci. U S A, 1997, 94:14083-14088.
[11] D. L. Choi-Lundberg, Q. Lin, T. Schallert, etal.C. Bohn. Behavioral and cellular protection of rat dopaminergic neurons by an adenoviral vector encoding glial cell line-derived neurotrophic factor. Exp. Neurol, 1998, 154:261-275.
[12] J. A. Gonzalez-Barrios, M. Lindahl, M. J. Bannon, etal. Neurotensin polyplex as an efficient carrier for delivering the human GDNF gene into nigral dopamine neurons of hemiparkinsonian rats.Mol. Ther, 2006(14):857-865.
[13] Y. Zhang, F. Schlachetzki, Y. F. Zhang,etal. Normalization of striatal tyrosine hydroxylase and reversal of motor impairment in experimental parkinsonism with intravenous nonviral gene therapy and a brain-specific promoter. Hum. Gene. Ther, 2004(15):339-350.
[14] M. J.During, J. R. Naegele,K.L.O .Malley, etal. Longterm behavioral recovery in parkinsonian rats by an HSV vector expressing tyrosine hydroxylase. Science, 1994,266:1399-1403.
[15] C. S. Hong, W. F. Goins, J. R. Goss, etal. Herpes simplex virus RNAi and neprilysin gene transfer vectors reduce accumulation of Alzheimer,s diseaserelated amyloid-beta peptide in vivo. Gene. Ther, 2006(13):1068-1079.
[16] Cho-Chung YS. Antiense DNAs as a targeted therapeutics for cancer : no longer a dream[J]. Curr Opin Investig Drugs,2002(3):934-939.
[17] D. J. Begley. Delivery of therapeutic agents to the central nervous system: theproblems and the possibilities. Pharmacol.Ther, 2004,104:29-45.
[18] R. J. Boado. RNA interference and nonviral targeted gene therapy of experimental brain cancer. NeuroRx, 2005, 2:139-150.
[19] W. M. Pardridge. Tyrosine hydroxylase replacement in experimental Parkinson_s disease with transvascular gene therapy.NeuroRx, 2005,2:129-138.
[20] W.M.Pardridge.Blood-brain barrier delivery. Drug Discov.Today, 2007,12:54-61.
[21] Chavany C,Sasion-Behmoaras T,le Don T,etal.Adsorption of oligonucleotides onto polyisohexylcyanoacrylate nanoparticles protects them against nuclease and increase their cellular uptake[J].Pharm Res,1994,11(9):1370.
[22] Andrei Maksimenko,Claude Malvy, Gregory Lambert, etal.Oligonucleotides Targeted Against a Junction Oncogene Are Made Efficient by Nanotechnologies. Pharmaceutical Research, 2003,10(20):1565-1567.
[1] Konmta T,Kanzawa T, Kondo Y ,et a1. Telomerase as a therapeutic target for malignant gliomas[J]. Oncogene,2002,21(4):656-663.
[2] Neidle S, Kellan d LR.Telomerase as an anti-cancer target:current status and future prospects[J].Anticancer Drug Des,1999,14(4):341-347.
[3] Singh A,Sharma H,Salhan S,et a1.Evaluation of expression of apoptosis-related proteins and their correlation with HPV,telomerase activity,and apoptotic index in cervical cancer[J].Pathobiology,2004,71(6):314-322.
[4] Iida A ,Yamaguchi A, Hirose K. Telomerase activity in colorectal cancer and itsrelationship to bel-2 expresion[J].J Surg Oncol,2000,73(4):219-223.
[5] Neidle S,Parkinson G.Telomere maintenan ce as a target for anticancer drug discovery[J].Nat Rev Drug Discov,2002,l(5):383-393.
[6] Kondo S,Tanaka Y,Kondo Y,et a1.Antisense telomerase treatment:induction of two distinct pathways,apoptosis and differentiation[J].FASEB J,1998,12(10):801-811.
[7]罗元辉,房殿存,畅仕明,等.端粒酶在原发肝癌中的表达及临床意义[J].中华肝脏病杂志,1998,6(3):133-135.
[8]许兰涛.端粒酶反义寡核苷酸抗肿瘤的实验研究[J].中华肝脏病杂志,2001, 9(2):80.
[9] Kondo Y, Koga S, Komata T, et al. Treatment of prostate cancer in vitro and vivo w ith 2-5A–anti-telomerase RNA component.Oncogene, 2000, 19:2205-2211.
[10] M ukai S, Kondo Y, Koga S, et al. 2-5A antisense telomerase RNA therapy for intracranial malignant gliomas. Cancer Res,2000, 60:4461-4467.
[1] Couvreur P,Kante B,Roland M,etal.Poly(alkylcyanoacrylate) nanoparticles as potential lysosomotropic carriers: preparation,morphological and sorptive properties.J Pharm Pharmacol,1979,31:331-332.
[2] Kreuter,J. Evaluation of nanoparticles as drug-delivery systems. I. preparations methods. Pharm.Acta Helv,1983,58,196-209.
[3] D. J. Begley, M. W. Bradbury, and J. Kreuter (eds.). The Blood-Brain Barrier and Drug Delivery to the CNS, Marcel DekkerNew York, 2000.
[4] Alyautdin,R.N,Gothier D,Petro V,Kharkevich D, etal.Analgesic activity of the hexapeptide dalargin on the surface of polysorbate 80-coated polybutylcyanoacrylate nanoparticles. Eur J.Biopharm,1998,41:44-48.
[5] Gulyaev AE, Gelperina SE, Skidan IN, Antropov AS, Kivman GY, Kreuter J. Significant transport of doxorubicin into the brain with polysorbate 80-coated nanoparticles. Pharm Res, 1999;16:1564-1569.
[6] Alyautdin RN, Petrov VE, Langer K, Berthold A, Kharkevich DA, Kreuter J. Delivery of loperamide across the blood-brain barrier with polysorbate 80-coated polybutylcyanoacrylate nanoparticles. Pharm Res, 1997,14:325-328.
[7] Alyautdin RN, Tezikov EB, Ramge P, etal. Significant entry of tubocurarine into the brain of rats by adsorption to polysorbate 80-coated polybutylcyanoacrylate nanoparticles: an in situ brain perfusion study. J Microencapsul, 1998;15:67-74.
[8] Kreuter J, Alyautdin RN, Kharkevich DA, etal. Passage of peptides through the blood-brain barrier with colloidal polymer particles(nanoparticles). Brain Res, 1995,674:171-174.
[9] Friese A, Seiller E, Quack G, Lorenz B, Kreuter J. Increase of the duration of the anticonvulsive activity of a novel NMDA receptor antagonist using poly (butylcyanoacrylate) nanoparticles as a parenteral controlled release system. Eur J Pharm Biopharm, 2000,49:103-109.
[10] Matsumura Y, Maeda H. A new concept for macromolecular therapeutics in cancer chemotherapy: mechanism of tumoritropic accumulation of proteins and the antitumor agent smancs. Cancer Res, 1986,46:6387-6392.
[11] Maeda H, Matsumura Y. Tumoritropic and lymphotropic principles of macromolecular drugs. Crit Rev Ther Drug Carrier Syst, 1989,6:193-210.
[12] Seymour LW, Miyamoto Y, Maeda H, etal. Influence of molecular weight on passive tumour accumulation of a soluble macromolecular drug carrier. Eur J Cancer, 1995,31A:766-770.
[13] Duncan R, Dimitrijevic S, Evagorou EG. The role of polymer conjugates in the diagnosis and treatment of cancer. Pharma Sci, 1996,6:237-263.
[14] Kreuter J, Shamenkov D, Petrov V, Ramge P, Cychutek K, Koch-Brandt C, Alyautdin R. Apolipoprotein-mediated transport of nanoparticle-bound drugs across the blood- brain barrier. J Drug Targeting, 2002,10:317-325.
[15] Olivier JC, Fenart L, Chauvet R,etal. Indirect evidence that drug brain targeting using polysorbate 80-coated polybutylcyanoacrylate nanoparticles is related to toxicity. Pharm Res, 1999,16:1836-1842.
[16] Alyaudtin RN, Reichel A, Lobenberg R, etal. Interaction of poly(butylcyanoacrylate) nanoparticles with the blood-brain barrier in vivo and in vitro. J Drug Target,2001, 9:209-221.
[17] Kreuter J, Ramge P, Petrov V, etal. Direct evidence that polysorbate-80-coated poly(butylcyanoacrylate) nanoparticles deliver drugs to the CNS via specific mechanisms requiring prior binding of drug to the nanoparticles. Pharm Res, 2003, 20:409 -416.
[18] S.C. Steiniger, J. Kreuter, A.S. Khalansky, etal. Chemotherapy of glioblastoma in rats using doxorubicin-loaded nanoparticles. Int. J. Cancer , 2004,109: 759-767.
[19] A. Ambruosi, S. Gelperina, A. Khalansky, etal. Antitumor effect of doxorubicin loaded in poly(butyl cyanoacrylate) nanoparticles in rat glioma model: influence of formulation parameters. J. Microencapsulation, 2006,23 :582-592.
[20] J. Kreuter, D. Shamenkov, V. Petrov, etal. Alyautdin, Apolipoprotein-mediated transport of nanoparticle-bound drugs across the blood–brain barrier.J. Drug Target, 2002,10:317-325.
[21] P. Ramge, R.E. Unger, J.B. Oltrogge, etal.Polysorbate 80-coating enhances uptake of polybutylcyanoacrylate(PBCA)-nanoparticles by human and bovine primary braincapillary endothelial cells. Eur. J. Neurosci, 2000,12:1931-1940.
[22] R.N.Alyautdin, A.Reichel, R.L?benberg,etal. Interaction of poly(butylcyanoacrylate) nanoparticles with the blood brain barrier in vivo and in vitro. J. Drug Target, 2001,9 :209-221.
[23] S.M. Moghimi, A.C. Hunter. Poloxamers and poloxamines in nanoparticle engineering and experimental medicine. Trends Biotechnol, 2000,18:412-420.
[24] A. Ambruosi, H. Yamamoto, J. Kreuter. Body distribution of polysorbate-80 and doxorubicin-loaded [14C]poly(butylcyanoacrylate) nanoparticles after i.v. administration in rats. J. Drug Target, 2005,13:535-542.
[25] A. Colin de Verdiere, C. Dubernet, F. Nemati, etal. Reversion of multidrug resistance with polyalkylcyanoacrylate nanoparticles: towards a mechanism of action.Br. J. Cancer, 1997,76:198-205.
[26] U. Panzenboeck, Z. Balazs, A. Sovic, A. Hrzenjak, etal. ABCA1 and scavenger receptor classB, type I, are modulators of reverse sterol transport at an in vitro blood brain barrier constituted of porcine brain capillary endothelial cells. J. Biol.Chem, 2002,277:42781-42789.
[27]V. Lenaerts, etal. Degradation of poly(isobutyl cyanoacrylate) nanoparticles. Biomaterials, 1984, 5 :65-68.
[28] M. Stein, E. Hamacher.Degradation of poly(butyl cyanoacrylate) microparticles. Int. J. Pharm,80:R11 -R13.
[29] M.A. El-Egakey, V. Bentele, J. Kreuter. Molecular weights of polycyanoacrylate nanoparticles. Int. J.Pharm,1983,13:349-352.
[30] S.J.Douglas,S.S.Davis, S.R.Holding. Molecular weights of poly(butyl 2-cyanoacrylate)produced during nanoparticle formation. Br. Polym. J, 1985, 17:339-342.
[31] L. Grislain, et al., Pharmacokinetics and distribution of a biodegradable drug-carrier, Int. J. Pharm. 1983, 15:335-345.
[32] J. Kreuter. Application of nanoparticles for the delivery of drugs to the brain. International Congress Series, 2005,1277: 85-94.
[2] Cristiano,Richard J.Targeted,non-viral gene delivery for cancer gene therapy [J].Front Biosci.1998,3:D1161-D1170.
[3] SONG CX,LABHASETWAR V,MURPHY H,etal.Formulation and characterization of biodegradable nanoparticales for intravascular local drug delivery[J]. J Contr Rel.1997,43(3):197-201.
[4] Cleland L. Solvent evaporation processes for the production of controlled release biodegradable microsphere formulations for therapeutics and vaccines[J]. Biotechnology Progerss.1998,14:102-107.
[5] Song C, Labhasetwar V,Levy J.Controlled release of U-86983 from double-layer biodegradable matrices: effect of additives on release mechanism and kinetics[J].J Controlled Release.1997,45:177-182.
[6] LUBBE AS, BERGEMANN C, RIESS H,etal.Clincal experiecses with magnetic drug targeting: a phase I study with 4'-epidoxorubicin in 14 patients with advanced aolid tumors [J].Cancer Res,1996,56(20):4686.
[7] GREIDER CW. Telomerase activity,cell proliferation and cancer[J]. Proc Natl Acad Sci USA,1998,95:90.
[8] OLIVIER Z, FRANCIS C, SZOKA J. Intracellular distribution and mechanism of delivery of oligonucleotides mediated by cationic lipid [J]. PharmRes. 1996,13(9): 1367-1372.
[9] Lambert G,Fattal E,Couvreur P,et al.Nanoparticulate systems for the delivery of Antisense oligonucleotides[J].Adv Drug Deliv Rev.2001,47(1):99-112.
[10] Cohen H,Levy RJ, Gao J, et al. Sustained delivery and expression of DNA encapsulated in polymeric nanoparticles [J].Gene Therapy.2000,7:1896-1905.
[11] Labhasetwar V et al. A DNA controlled-release coating for gene transfer: transfection in skeletal and cardiac mulscle[J].J PHarm Sci.1998,87:1347-1350.
[12] Leong KW, et a1.DNA-polycation nanospheres as non-virus gene delivery vehicles[J]. J Control Release. 1998,53:183-193.
[13]De Verdiere AC, Dubernet C, Nemati F, et al. Revertion of muitidrug resistance with polyalkylcyanoacrylate nanoparticles: towards a mechanism of action[J]. British J Cancer,1997,76(2):198-205.
[14]Gonzalez-Martin G, Isabel Merino M, Rodriguez-Cabezas N,et al.Characterization and trypanocidal activity of Nifurtimox-containing and empty nanoparticles of polyethylcyanoacrylate[J].J PHarm PHarmacol,1998,50(1):29-35.
[1] Stan AC, Casares S, Radu D, Walter GF, Brumeanu TD. Doxorubicin induced cell death in highly invasive human gliomas. Anticancer Res. 1999;19:941-50.
[2] Walter KA,Tamargo RJ, Qlivi A, Bueger PC,Brem H. Intratumoral chemotheraphy.Neurosurgery .1995,37:1128-45.
[3] von Holst H, Knochenhauer E, Blomgren H, Collins VP, Ehn L, Lindquist M, Noren G, Peterson C. Uptake of adriamycin in tumour and surrounding brain tissue in patients with malignant gliomas. Acta Neurochir 1990;104:13-16.
[4] Steiniger SC, Kreuter J, Khalansky AS, Skidan IN, Bobruskin AI, etal. Chemotherapy of glioblastoma in rats using doxorubicin-loaded nanoparticles. Int J Cancer. 2004,109:759-767.
[5]M.Simeonova,R.Velichkova,G.Ivanova,etal.Poly(butyalcyanoacrylate)nanoparticles for topical delivery of 5-fluorouacil.Int.J.Pharm. 2003,263:133-140.
[6]Alyaudtin.R,Reichel.A,L?benberg.R,etal.Internation of poly(butyalcyanoacrylate) nanoparticles with the Blood-Brain Barrier in vivo and in vitro.Journal of Drug Targeting.No.3,Vol.9:209-221.
[7]杨西晓,陈建海,郭丹.聚氰基丙烯酸正丁酯纳米粒的生物相容性研究[J].第一军医大学学报,2005,25(10):1261-1262.
[8] S. I. Park, J. H. Lim, Y. H. Hwang etal. In vivo and in vitro antitumor activity of doxorubicin-loaded magnetic fluids. phys. stat. sol. (c) 4, No. 12, 4345-4351 (2007).
[9] J?rg Kreuter,Peter Ramge,Valery Petrovetal. Direct Evidence that Polysorbate-80- Coated Poly(Butylcyanoacrylate) Nanoparticles Deliver Drugs to the CNS via Specific Mechanisms Requiring Prior Binding of Drug to the Nanoparticles. Pharmaceutical Research,2003 , No. 3,Vol. 20:409-416.
[10]Dunn SE, Coombes AGA, Garnett MC, Davis SS, Davies MC, Illum L. In vitro cell interaction and in vivo biodistribution of poly(lactide-co-glycolide) nanospheres surface modified by poloxamer and poloxamine copolymers. J Controlled Release 1997;44:65–76.
[11]Lourenco C, Teixeira M, Simoes S, Gaspar R. Steric stabilization of nanoparticles: Size and surface properties. Int J Pharm1996;138:1–12.
[12] Coombes AGA, Yeh MK, Lavelle EC, Davis SS. The control of protein release from poly(DL-lactide-co-glycolide) microparticles by variation of the external aqueousphase surfactant in the water-in oil-in-water method. J Controlled Release. 1998;52:311–320.
[13] Quintanar–Guerrero D, Ganem–Quintanar A, Allemann E, etal. Influence of the stabilizer coating layer on the purification and freeze-drying of poly(D,L-lactic acid) nanoparticles prepared by an emulsion-diffusion technique. J Microencapsulation 1998;15:107-119.
[14]Bouillot P, Babak V, Dellacherie E. Novel bioresorbable and bioeliminable surfactants for microsphere preparation. Pharm Res, 1999,16:148–154.
[1] Stan AC, Casares S, Radu D,Walter GF, Brumeanu TD. Doxorubicin induced cell death in highly invasive human gliomas. Anticancer Res ,1999,19:941-950.
[2] Brigger I,Dubernet C,Courveur P. Nanoparticles in cancer therapy and diagnosis. Advdrug Del Rev, 2002,54:631-651.
[3] Ferrari M.Cancer nanotechnology.Opportunities and challenges. Nat Rev Cancer, 2005, 5:161-171..
[4] Hillyer J,Albrecht R.Gastrointestinal persorption and tissue distribution of differently sized colloidal gold nanoparticles.J Pharm Sci, 2001,90:1927-1936.
[5] Kayser O,Lemke A,Hernandez-Trejo N.The impact of nanobiotechnology on the development of new drug delivery systems.Curr Pharm Biotechnol, 2005,6:3-5..
[6] Kreuter J.Nanoparticulate systems for the brain delivery of drugs. Advdrug Deliv Rev, 2001, 47:65-81.
[7] Kreuter J,Ramge P,Petrov V,etal.Direct evidence that polysorbate-80-coated (Polybutylcyanoacrylate) nanoparticles deliver drugs to the CNS via specific mechanisms requiring prior binding of drugs to the nanoparticles. Pharm Res, 2003,20 :409-416.
[8] Vauthier C, Dubernet C, Fattal E,etal. Poly(butylcyanoacrylate ) as biodegradable materials for biomedical applications. Advdrug Deliv Rev, 2003,55:519-548.
[9]杨西晓,陈建海,郭丹.聚氰基丙烯酸正丁酯纳米粒的生物相容性研究[J].第一军医大学学报,2005,25(10):1261-1262.
[10] 1. R. Alyautdin, D. Gothier, V. Petrov, D. Kharkevich, and J.Kreuter. Analgesic activity of the hexapeptide dalargin adsorbed on the surface of polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles. Eur. J. Pharm. Biopharm, 1995 ,41:44-48.
[11] R. N. Alyautdin, V. E. Petrov, K. Langer, etal. Kreuter. Delivery of loperamide across the blood–brain barrier with poly-sorbate 80-coated polybutylcyanoacrylate nanoparticles. Pharm. Res, 1997,14:325-328.
[12] R. N. Alyautdin, E. B. Tezikov, P. Ramge, etal. Significant entry of tubocurarine into the brain of rats by absorption to polysorbate 80-coated polybutyl-cyanoacrylate nanoparticles: an in situ brain perfusion study.J. Microencapsul, 1998, 15:67-74 .
[13] A. E. Gulyaev, S. E. Gelperina, I. N. Skidan,etal. Kreuter. Significant transport of doxorubicin into the brain with polysorbate 80-coated nanoparticles. Pharm.Res, 1999,16:1564-1569.
[14] A. Friese, E. Seiler, G. Quack, B. Lorenz, and J. Kreuter. Enhancement of the duration of the anticonvulsive activity of a novel NMDA receptor antagonist using poly(butylcyanoacylate) nanoparticles as a parenteral controlled release delivery system.Eur. J. Pharm. Biopharm, 2000, 49:103-109.
[15] Sebastian C.J. Steiniger, Jorg Kreuter, Alexander S,etal. Chemotherphy of glioblastoma in rats using doxorubicin-loaded nanoparticles. Cancer, 2004,109: 759-767.
[16] Mahaley MS Jr. Immunological considerations and the malignant gliomas problem. Clin Neurosurg, 1968,15:175-189.
[17] Giometto B, Bozza F, Faresin F, etal.Immune infiltrates and cytokines in gliomas.Acta Neurochir, 1996,138:50-56.
[18] S, Mehraein P, Graeber MB. Differential expression of MHC class II molecules by microglia and neoplastic astroglia: relevance for the escape of astrocytoma cells from immune surveillance. Neuropathol Appl Neurobiol,1998,24:293-230.
[19] Aldskogius H, Liu L, Svensson M. Glial responses to synaptic damage and plasticity. J Neurosci Res,1999,58:33-41.
[20] Badie B, Schartner J. Role of microglia in glioma biology. Microsc Res Tech, 2001,54:106 -113.
[21] Boiardi A, Pozzi A, Salmaggi A, etal.Safety and potential effectiveness of daunorubicin-containing liposomesin patients with advanced recurrent malignant CNS tumors.Cancer Chemother Pharmacol, 1999,43:178-179.
[22] Lippens RJ. Liposomal daunorubicin (DaunoXome) in children with recurrent orprogressive brain tumors. Pediatr Hematol Oncol, 1999,16:131-139.
[23] Koukourakis MI, Koukouraki S, Fezoulidis I, etal. High intratumoural accumulation of stealth liposomal doxorubicin (Caelyx) in glioblastomas and in metastatic brain tumours. Br J Cancer, 2000,83:1281-1286.
[24] Gelperina SE, Khalansky AS, Skidan IN, etal.Toxicological studies of doxorubicin bound to polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles in healthy rats and rats with intracranial glioblastoma.Toxicol Lett,2002,126:131-141.
[25] Neuwelt EA, Pagel M, Barnett P, etal. Pharmacology and toxicity of intracarotid adriamycin administration following osmotic blood-brain barrier modification. Cancer Res,1981,41:4466-4470.
[1] HANEY C B,FUTCHER C W,GREIDER C W.Telomeres shorten during of human fibroblasts[J].Natttre,1990,245:458-460.
[2] Makarov VL, Hirose Y, Langmore JP: Long G tails at both ends of human chromosomes suggest a C strand degradationmechanism for telomere shortening. Cell 1997, 88:657-666.
[3] Wright WE, Tesmer VM, Huffman KE, Levene SD, Shay JW: Normal human chromosomes have long G-rich telomeric overhangs at one end. Genes Dev, 1997, 11:2801-2809.
[4] Colgin LM, Reddel RR: Telomere maintenance mechanisms and cellular immortalization. Curr Opin Genet Dev,1999, 9:97-103.
[5] Fattal E, Vauthier C, Ayine I, et al. Biadegadable polyalkylcyanoacrylate nano- particles for the delivery of oligonucleotides[J]. J Controlled Release,1998,53:137.
[1] Bennett WP, Hollstein MC, Hsu IC,etal.Mutational spectra and immunohistochemical analysis of p53 in human cancer [J]. Chest,1992,101:19-20.
[2]杜子威,徐庚达,王尧等.人脑恶性胶质瘤体外细胞系的建立及其特征.中华肿瘤杂志.1984;6(4):241-245.
[3] W. J. Bowers, D. F. Howard, and H. J. Federoff. Gene therapeutic strategies for neuroprotection: implications for Parkinson,s disease. Exp. Neurol, 1997,144:58–68.
[4] B. Blits, and M. B. Bunge. Direct gene therapy for repair of the spinal cord. J. Neurotrauma, 2006,23:508-520.
[5] J. M. Alisky, and B. L. Davidson. Gene therapy for amyotrophic lateral sclerosis and other motor neuron diseases. Hum.Gene. Ther, 2000, 11:2315-2329.
[6] R. Tinsley, and P. Eriksson. Use of gene therapy in central nervous system repair. Acta. Neurol. Scand, 2004,109:1-8.
[7] T. Federici, and N. M. Boulis. Gene-based treatment of motor neuron diseases. Muscle Nerve, 2006,33:302-323.
[8] B. K. Kaspar, J. Llado, N. Sherkat, etal. Retrograde viral delivery of IGF-1 prolongs survival in amouse ALS model. Science, 2003, 301:839-842.
[9] A. P. Kells, D. M. Fong, M. Dragunow,etal. AAV-mediated gene delivery of BDNF or GDNF is neuroprotective in a model of Huntington disease. Mol. Ther, 2004,9:682-688.
[10] R. J. Mandel, S. K. Spratt, R. O. Snyder,etal.Midbrain injection of recombinant adeno-associated virus encoding rat glial cell line-derived neurotrophic factor protects nigral neurons in a progressive 6-hydroxydopamine-induced degeneration model of Parkinson,s disease in rats. Proc. Natl.Acad. Sci. U S A, 1997, 94:14083-14088.
[11] D. L. Choi-Lundberg, Q. Lin, T. Schallert, etal.C. Bohn. Behavioral and cellular protection of rat dopaminergic neurons by an adenoviral vector encoding glial cell line-derived neurotrophic factor. Exp. Neurol, 1998, 154:261-275.
[12] J. A. Gonzalez-Barrios, M. Lindahl, M. J. Bannon, etal. Neurotensin polyplex as an efficient carrier for delivering the human GDNF gene into nigral dopamine neurons of hemiparkinsonian rats.Mol. Ther, 2006(14):857-865.
[13] Y. Zhang, F. Schlachetzki, Y. F. Zhang,etal. Normalization of striatal tyrosine hydroxylase and reversal of motor impairment in experimental parkinsonism with intravenous nonviral gene therapy and a brain-specific promoter. Hum. Gene. Ther, 2004(15):339-350.
[14] M. J.During, J. R. Naegele,K.L.O .Malley, etal. Longterm behavioral recovery in parkinsonian rats by an HSV vector expressing tyrosine hydroxylase. Science, 1994,266:1399-1403.
[15] C. S. Hong, W. F. Goins, J. R. Goss, etal. Herpes simplex virus RNAi and neprilysin gene transfer vectors reduce accumulation of Alzheimer,s diseaserelated amyloid-beta peptide in vivo. Gene. Ther, 2006(13):1068-1079.
[16] Cho-Chung YS. Antiense DNAs as a targeted therapeutics for cancer : no longer a dream[J]. Curr Opin Investig Drugs,2002(3):934-939.
[17] D. J. Begley. Delivery of therapeutic agents to the central nervous system: theproblems and the possibilities. Pharmacol.Ther, 2004,104:29-45.
[18] R. J. Boado. RNA interference and nonviral targeted gene therapy of experimental brain cancer. NeuroRx, 2005, 2:139-150.
[19] W. M. Pardridge. Tyrosine hydroxylase replacement in experimental Parkinson_s disease with transvascular gene therapy.NeuroRx, 2005,2:129-138.
[20] W.M.Pardridge.Blood-brain barrier delivery. Drug Discov.Today, 2007,12:54-61.
[21] Chavany C,Sasion-Behmoaras T,le Don T,etal.Adsorption of oligonucleotides onto polyisohexylcyanoacrylate nanoparticles protects them against nuclease and increase their cellular uptake[J].Pharm Res,1994,11(9):1370.
[22] Andrei Maksimenko,Claude Malvy, Gregory Lambert, etal.Oligonucleotides Targeted Against a Junction Oncogene Are Made Efficient by Nanotechnologies. Pharmaceutical Research, 2003,10(20):1565-1567.
[1] Konmta T,Kanzawa T, Kondo Y ,et a1. Telomerase as a therapeutic target for malignant gliomas[J]. Oncogene,2002,21(4):656-663.
[2] Neidle S, Kellan d LR.Telomerase as an anti-cancer target:current status and future prospects[J].Anticancer Drug Des,1999,14(4):341-347.
[3] Singh A,Sharma H,Salhan S,et a1.Evaluation of expression of apoptosis-related proteins and their correlation with HPV,telomerase activity,and apoptotic index in cervical cancer[J].Pathobiology,2004,71(6):314-322.
[4] Iida A ,Yamaguchi A, Hirose K. Telomerase activity in colorectal cancer and itsrelationship to bel-2 expresion[J].J Surg Oncol,2000,73(4):219-223.
[5] Neidle S,Parkinson G.Telomere maintenan ce as a target for anticancer drug discovery[J].Nat Rev Drug Discov,2002,l(5):383-393.
[6] Kondo S,Tanaka Y,Kondo Y,et a1.Antisense telomerase treatment:induction of two distinct pathways,apoptosis and differentiation[J].FASEB J,1998,12(10):801-811.
[7]罗元辉,房殿存,畅仕明,等.端粒酶在原发肝癌中的表达及临床意义[J].中华肝脏病杂志,1998,6(3):133-135.
[8]许兰涛.端粒酶反义寡核苷酸抗肿瘤的实验研究[J].中华肝脏病杂志,2001, 9(2):80.
[9] Kondo Y, Koga S, Komata T, et al. Treatment of prostate cancer in vitro and vivo w ith 2-5A–anti-telomerase RNA component.Oncogene, 2000, 19:2205-2211.
[10] M ukai S, Kondo Y, Koga S, et al. 2-5A antisense telomerase RNA therapy for intracranial malignant gliomas. Cancer Res,2000, 60:4461-4467.
[1] Couvreur P,Kante B,Roland M,etal.Poly(alkylcyanoacrylate) nanoparticles as potential lysosomotropic carriers: preparation,morphological and sorptive properties.J Pharm Pharmacol,1979,31:331-332.
[2] Kreuter,J. Evaluation of nanoparticles as drug-delivery systems. I. preparations methods. Pharm.Acta Helv,1983,58,196-209.
[3] D. J. Begley, M. W. Bradbury, and J. Kreuter (eds.). The Blood-Brain Barrier and Drug Delivery to the CNS, Marcel DekkerNew York, 2000.
[4] Alyautdin,R.N,Gothier D,Petro V,Kharkevich D, etal.Analgesic activity of the hexapeptide dalargin on the surface of polysorbate 80-coated polybutylcyanoacrylate nanoparticles. Eur J.Biopharm,1998,41:44-48.
[5] Gulyaev AE, Gelperina SE, Skidan IN, Antropov AS, Kivman GY, Kreuter J. Significant transport of doxorubicin into the brain with polysorbate 80-coated nanoparticles. Pharm Res, 1999;16:1564-1569.
[6] Alyautdin RN, Petrov VE, Langer K, Berthold A, Kharkevich DA, Kreuter J. Delivery of loperamide across the blood-brain barrier with polysorbate 80-coated polybutylcyanoacrylate nanoparticles. Pharm Res, 1997,14:325-328.
[7] Alyautdin RN, Tezikov EB, Ramge P, etal. Significant entry of tubocurarine into the brain of rats by adsorption to polysorbate 80-coated polybutylcyanoacrylate nanoparticles: an in situ brain perfusion study. J Microencapsul, 1998;15:67-74.
[8] Kreuter J, Alyautdin RN, Kharkevich DA, etal. Passage of peptides through the blood-brain barrier with colloidal polymer particles(nanoparticles). Brain Res, 1995,674:171-174.
[9] Friese A, Seiller E, Quack G, Lorenz B, Kreuter J. Increase of the duration of the anticonvulsive activity of a novel NMDA receptor antagonist using poly (butylcyanoacrylate) nanoparticles as a parenteral controlled release system. Eur J Pharm Biopharm, 2000,49:103-109.
[10] Matsumura Y, Maeda H. A new concept for macromolecular therapeutics in cancer chemotherapy: mechanism of tumoritropic accumulation of proteins and the antitumor agent smancs. Cancer Res, 1986,46:6387-6392.
[11] Maeda H, Matsumura Y. Tumoritropic and lymphotropic principles of macromolecular drugs. Crit Rev Ther Drug Carrier Syst, 1989,6:193-210.
[12] Seymour LW, Miyamoto Y, Maeda H, etal. Influence of molecular weight on passive tumour accumulation of a soluble macromolecular drug carrier. Eur J Cancer, 1995,31A:766-770.
[13] Duncan R, Dimitrijevic S, Evagorou EG. The role of polymer conjugates in the diagnosis and treatment of cancer. Pharma Sci, 1996,6:237-263.
[14] Kreuter J, Shamenkov D, Petrov V, Ramge P, Cychutek K, Koch-Brandt C, Alyautdin R. Apolipoprotein-mediated transport of nanoparticle-bound drugs across the blood- brain barrier. J Drug Targeting, 2002,10:317-325.
[15] Olivier JC, Fenart L, Chauvet R,etal. Indirect evidence that drug brain targeting using polysorbate 80-coated polybutylcyanoacrylate nanoparticles is related to toxicity. Pharm Res, 1999,16:1836-1842.
[16] Alyaudtin RN, Reichel A, Lobenberg R, etal. Interaction of poly(butylcyanoacrylate) nanoparticles with the blood-brain barrier in vivo and in vitro. J Drug Target,2001, 9:209-221.
[17] Kreuter J, Ramge P, Petrov V, etal. Direct evidence that polysorbate-80-coated poly(butylcyanoacrylate) nanoparticles deliver drugs to the CNS via specific mechanisms requiring prior binding of drug to the nanoparticles. Pharm Res, 2003, 20:409 -416.
[18] S.C. Steiniger, J. Kreuter, A.S. Khalansky, etal. Chemotherapy of glioblastoma in rats using doxorubicin-loaded nanoparticles. Int. J. Cancer , 2004,109: 759-767.
[19] A. Ambruosi, S. Gelperina, A. Khalansky, etal. Antitumor effect of doxorubicin loaded in poly(butyl cyanoacrylate) nanoparticles in rat glioma model: influence of formulation parameters. J. Microencapsulation, 2006,23 :582-592.
[20] J. Kreuter, D. Shamenkov, V. Petrov, etal. Alyautdin, Apolipoprotein-mediated transport of nanoparticle-bound drugs across the blood–brain barrier.J. Drug Target, 2002,10:317-325.
[21] P. Ramge, R.E. Unger, J.B. Oltrogge, etal.Polysorbate 80-coating enhances uptake of polybutylcyanoacrylate(PBCA)-nanoparticles by human and bovine primary braincapillary endothelial cells. Eur. J. Neurosci, 2000,12:1931-1940.
[22] R.N.Alyautdin, A.Reichel, R.L?benberg,etal. Interaction of poly(butylcyanoacrylate) nanoparticles with the blood brain barrier in vivo and in vitro. J. Drug Target, 2001,9 :209-221.
[23] S.M. Moghimi, A.C. Hunter. Poloxamers and poloxamines in nanoparticle engineering and experimental medicine. Trends Biotechnol, 2000,18:412-420.
[24] A. Ambruosi, H. Yamamoto, J. Kreuter. Body distribution of polysorbate-80 and doxorubicin-loaded [14C]poly(butylcyanoacrylate) nanoparticles after i.v. administration in rats. J. Drug Target, 2005,13:535-542.
[25] A. Colin de Verdiere, C. Dubernet, F. Nemati, etal. Reversion of multidrug resistance with polyalkylcyanoacrylate nanoparticles: towards a mechanism of action.Br. J. Cancer, 1997,76:198-205.
[26] U. Panzenboeck, Z. Balazs, A. Sovic, A. Hrzenjak, etal. ABCA1 and scavenger receptor classB, type I, are modulators of reverse sterol transport at an in vitro blood brain barrier constituted of porcine brain capillary endothelial cells. J. Biol.Chem, 2002,277:42781-42789.
[27]V. Lenaerts, etal. Degradation of poly(isobutyl cyanoacrylate) nanoparticles. Biomaterials, 1984, 5 :65-68.
[28] M. Stein, E. Hamacher.Degradation of poly(butyl cyanoacrylate) microparticles. Int. J. Pharm,80:R11 -R13.
[29] M.A. El-Egakey, V. Bentele, J. Kreuter. Molecular weights of polycyanoacrylate nanoparticles. Int. J.Pharm,1983,13:349-352.
[30] S.J.Douglas,S.S.Davis, S.R.Holding. Molecular weights of poly(butyl 2-cyanoacrylate)produced during nanoparticle formation. Br. Polym. J, 1985, 17:339-342.
[31] L. Grislain, et al., Pharmacokinetics and distribution of a biodegradable drug-carrier, Int. J. Pharm. 1983, 15:335-345.
[32] J. Kreuter. Application of nanoparticles for the delivery of drugs to the brain. International Congress Series, 2005,1277: 85-94.