纳米载体在肿瘤靶向治疗中的应用

近年来纳米材料的研究正成为医药学者研究的焦点,纳米载体作为控释系统所具有的特殊性质,使其在药物输送方面具有高效,稳定,特异性强等优点,因其粒径呈纳米态,可透过细胞膜和穿透血脑屏障等,能更好的向靶器官,靶细胞,靶分子输送药物,起到更好的治疗效果.文章针对纳米载体的结构和特点,综述了纳米载体在肿瘤靶向治疗方面的国内外最新研究进展.     

纳米靶向给药系统载体材料的研究进展

目的:综述及讨论近年来新材料在靶向给药系统载体材料方面的应用及存在问题。资料来源:以网络数据库资源为主,查询Science Direct,Pubmed,Springer,Ovid等数据库1995-01/2006-06关于靶向给药系统载体材料方面的相关文献,检索词“Targetdrug delivery system”and“Materials”,限定文章的语种种类为“English”;同时检索CNKI数据库2001-01/2006-06的相关文章为补充,限定文章语种种类为中文,检索关键词“靶向给药系统,载体材料”。资料选择:对资料进行初审,纳入标准:①关于靶向给药系统的研究。②应用的较新的材料作为药物载体。排除标准:重复性研究。资料提炼:共收集到122篇文献,98篇符合纳入标准,取其中有关新型载体材料应用于靶向给药系统的30篇相关文献进行归纳总结。资料综合:靶向给药是目前药剂学领域的研究热点之一,但多年来转化为产品的研究成果极为少见。其中关键问题之一就是药物载体材料的研究瓶颈,药剂学研究与材料学研究间学科交叉与衔接的严重滞后阻碍了新型给药系统的产业化。结论:新型给药系统载体材料研究开发尤为重要,载体材料应用的安全性一旦解决,必将带来新型给药系统全面用于临床的新局面,从而使药效得到更好的发挥,提高疾病的预防治疗能力。     

多功能靶向纳米载体的制备及其对神经母细胞瘤的靶向性和MRI示踪性观察

目的 制备集靶向siRNA传递及MRI显像功能于一体的多功能靶向纳米载体,并验证其对神经母细胞瘤的靶向性及MRI示踪性.方法 制备PEG-PEI-SPION,scAbGD2 -PEG-PEI-SPION并检测其基本特性.利用凝胶阻滞检测纳米载体与siRNA的复合能力;以CLSM及MRI验证纳米载体的靶向性.结果 PEG-PEI-SPION,scAbGD2-PEG-PEI-SPION平均粒径分别为(60.0±2.3)nm及(90.0±7.8)nm.氮磷比(N/P)≥2.2时siRNA与PEG-PEI-SPION,scAbGD2 -PEG-PEI-SPION完全复合.CLSM实验及MRI结果证实scAbGD2-PEG-PEI-SPION具有靶向性.结论 scAbGD2-PEG-PEI-SPION对神经母细胞瘤具有靶向传递siRNA及MRI显影的功能.     

超声靶向微泡破碎联合半乳糖聚乙烯亚胺介导凋亡素基因诱导肝癌细胞凋亡的研究

目的:探讨超声靶向微泡破碎(ultrasound targeted microbubble destruction,UTMD)联合半乳糖聚乙烯亚胺(PEI-Gal)介导凋亡素基因(VP3)诱导肝癌细胞HepG2凋亡的影响。方法:体外培养HepG2细胞,随机分为4组:(1)对照组;(2)PEI-Gal组;(3)PEI-Gal+超声组;(4)PEI-Gal+超声+微泡组。转染24h后荧光显微镜观察pEGFP-VP3在HepG2细胞中的表达,流式细胞仪检测转染效率,RT-PCR和Western blot分别检测VP3 mRNA和蛋白表达水平,AnnexinV-FITC/PI检测细胞凋亡率。结果:流式细胞仪、RT-PCR和Western blot分析表明,PEI-Gal+超声+微泡组的转染效率为(28.83±2.07)%、VP3 mRNA水平为0.92±0.02,蛋白水平为1.65±0.06,HepG2凋亡率为(37.40±2.12)%,均显著高于其他各组(均P<0.01)。结论:UTMD联合PEI-Gal可明显提高VP3基因转染HepG2细胞的效率及在HepG2细胞内的表达,增加HepG2细胞的凋亡率。     

人骨髓间充质干细胞靶向纳米基因载体制备及体外细胞磁共振成像

目的 探讨靶向纳米基因载体单链神经节苷脂抗体-聚乙二醇-聚乙烯亚胺-超顺磁性氧化铁（scAbGD2-PEG-g-PEI-SPION）转染人骨髓间充质干细胞（hBMSCs）的可行性、效率及体外细胞MR显像能力。方法 合成scAbGD2-PEG-g-PEI-SPION后,采用凝胶阻滞实验评估其复合外源性基因的能力;动态光散射法测量scAbGD2-PEG-g-PEI-SPION/pDNA纳米复合物的粒径大小及表面电位;体外细胞毒性实验检测其对hBMSCs的细胞毒性。采用流式细胞仪检测scAbGD2-PEG-g-PEI-SPION靶向转染hBMSCs的效率,并设置PEG-g-PEI-SPION组、cAbGD2-PEG-g-PEI-SPION组、抗体竞争抑制（scAbGD2-PEG-g-PEI-SPION+free AbGD2）组和同型抗体（scAbIgG2a-PEG-g-PEI-SPION）组,通过激光共聚焦显微镜及普鲁士蓝染色观察hBMSCs对纳米复合物的摄入。通过体外细胞MR扫描验证scAbGD2-PEG-g-PEI-SPION的MR成像功能。结果 scAbGD2-PEG-g-PEI-SPION细胞毒性小,复合外源性基因后能够形成稳定的纳米复合物,粒径80~100 nm。在相同的N/P比值下,scAbGD2-PEG-g-PEI-SPION组的转染率明显高于其他组（P<0.001）。N/P=20时,靶向组具有最高转染率[（59.60±4.50）%]。同时,scAbGD2-PEG-g-PEI-SPION中的SPION可有效标记hBMSCs,在MR T2/T2^*加权图像上呈低信号。结论 scAbGD2-PEG-g-PEI-SPION是一种MRI可视的、可有效转染hBMSCs的靶向纳米基因载体。     

无法手术的肝细胞性肝癌的分子靶向治疗

肝细胞性肝癌严重威胁着人类健康。迄今为止,索拉非尼仍是唯一一个被批准用于无法手术的肝细胞性肝癌的药物。近年来,分子靶向治疗药物层出不穷,但其在肝细胞性肝癌的治疗方面却停滞不前,多个III期临床研究都以失败告终。本文详细的回顾了过去2年内公开报道的分子靶向药物在肝癌中的II期和III期临床研究结果,就其研究失败的结果进行了总结,为接下来在肝癌中开展临床研究的药物的研发给出了一点意见和建议。     

纳米药物的靶向作用及不良反应

目的:探讨纳米药物的靶向作用及其在疾病治疗中的不良反应.方法:由第一作者以"纳米药物,纳米微粒,药物载体,靶向作用,不良反应"为检索词,在中国期刊全文数据库(CNKI:2001/2009)中,采用电子检索的方式进行文献检索,语言设定为中文.纳入主题内容与纳米药物的靶向作用及不良反应联系紧密的文章,排除Meta分析、与主题无关的文章及重复性研究.共纳入18篇文章,归纳总结纳米药物靶向作用及不良反应的研究现状.结果:纳米药物与普通制剂的药物相比,具有较大的表面积、较强的化学活性、较快的吸收速度,在通过生物体的各种屏障、控制药物的释放速度、设定药物的靶向性等许多方面,纳米药物都具有一般药物不可替代的优越性.结论:靶向性纳米药物制剂与以往药物剂型比较,最突出的优点是具有明显的靶向性,也就是说它能将药按设计途径输送到药物的靶位.不仅可提高疗效,而且可降低药物的不良反应.纳米药物作为崭新的一类制剂,为治疗一些难治性疾病提供了崭新的思路,给医药界带来了观念性变革.但一些纳米材料显示出毒性特征,特别是纳米材料与人体或环境相互作用的后果还不清楚.因此在纳米材料被加工成商业化产品之前,应充分研究其生物学效应、作用机制及其毒性的消除措施,为合理利用纳米材料奠定理论基础.     

Gal-PEG-PEI/psiRNA肝靶向性纳米基因载体对人胎肝细胞系L-02细胞的转染效率检测

目的：检测半乳糖-聚乙二醇-聚乙烯亚胺/小干扰RNA质粒（Gal-PEG-PEI/psiRNA）肝靶向性纳米基因载体对人胎肝细胞系L-02细胞的转染效率。方法：合成Gal-PEG-PEI/psiRNA纳米复合物,采用纳米粒径仪测定复合物的粒径和zeta电位,用不同Gal-PEG-PEI氨基与psiRNA磷酸基比例（N/P）的Gal-PEG-PEI/psiRNA纳米复合物转染L-02细胞,以Lipofectamine2000、非靶向性载体PEG-PEI/psiRNA和裸psiRNA转染为对照组,48 h后用流式细胞仪测定转染效率。转染前加入1 mg半乳糖观察其半乳糖竞争拮抗结果。结果：Gal-PEG-PEI/psiRNA纳米复合物的粒径随N/P增大而减小,N/P≥1/15时达最小粒径,约为80 nm。Gal-PEG-PEI/psiRNA纳米基因载体的转染率为（20.4±0.9）%,明显高于非肝靶向性载体PEG-PEI/psiRNA（P〈0.01）及裸psiRNA（P〈0.01）,但低于Lipofectamine 2000（P〈0.05）;加入竞争性拮抗剂半乳糖后Gal-PEG-PEI/psiRNA纳米基因载体的转染率下降至（4.8±2.1）%,PEG-PEI/psiRNA介导的转染率则不受影响。结论：Gal-PEG-PEI/psiRNA纳米基因载体可明显提高人胎肝细胞系L-02细胞的转染效率,且具有良好的肝细胞靶向性。     

靶向非病毒载体的构建及介导外源基因转导卵巢癌细胞的效率的研究

目的 探索靶向非病毒载体（GE7）系统在人卵巢癌细胞的转导效率。方法 组建由表皮生长因子受体（EGF．R）介导的靶向非病毒载体（GE7）系统，分别与标志基因β-半乳糖苷酶（β-gal）和治疗基因I型单纯疱疹病毒胸腺嘧啶核苷激酶基因（HSV1-TK）构成载体复合物，体外转导卵巢癌细胞CaOV3，通过X-gal染色、Northernblot分析，观察了GE7系统对外源基因的转导效率。结果 X-gal染色显示，外源基因的导入效率可达80％，呈瞬时表达；Northern杂交结果表示转导了HSV1-TK基因的卵巢癌细胞提取的RNA有阳性条带的显示。结论 GE7载体系统能高效靶向安全地将外源基因导入卵巢癌细胞。     

生物载体硫交联聚乙烯亚胺的合成及递送小干扰RNA的实验研究

目的合成可生物降解的基因载体,并分析其生物毒性及转染率。方法低分子量聚乙烯亚胺(PEI)通过双硫键交联合成可降解的高分子量PEI衍生物(SS-PEI),通过红外光谱和核磁波谱分析技术分析其化学结构,采用细胞活力实验和检测大鼠肝肾功能指标分析其细胞和体内毒性,并转染羟基荧光素修饰的siRNA(FAM-siRNA)分析细胞转染率。结果红外波谱和核磁波谱分析可见酰胺键的特征波谱,噻唑蓝法和肝肾功能指标显示SS-PEI不同剂量组与对照组的差异均无统计学意义(P>0.05),SS-PEI/FAM-siRNA转染率为(76.0±2.8)%。结论 SS-PEI的合成可明显提高装载siRNA的效率,具有安全、高效等特点。     

Galactosylated chitosan-graft-poly(ethylene glycol) as hepatocyte-targeting DNA carrier.

Lactobionic acid bearing galactose group was coupled with chitosan for liver specificity, and poly(ethylene glycol) (PEG) was grafted to galactosylated chitosan (GC) for stability in water and enhanced cell permeability. Complex formation of galactosylated chitosan-graft-PEG (GCP)/DNA complexes was confirmed by agarose gel electrophoresis. Compared to GC/DNA complex, the stability of GCP/DNA complex could be enhanced. Particle sizes of GCP/DNA complexes decreased as the charge ratio of GCP to DNA increased and had a minimum value around 27 nm at the charge ratio of 5. Conformational change of DNA did not occur after complex formation with GCP compared to conformation of DNA itself. GCP/DNA complexes were only transfected into Hep G2 having asialoglycoprotein receptors (ASGR), indicative of specific interaction of ASGR on cells and galactose ligands on GCP.     

Synthesis of biodegradable multi-block copolymers of poly(L-lysine) and poly(ethylene glycol) as a non-viral gene carrier.

Biodegradable and non-toxic multi-block copolymers based on poly(L-lysine) and poly(ethylene glycol) were synthesized. Synthesized copolymers showed almost negligible cytotoxicity above 95% cell viability and transfection efficiency compared to the PLL homopolymer with molecular weight of 25,700. Biodegradation under physiological conditions revealed that the molecular weight of copolymers decreased to 20% of the initial molecular weight within 72 h. Transfection efficiencies of copolymers were not affected by the presence of serum, while that of PLL homopolymer decreased to the level of naked DNA in the presence of serum. Based on the results, the new copolymers are believed to be a potentially efficient carrier for the delivery of bioactive agents.     

Injectable microfluidic hydrogel microspheres based on chitosan and poly(ethylene glycol) diacrylate (PEGDA) as chondrocyte carriers

Direct injection of chondrocytes in a minimally invasive way has been regarded as the significantly potential treatment for cartilage repair due to their ability to fill various irregular chondral defects. However, the low cell retention and survival after injection still limited their application in clinical transformation. Herein, we present chondrocyte-laden microspheres as cell carriers based on a double-network hydrogel by the combination of the chitosan and poly(ethylene glycol) diacrylate (PEGDA). The microfluidic technique was applied to prepare size-controllable chitosan/PEGDA hydrogel microspheres (CP-MSs) via the water-in-oil approach after photo-crosslinking and physical-crosslinking. The chondrocytes were laden on CP-MSs, which showed good cell viability and proliferation after long-term cell cultivation. The in vitro investigation further demonstrated that chondrocyte-laden CP-MSs were injectable and the cell viability was still high after injection. In particular, these cell-laden microspheres were self-assembled into a 3D cartilage-like scaffold by a bottom-up strategy based on cell–cell interconnectivity, which suggested that these injectable chondrocyte-laden microspheres showed potential applications as chondrocyte carriers for bottom-to-up cartilage tissue engineering.

Chitosan/PEGDA double-network hydrogel microspheres prepared by microfluidic method as chondrocyte carriers for bottom-up cartilage tissue engineering.     

Degradable polyethylenimine-alt-poly(ethylene glycol) copolymers as novel gene carriers.

An ideal gene carrier requires both safety and transfection efficiency. Polyethylenimine (PEI) is a well-known cationic polymer, which has high transfection efficiency owing to its buffering capacity. But it has been reported that PEI is cytotoxic in many cell lines and non-degradable. In this study, we synthesized degradable PEI-alt-poly(ethylene glycol) (PEG) copolymers using Michael-type addition reactions as a new gene carrier and characterized them. These copolymers were complexed with plasmid DNA and the resulting complexes were characterized by dynamic light scattering, gel retardation and atomic force microscopy to determine particle sizes, complex formation and complex shape, respectively. Cytotoxicity and transfection efficiency of the copolymers were also checked in cultured HeLa human cervix epithelial carcinoma cells, HepG2 human hepatoblastoma cell line and MG63 human osteosarcoma cells. PEG to PEI ratio in the copolymers was near 1 and the molecular weight of the copolymer ranged from around 8000 to 12,900. These copolymers degraded rapidly at 37 degrees C in 0.1 M phosphate buffered saline (PBS, pH 7.4). The complete copolymer/DNA complex was formed at an N/P ratio of 12, producing a complex resistant to DNase I. Particle sizes decreased with increasing N/P ratio and PEG molecular weight, exhibiting a minimum value of 75 nm at an N/P ratio of 45 with PEI-alt-PEG (700). Cytotoxicity study showed that copolymers exhibited no cytotoxic effects on cells even at high copolymer concentration. Also, transfection efficiency was influenced by PEG molecular weight and, in case of PEI-alt-PEG (258), the transfection efficiency was higher than that for PEI 25 K in HepG2 and MG63, whereas it was lower than that for PEI 25K in HeLa cells.     

Micellar carriers based on block copolymers of poly(epsilon-caprolactone) and poly(ethylene glycol) for doxorubicin delivery.

Diblock copolymers of poly(epsilon-caprolactone) (PCL) and monomethoxy poly(ethylene glycol) (MPEG) with various compositions were synthesized. The amphiphilic block copolymers self-assembled into nanoscopic micelles and their hydrophobic cores encapsulated doxorubicin (DOX) in aqueous solutions. The micelle diameter increased from 22.9 to 104.9 nm with the increasing PCL block length (2.5-24.7 kDa) in the copolymer composition. Hemolytic studies showed that free DOX caused 11% hemolysis at 200 microg ml(-1), while no hemolysis was detected with DOX-loaded micelles at the same drug concentration. An in vitro study at 37 degrees C demonstrated that DOX-release from micelles at pH 5.0 was much faster than that at pH 7.4. Confocal laser scanning microscopy (CLSM) demonstrated that DOX-loaded micelles accumulated mostly in cytoplasm instead of cell nuclei, in contrast to free DOX. Consistent with the in vitro release and CLSM results, a cytotoxicity study demonstrated that DOX-loaded micelles exhibited time-delayed cytotoxicity in human MCF-7 breast cancer cells.     

Hyaluronic acid grafted with poly(ethylene glycol) as a novel peptide formulation.

Hyaluronic acids (HA) grafted with poly(ethylene glycol) (PEG) (PEG-g-HA) were synthesized. The materials characterization, enzymatic degradability and peptide (insulin) release from solutions of the copolymers were examined. Distribution of bioactive peptides within the polymer chain is well-known for combinations of PEG and polysaccharides as aqueous polymer two-phase systems. Insulin was preferentially partitioned into the PEG phase in a PEG/HA solution system. Enzymatic degradation of the copolymers was strongly dependent on the PEG content. Thermal analysis revealed that PEG-g-HA exhibited a variation in phase-separated structures depending on the PEG content. The solution of PEG-g-HA enabled insulin to remain in the PEG moieties dispersed in the HA matrix. Leakage of insulin from the copolymers was dependent upon the PEG content. Leakage rate of insulin from copolymer containing between 7 and 39% by weight of PEG were similar. A dramatic increase in leakage rate occurred when the PEG content was increased to greater than 39% by weight. It is considered that the loaded insulin was partitioned into the PEG moieties and became entangled with the PEG chains. The conformational change of insulin was effectively prevented in PEG-g-HA solutions, although insulin was denatured in storage of both phosphate buffered solution and HA solution. Such a heterogeneous-structured polymeric solution may be advantageous as an injectable therapeutic formulation for ophthalmic or arthritis treatment.     

Poly(ethylene glycol)–poly(propylene glycol)–poly(ethylene glycol) and polyvinylidene fluoride blend doped with oxydianiline-based thiourea derivatives as a novel and modest gel electrolyte system for dye-sensitized solar cell applications

Unique symmetrical thiourea derivatives with an oxydianiline core were synthesized using cost-effective and simple methods. A new gel electrolyte system was prepared using these thiourea additives along with a highly conductive PEG–PPG–PEG block copolymer, PVDF, and an iodide/triiodide redox couple. The PEG units present in the electrolyte are well-known for their intense segmental motion of ions, which can degrade the recombination rate and favour the charge transfer. The thiourea additives interacted well with the redox couple to limit iodine sublimation and their adsorption induced a negative potential shift for TiO₂. The highest efficiency attained by utilizing such gel polymer electrolytes was 5.75%, especially with 1,1′-(oxybis(4,1-phenylene))bis(3-(6-methylpyridin-2-yl) thiourea) (OPPT), under an irradiation of 100 mW cm⁻². The electrochemical impedance spectroscopy, UV-vis absorption spectroscopy, differential scanning calorimetry, and FTIR spectroscopy data of such gel polymer electrolytes favoured the PCE order of the additives used in DSSCs. The improvement in the DSSC performance with symmetrical thioureas having electron-rich atoms was practically attributed to the reduction of back electron transfer, dye regeneration, and hole transport.

A unique gel polymer electrolyte was prepared using PVDF and PEG–PPG–PEG block copolymer with I⁻/I₃⁻ for DSSC application. This is a cost-effective method used for the synthesis of thiourea additives. The GPE with OPPT thiourea additive achieved a good efficiency of 5.7%.