CpG oligodeoxynucleotide-loaded PAMAM dendrimer-coated magnetic nanoparticles promote apoptosis in breast cancer cells

One major application of nanotechnology in cancer treatment involves designing nanoparticles to deliver drugs, oligonucleotides, and genes to cancer cells. Nanoparticles should be engineered so that they could target and destroy tumor cells with minimal damage to healthy tissues. This research aims to develop an appropriate and efficient nanocarrier, having the ability of interacting with and delivering CpG-oligodeoxynucleotides (CpG-ODNs) to tumor cells. CpG-ODNs activate Toll-like receptor 9 (TLR9), which can generate a signal cascade for cell death. In our study, we utilized three-layer magnetic nanoparticles composed of a Fe₃O₄ magnetic core, an aminosilane (APTS) interlayer and a cationic poly(amidoamine) (PAMAM) dendrimer. This will be a novel targeted delivery system to enhance the accumulation of CpG-ODN molecules in tumor cells. The validation of CpG-ODN binding to DcMNPs was performed using agarose gel electrophoresis, UV-spectrophotometer, XPS analyses. Cytotoxicity of conjugates was assessed in MDA-MB231 and SKBR3 cancer cells based on cell viability by XTT assay and flow cytometric analysis. Our results indicated that the synthesized DcMNPs having high positive charges on their surface could attach to CpG-ODN molecules via electrostatic means. These nanoparticles with the average sizes of 40 ± 10 nm bind to CpG-ODN molecules efficiently and induce cell death in MDA-MB231 and SKBR3 tumor cells and could be considered a suitable targeted delivery system for CpG-ODN in biomedical applications. The magnetic core of these nanoparticles represents a promising option for selective drug targeting as they can be concentrated and held in position by means of an external magnetic field.     

Effect of gemcitabine and retinoic acid loaded PAMAM dendrimer-coated magnetic nanoparticles on pancreatic cancer and stellate cell lines

Gemcitabine is an anticancer drug used in the treatment of different cancer types, including pancreatic ductal adenocarcinoma. The maximum tolerated dose in humans is restricted by its side effects on healty cells. Furthermore, the fibrotic stroma produced by the pancreatic stellate cells prevents effective delivery of chemotherapeutic agents providing a safe-haven for the cancer cells. This becomes more of a problem considering the short half-life of this drug. Magnetic nanoparticle-based targeted drug delivery systems are a promising alternative to overcome the limitations of classical chemotherapies. The aim of this study is to obtain an effective targeted delivery system for gemcitabine using magnetic nanoparticles (MNPs) and all-trans retinoic acid (ATRA). This dual approach targets the tumor cells and its infrastructure – stellate cells – simultaneously. Gemcitabine and ATRA were loaded onto the PAMAM dendrimer-coated magnetic nanoparticles (DcMNPs), which were synthesized and characterized previously. Drug loading and release characteristics, and stability of the nanoparticles were investigated. Gemcitabine and ATRA loaded MNPs are efficiently taken up by pancreatic cancer and stellate cells successfully targeting and eliminating both cells. Results of this study can provide new insights on pancreatic cancer therapy where tumor is seen as a system with its stroma insead of epithelial cells alone.     

Dynamics of cellular entry and drug delivery by dendritic polymers into human lung epithelial carcinoma cells

Dendrimers and hyperbranched polymers are emerging as potentially ideal drug delivery vehicles because they provide a significant amount of tailorability and a large density of functional groups. This study explores the dynamics of cellular entry of dendrimers and hyperbranched polymers alone, and in the complexed form with ibuprofen, into A549 human lung epithelial carcinoma cells using UV/Vis spectroscopy, flow cytometry and fluorescence microscopy. Both dendrimers and hyperbranched polymers appear to enter these cells rapidly. The polyamidoamine (PAMAM) dendrimers, with NH₂ and OH end functionalities appear to enter cells (in approx. 1 h) faster than the hyperbranched polyol (OH functionality) (in approx. 2 h). Cellular entry of PAMAM-NH₂ was detected as early as 5 min. All branched polymers and their ibuprofen complexes entered A549 lung epithelial cells rapidly when compared to the pure drug. The drug payload was about 50% by weight in the complexes formed by PAMAM-NH₂ dendrimers and was about 30% in the encapsulated form for Polyol-OH and PAMAM-OH. The complexation and encapsulation of ibuprofen with the polymers appear to facilitate rapid cellular entry of ibuprofen. The anti-inflammatory effect of the polymer-complexed drug was demonstrated by more rapid suppression of COX-2 mRNA levels than that achieved by the pure drug. This suggests that these dendritic polymers can act as efficient drug carriers, delivering high 'payloads' of drug even with complexation and encapsulation.     

Biomimetic mineralization of collagen fibrils induced by amine-terminated PAMAM dendrimers – PAMAM dendrimers for remineralization

Objective: Achieving biomimetic mineralization of collagen fibrils by mimicking the role of non-collagenous proteins (NCPs) with biomimetic analogs is of great interest in the fields of material science and stomatology. Amine-terminated PAMAM dendrimer (PAMAM-NH₂), which possesses a highly ordered architecture and many calcium coordination sites, may be a desirable template for simulating NCPs to induce mineralization of collagen fibrils. In this study, we focused on the ability of PAMAM-NH₂ to mineralize collagen fibrils. Design: Type-I collagen fibrils were reconstituted over 400-mesh formvar-and-carbon-coated gold grids and treated with a third-generation PAMAM-NH₂ (G3-PAMAM-NH₂) solution. The treated collagen fibrils were immersed in artificial saliva for different lengths of time. The morphologies of the mineralized reconstituted type-I collagen fibrils were characterized by transmission electron microscopy. Results: No obvious mineralized collagen fibrils were detected in the control group. On the contrary, collagen fibrils were heavily mineralized in the experimental group. Most importantly, intrafibrillar mineralization was achieved within the reconstituted type-I collagen fibrils. Conclusions: In this study, we successfully induced biomimetic mineralization within type-I collagen fibrils using G3-PAMAM-NH₂. This strategy may serve as a potential therapeutic technique for restoring completely demineralized collagenous mineralized tissues.     

Electrochemistry at gold nanoparticles deposited on dendrimers assemblies adsorbed onto gold and platinum surfaces

Cyclic voltammetry and atomic force microscopy (AFM) were used to investigate the electrochemical communication between gold nanoparticles deposited onto adsorbed dendrimers of 4th generation poly(amido amine) (PAMAM) and gold or platinum electrode surfaces. Combination of these techniques evidenced that adsorption of dendrimers depends on both their chemical structures and the nature of the metallic surface. The adsorption process could be controlled as a function of either the dendrimer concentration or the time during which the electrodes were soaked in the dendrimer containing solution. Accordingly, mono- and multi-layer films of dendrimers could be obtained. The presence of dendrimer multilayer film did not prevent redox species such as ferrocene to reach the electrode surface presumably through pinholes in the dendrimers array. Interestingly, the dendrimer layer did not behave as an insulating film but was “electrochemically transparent” over the time ranges investigated. Thus, depositing gold nanoparticles over the dendrimer layers and constructing a self-assembled monolayer (SAM) consisting of 6-ferrocenyl hexanethiol onto the gold surfaces still allowed electrochemical communication with the ferrocene groups. This opens the future possibility of using dendrimers as anchors for gold nanoparticles so as to construct 2D-arrays of metal nanoparticles for electronic applications.     

聚酰胺-胺对永生化的人角膜上皮细胞的毒性

目的考察一种新型药物载体——聚酰胺-胺[poly(amidoamine),PAMAM]对体外培养的角膜上皮细胞的毒性作用,为其在眼部药物载体的应用提供理论依据。方法建立应用于PAMAM毒性研究的角膜上皮细胞模型,通过细胞毒性分析测定不同浓度(10g·L-1、1g·L-1、0.1g·L-1、0.01g·L-1、0.001g·L-1)、不同代数(G3.0、G3.5、G4.0、G4.5、G5.0)的PAM-AM在共同孵育5min、15min、60min对角膜上皮细胞存活率的影响并计算其IC50值。结果整代PAMAM(G3.0、G4.0、G5.0)和半代PAMAM(G3.5、G4.5)均随着代数的增大,细胞毒性增加,且在一定浓度范围内细胞毒性随着药物浓度增大、药物接触的时间延长而增加。半代PAMAM细胞毒性均低于整代。G3.0、G3.5、G4.5PAMAM在1g·L-1及以下浓度毒性反应均为合格,G4PAMAM在0.1g·L-1浓度及以下毒性评级大部分为合格,而G5PAMAM只在0.01g·L-1和0.001g·L-1下部分毒性评级合格。结论低代或半代PAMAM可能更适合作为眼部药物给药的载体,也可考虑通过结构修饰等手段降低高代PAMAM毒性。     

PAMAM/sh-miR-34a纳米复合物对黑素瘤细胞的抑制作用

目的 将树枝状聚合物聚酰胺-胺(PAMAM)包载可以表达具有抑制人恶性黑素瘤细胞A375增殖、侵袭和转移的miR-34a的质粒sh-miR-34a,构建聚阳离子纳米复合物PAMAM/sh-miR-34a.考察形成的复合物的粒径、zeta电位、细胞摄取,以及对A375细胞的抑制作用.方法 利用粒度测定仪测定纳米复合物的粒径和电位,凝胶电泳实验测定PAMAM对sh-miR-34a的包裹能力;利用罗丹明标记的sh-miR-34a考察A375细胞对纳米复合物的摄取;CCK8法测定PAMAM/sh-miR-34a对A375细胞的抑制作用;Transwell法测定PAMAM/sh-miR-34a抑制A375细胞迁移和侵袭作用;蛋白质印迹法测定PAMAM/sh-miR-34a对A375细胞中pAkt、pRb、pERK1/2蛋白表达的阻滞作用.结果 PAMAM包裹sh-miR-34a可形成稳定的纳米复合物,在N/P=20时,细胞对PAMAM/sh-miR-34a的摄取达最高(P＜0.05).PAMAM可以有效携带sh-miR-34a进入A375细胞,体外抑制A375细胞的增殖、侵袭和迁移,并且可以阻滞A375细胞中pAkt、pRb和pERK1/2蛋白的表达.结论 PAMAM可以包裹sh-miR-34a并对人恶性黑素瘤A375细胞起到抑制作用.     

生物材料聚酰胺-胺树状大分子合成及其体外细胞相容性评价

主要通过Michael加成和酰胺化逐步迭代合成反应得到聚酰胺-胺（PAMAM）树状大分子并对合成产物进行了结构表征。对生物医学有潜在应用价值的第五代（G5．0）提出了廉价易行的合成提纯步骤并通过细胞计数、MTF检测和FCM对L-929细胞系进行了体外生物相容性评价分析。结果表明，按照本试验的合成及纯化所得PAMAM G5．0对L-929细胞系在给药范围内无细胞毒性，无致瘤性，有良好的细胞相容性。     

Scaling Behaviour of PAMAM Dendrimers Determined by Diffusion NMR

Summary: The hydrodynamic radius of PAMAM dendrimers as a function of molar mass is investigated by diffusion NMR. As a characteristic length, the hydrodynamic radius is calculated by Stokes‐Einstein equation. Poly(amidoamine) (PAMAM) dendrimers of generations 0 to 7 of two different terminal groups (NH₂, COONa) have been investigated. This dependence of the hydrodynamic radius from molar mass is compared with a scaling model and statistical model. A scaling exponent of 3.7 has been found in both cases. The scaling exponent found exceeds the dimension of the embedding space, thus the possible growth of dendrimers of this structure is limited.

Hydrodynamic radius of PAMAM‐NH₂ □, PAMAM‐COONa ○ as a function of molar mass.     

Enhanced anti-hepatocarcinoma efficacy by GLUT1 targeting and cellular microenvironment-responsive PAMAM–camptothecin conjugate

The efficient targeting of drugs to tumor cell and subsequent rapid drug release remain primary challenges in the development of nanomedicines for cancer therapy. Here, we constructed a glucose transporter 1 (GLUT1)-targeting and tumor cell microenvironment-sensitive drug release Glucose–PEG–PAMAM-s-s–Camptothecin-Cy7 (GPCC) conjugate to tackle the dilemma. The conjugate was characterized by a small particle size, spherical shape, and glutathione (GSH)-sensitive drug release. In vitro tumor targeting was explored in monolayer (2D) and multilayer tumor spheroid (3D) HepG2 cancer cell models (GLUT1⁺). The cellular uptake of GPCC was higher than that in the control groups and that in normal L02 cells (GLUT1^?), likely due to the conjugated glucose moiety. Moreover, the GPCC conjugate exhibited stronger cytotoxicity, higher S arrest and enhanced apoptosis and necrosis rate in HepG2 cells than control groups but not L02 cells. However, the cytotoxicity of GPCC was lower than that of free CPT, which could be explained by the slower release of CPT from the GPCC compared with free CPT. Additional in vivo tumor targeting experiments demonstrated the superior tumor-targeting ability of the GPCC conjugate, which significantly accumulated in tumor meanwhile minimize in normal tissues compared with control groups. The GPCC conjugate showed better pharmacokinetic properties, enabling a prolonged circulation time and increased camptothecin area under the curve (AUC). These features contributed to better therapeutic efficacy and lower toxicity in H22 hepatocarcinoma tumor-bearing mice. The GLUT1-targeting, GSH-sensitive GPCC conjugate provides an efficient, safe and economic approach for tumor cell targeted drug delivery.