Nanoparticles for Nucleic Acids Delivery - Tecrea Ltd

Nanoparticles and Nucleic Acids Delivery is a group in Biological Sciences on Mendeley.

Beyerle, A. et al., 2009. In vitro cytotoxic and immunomodulatory profiling of low molecular weight polyethylenimines for pulmonary application. Toxicology in vitro : an international journal published in association with BIBRA, 23(3), pp.500-8. Available at: http://www.ncbi.nlm.nih.gov/pubmed/19444927 [Accessed April 7, 2012].

Abstract: Polyethylenimines (PEI) are potent non-viral nucleic acid delivery vehicles used for gene delivery and RNA interference (RNAi). For non-invasive pulmonary RNAi therapy the respiratory tissue is an attractive application route, but offers particularly unwanted side-effects like cytotoxicity as well as inflammatory and immune responses. In the current study, we determined the most crucial issues of pulmonary applications for two low molecular weight PEIs in comparison to the well-known lung toxic crystalline silica. Cytotoxic effects and inflammatory responses were evaluated in three murine pulmonary target cell lines, the alveolar epithelial (LA4), the alveolar macrophage (MH-S) and the macrophage-monocyte-like (RAW 264.7) cell line. For both PEIs, cytotoxicity was detected most prominently in the alveolar epithelial cells and only at high doses. Cytokine responses, in contrast were observed already at low PEI concentrations and could be divided into three groups, induced (i) by free PEI (IL-6, TNF-alpha, G-CSF), (ii) by PEI/siRNA complexes (CCL2, -5, CXCL1, -10), or (iii) unaffected by either treatment (IL-2, -4,-7, -9, and CCL3). We conclude that even for the respiratory tissue both PEIs represent powerful siRNA delivery tools with reduced cytotoxicity and minor proinflammatory potency. However, in relation to response levels observed upon crystalline silica exposures, some PEI induced proapoptotic and proinflammatory responses might not be considered completely harmless, therefore further in vivo investigations are advisable.

Davis, M.E., 2009. The first targeted delivery of siRNA in humans via a self-assembling, cyclodextrin polymer-based nanoparticle: from concept to clinic. Molecular pharmaceutics, 6(3), pp.659-68. Available at: http://dx.doi.org/10.1021/mp900015y [Accessed April 7, 2012].

Abstract: Experimental therapeutics developed to exploit RNA interference (RNAi) are now in clinical studies. Here, the translation from concept to clinic for the first experimental therapeutic to provide targeted delivery of synthetic, small interfering RNA (siRNA) in humans is described. This targeted, nanoparticle formulation of siRNA, denoted as CALAA-01, consists of a cyclodextrin-containing polymer (CDP), a polythethylene glycol (PEG) steric stabilization agent, and human transferrin (Tf) as a targeting ligand for binding to transferrin receptors (TfR) that are typically upregulated on cancer cells. The four component formulation is self-assembled into nanoparticles in the pharmacy and administered intravenously (iv) to patients. The designed features of this experimental therapeutic are described, and their functions illustrated.

Douglas, K.L., Piccirillo, C.A. & Tabrizian, M., 2008. Cell line-dependent internalization pathways and intracellular trafficking determine transfection efficiency of nanoparticle vectors. European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft für Pharmazeutische Verfahrenstechnik e.V, 68(3), pp.676-87. Available at: http://dx.doi.org/10.1016/j.ejpb.2007.09.002 [Accessed March 26, 2012].

Abstract: It has been suggested that cell physiology may affect the internalization pathways of non-viral vectors, leading to cell line-dependent transfection efficiency. To verify this hypothesis, fluorescently labeled alginate-chitosan nanoparticle complexes were used as non-viral vectors to transfect 293T, COS7, and CHO cells and to observe the cellular interactions and internalization mechanisms of the complexes in each cell line. 293T cells, which demonstrate the highest transfection efficiency, internalize complexes primarily through clathrin-mediated processes. COS7 cells also demonstrate some internalization of complexes through the clathrin-dependent pathway, explaining the moderate transfection exhibited. In contrast, CHO cells internalize complexes predominantly through caveolin-mediated pathways and are not transfected. Results suggest that following clathrin-mediated endocytosis, complexes are trafficked to the endo-lysosomal pathway, where the proton-sponge effect leads to their release into the cytosol. Contrarily, the absence of trafficking to this pathway following caveolin-mediated endocytosis results in vesicle-entrapped complexes that become transfection-incompetent. These results demonstrate that cell physiology is a critical factor in efficient transfection, and that trafficking to the endo-lysosomal pathway through specific internalization mechanisms is essential for transfection with alginate-chitosan nanoparticle complexes.

Eliyahu, H., Barenholz, Y. & Domb, A.J., 2005. Polymers for DNA delivery. Molecules (Basel, Switzerland), 10(1), pp.34-64. Available at: http://www.ncbi.nlm.nih.gov/pubmed/18007276 [Accessed April 7, 2012].

Abstract: Nucleic acid delivery has many applications in basic science, biotechnology, agriculture, and medicine. One of the main applications is DNA or RNA delivery for gene therapy purposes. Gene therapy, an approach for treatment or prevention of diseases associated with defective gene expression, involves the insertion of a therapeutic gene into cells, followed by expression and production of the required proteins. This approach enables replacement of damaged genes or expression inhibition of undesired genes. Following two decades of research, there are two major methods for delivery of genes. The first method, considered the dominant approach, utilizes viral vectors and is generally an efficient tool of transfection. Attempts, however, to resolve drawbacks related with viral vectors (e.g., high risk of mutagenicity, immunogenicity, low production yield, limited gene size, etc.), led to the development of an alternative method, which makes use of non-viral vectors. This review describes non-viral gene delivery vectors, termed “self-assembled” systems, and are based on cationic molecules, which form spontaneous complexes with negatively charged nucleic acids. It introduces the most important cationic polymers used for gene delivery. A transition from in vitro to in vivo gene delivery is also presented, with an emphasis on the obstacles to achieve successful transfection in vivo.

Giljohann, D.A. et al., 2009. Gene regulation with polyvalent siRNA-nanoparticle conjugates. Journal of the American Chemical Society, 131(6), pp.2072-3. Available at: http://dx.doi.org/10.1021/ja808719p [Accessed April 7, 2012].

Abstract: We report the synthesis and characterization of polyvalent RNA-gold nanoparticle conjugates (RNA-Au NPs), nanoparticles that are densely functionalized with synthetic RNA oligonucleotides and designed to function in the RNAi pathway. The particles were rationally designed and synthesized to be free of degrading enzymes, have a high surface loading of siRNA duplexes, and contain an auxiliary passivating agent for increased stability in biological media. The resultant conjugates have a half-life six times longer than that of free dsRNA, readily enter cells without the use of transfection agents, and demonstrate a high gene knockdown capability in a cell model.

Guo, S. et al., 2010. Enhanced gene delivery and siRNA silencing by gold nanoparticles coated with charge-reversal polyelectrolyte. ACS nano, 4(9), pp.5505-11. Available at: http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=3044603&tool=pmcentrez&rendertype=abstract [Accessed April 7, 2012].

Abstract: Charge-reversal functional gold nanoparticles first prepared by layer-by-layer technique were employed to deliver small interfering RNA (siRNA) and plasmid DNA into cancer cells. Polyacrylamide gel electrophoresis measurements of siRNA confirmed the occurrence of the charge-reversal property of functional gold nanoparticles. The expression efficiency of enhanced green fluorescent protein (EGFP) was improved by adjuvant transfection with charge-reversal functional gold nanoparticles, which also had much lower toxicity to cell proliferation. Lamin A/C, an important nuclear envelope protein, was effectively silenced by lamin A/C-siRNA delivered by charge-reversal functional gold nanoparticles, whose knockdown efficiency was better than that of commercial Lipofectamine 2000. Confocal laser scanning microscopic images indicated that there was more cy5-siRNA distributed throughout the cytoplasm for cyanine 5-siRNA/polyethyleneimine/cis-aconitic anhydride-functionalized poly(allylamine)/ polyethyleneimine/11-mercaptoundecanoic acid-gold nanoparticle (cy5-siRNA/PEI/PAH-Cit/PEI/MUA-AuNP) complexes. These results demonstrate the feasibility of using charge-reversal functional gold nanoparticles as a means of improving the nucleic acid delivery efficiency.

Hallaj-Nezhadi, S., Lotfipour, F. & Dass, C.R., 2010. Delivery of nanoparticulate drug delivery systems via the intravenous route for cancer gene therapy. Die Pharmazie, 65(12), pp.855-9. Available at: http://www.ncbi.nlm.nih.gov/pubmed/21284252 [Accessed April 7, 2012].

Abstract: While the systemic route of administration enables therapeutic genes to spread through the bloodstream and access target cells, it is a challenge to achieve this. Several studies demonstrate that systemic administration of therapeutic genes or other nucleic acid-based constructs such as siRNA to solid tumors as well as cancer metastases are better with nanoparticulate systems compared to administration of free (uncomplexed) nucleic acids. Nanoparticle-based nucleic acid delivery systems might be more pertinent, due to the several privileges in terms of enhanced tissue penetrability, improved cellular uptake and to a lesser extent, targeted gene delivery to the cells of interest provided targeting ligands are used. Systemic delivery of nanoplexes has already been reported with different nanoparticles containing DNA via various routes of administration. The goal of the present article is to review the current state of intravenous delivery of nanoparticles for gene therapy of cancer.

Hayes, M.E. et al., 2006. Genospheres: self-assembling nucleic acid-lipid nanoparticles suitable for targeted gene delivery. Gene therapy, 13(7), pp.646-51. Available at: http://www.ncbi.nlm.nih.gov/pubmed/16341056 [Accessed March 15, 2012].

Abstract: We describe the assembly of a cationic lipid-nucleic acid nanoparticle from a liquid monophase containing water and a water miscible organic solvent where both lipid and DNA components are separately soluble prior to their combination. Upon removal of the organic solvent, stable and homogenously sized (70-100 nm) lipid-nucleic acid nanoparticles (Genospheres) were formed. The low accessibility (<15%) of the nanoparticle-encapsulated DNA to a DNA intercalating dye indicated well-protected nucleic acids and high DNA incorporation efficiencies. It was demonstrated that Genospheres could be stably stored under a variety of conditions including a lyophilized state where no appreciable increase in particle size or DNA accessibility was observed following reconstitution. Finally, Genospheres were made target-specific by insertion of an antibody-lipopolymer (anti-HER2 scFv (F5)-PEG-DSPE) conjugate into the particle. The target specificity (>100-fold) in HER2 overexpressing SK-BR-3 breast cancer cells was dependent on the degree of PEGylation, where the incorporation of high amounts of PEG-lipid on the particle surface (up to 5 mol%) had only a minor effect on the transfection activity of the targeted Genospheres. In summary, this work describes a novel, readily scalable method for preparing highly stable immunotargeted nucleic acid delivery vehicles capable of achieving a high degree of specific transfection activity.

Howard, K.A. et al., 2006. RNA interference in vitro and in vivo using a novel chitosan/siRNA nanoparticle system. Molecular therapy : the journal of the American Society of Gene Therapy, 14(4), pp.476-84. Available at: http://dx.doi.org/10.1016/j.ymthe.2006.04.010 [Accessed March 16, 2012].

Abstract: This work introduces a novel chitosan-based siRNA nanoparticle delivery system for RNA interference in vitro and in vivo. The formation of interpolyelectrolyte complexes between siRNA duplexes (21-mers) and chitosan polymer into nanoparticles, ranging from 40 to 600 nm, was shown using atomic force microscopy and photon correlation spectroscopy. Rapid uptake (1 h) of Cy5-labeled nanoparticles into NIH 3T3 cells, followed by accumulation over a 24 h period, was visualized using fluorescence microscopy. Nanoparticle-mediated knockdown of endogenous enhanced green fluorescent protein (EGFP) was demonstrated in both H1299 human lung carcinoma cells and murine peritoneal macrophages (77.9% and 89.3% reduction in EGFP fluorescence, respectively). In addition, Western analysis showed approximately 90% reduced expression of BCR/ABL-1 leukemia fusion protein while BCR expression was unaffected in K562 (Ph(+)) cells after transfection using nanoparticles containing siRNA specific to the BCR/ABL-1 junction sequence. Effective in vivo RNA interference was achieved in bronchiole epithelial cells of transgenic EGFP mice after nasal administration of chitosan/siRNA formulations (37% and 43% reduction compared to mismatch and untreated control, respectively). These findings highlight the potential application of this novel chitosan-based system in RNA-mediated therapy of systemic and mucosal disease.

Jensen, L.B. et al., 2011. Elucidating the molecular mechanism of PAMAM-siRNA dendriplex self-assembly: effect of dendrimer charge density. International journal of pharmaceutics, 416(2), pp.410-8. Available at: http://www.ncbi.nlm.nih.gov/pubmed/21419201 [Accessed April 7, 2012].

Abstract: Dendrimers are attractive vehicles for nucleic acid delivery due to monodispersity and ease of chemical design. The purpose of this study was to elucidate the self-assembly process between small interfering RNA (siRNA) and different generation poly(amidoamine) dendrimers and to characterize the resulting structures. The generation 4 (G4) and G7 displayed equal efficiencies for dendriplex aggregate formation, whereas G1 lacked this ability. Nanoparticle tracking analysis and dynamic light scattering showed reduced average size and increased polydispersity at higher dendrimer concentration. The nanoparticle tracking analysis indicated that electrostatic complexation results in an equilibrium between differently sized complex aggregates, where the centre of mass depends on the siRNA:dendrimer ratio. Isothermal titration calorimetric data suggested a simple binding for G1, whereas a biphasic binding was evident for G4 and G7 with an initial exothermic binding and a secondary endothermic formation of larger dendriplex aggregates, followed by agglomeration. The initial binding became increasingly exothermic as the generation increased, and the values were closely predicted by molecular dynamics simulations, which also demonstrated a generation dependent differences in the entropy of binding. The flexible G1 displayed the highest entropic penalty followed by the rigid G7, making the intermediate G4 the most suitable for dendriplex formation, showing favorable charge density for siRNA binding.

Lentacker, I. et al., 2008. New strategies for nucleic acid delivery to conquer cellular and nuclear membranes. Journal of controlled release : official journal of the Controlled Release Society, 132(3), pp.279-88. Available at: http://www.ncbi.nlm.nih.gov/pubmed/18655814 [Accessed April 7, 2012].

Abstract: After administration to the body, nucleic acid containing nanoparticles (NANs) need to cross several extra- and intracellular barriers to reach the cytoplasm or nucleus of the target cells. In the last decade several groups tried to overcome these barriers by arming non-viral delivery systems with targeting moieties, polyethylene glycol chains, fusogenic peptides and so forth. However, the drawback of this upgrading strategy is that each of the encountered barriers requires a new functionality, leading to very complex multi-component NANs. Moreover, there are currently no components available that can efficiently transport genes or NANs inside the nucleus of non-dividing cells. In this article a new, ultrasound based delivery system that possesses the capacity to simultaneously overcome several key barriers in non-viral nucleic acid delivery is presented. Additionally, a small amphiphilic compound that induces nuclear uptake of plasmid DNA and enhances non-viral gene transfer is presented.

Li, W. & Szoka, F.C., 2007. Lipid-based nanoparticles for nucleic acid delivery. Pharmaceutical research, 24(3), pp.438-49. Available at: http://www.ncbi.nlm.nih.gov/pubmed/17252188 [Accessed March 6, 2012].

Abstract: Lipid-based colloidal particles have been extensively studied as systemic gene delivery carriers. The topic that we would like to emphasize is the formulation/assembly of lipid-based nanoparticles (NP) with diameter under 100 nm for delivering nucleic acid in vivo. NP are different from cationic lipid-nucleic acid complexes (lipoplexes) and are vesicles composed of lipids and encapsulated nucleic acids with a diameter less than 100 nm. The diameter of the NP is an important attribute to enable NP to overcome the various in vivo barriers for systemic gene delivery such as: the blood components, reticuloendothelial system (RES) uptake, tumor access, extracellular matrix components, and intracellular barriers. The major formulation factors that impact the diameter and encapsulation efficiency of DNA-containing NP include the lipid composition, nucleic acid to lipid ratio and formulation method. The particle assembly step is a critical one to make NP suitable for in vivo gene delivery. NP are often prepared using a dialysis method either from an aqueous-detergent or aqueous-organic solvent mixture. The resulting particles have diameters about 100 nm and nucleic acid encapsulation ratios are >80%. Additional components can then be added to the particle after it is formed. This ordered assembly strategy enables one to optimize the particle physico-chemical attributes to devise a biocompatible particle with increased gene transfer efficacy in vivo. The components included in the sequentially assembled NP include: poly(ethylene glycol) (PEG)-shielding to improve the particle pharmacokinetic behavior, a targeting ligand to facilitate the particle-cell recognition and in some case a bioresponsive lipid or pH-triggered polymer to enhance nucleic acid release and intracellular trafficking. A number of groups have observed that a PEG-shielded NP is a robust and modestly effective system for systemic gene or small interfering RNA (siRNA) delivery.

Li, W. & Szoka, F.C., 2007. Lipid-based nanoparticles for nucleic acid delivery. Pharmaceutical research, 24(3), pp.438-49. Available at: http://www.springerlink.com/content/x74478m75m165672/ [Accessed March 6, 2012].

M.N.V. Ravi Kumar et al., 2004. Cationic Silica Nanoparticles as Gene Carriers: Synthesis, Characterization and Transfection Efficiency In vitro and In vivo. Journal of Nanoscience and Nanotechnology, 4(7), p.6. Available at: http://www.ingentaconnect.com/content/asp/jnn/2004/00000004/00000007/art00031 [Accessed April 7, 2012].

Abstract: The potential of cationic SiO2 nanoparticles was investigated for in vivo gene transfer in this study. Cationic SiO2 nanoparticles with surface modification were generated using amino-hexyl-aminopropyltri-methoxysilane (AHAPS). The zeta potential of the nanoparticles at pH = 7.4 varied from -31.4 mV (unmodified particles; 10 nm) to +9.6 mV (modified by AHAPS). Complete immobilization of DNA at the nanoparticle surface was achieved at a particle ratio of 80 (w/w nanoparticle/DNA ratio). The surface modified nanoparticle had a size of 42 nm with a distribution from 10–100 nm. The ability of these particles to transfect pCMV reporter gene was tested in Cos1 cells, and optimum results were obtained in the presence of FCS and chloroquine at a particle ratio of 80. These nanoparticles were tested for their ability to transfer genes in vivo in the mouse lung, and a two-times increase in the expression levels was found with silica particles in comparison to EGFP alone. Very low or no cell toxicity was observed, suggesting silica nanoparticles as potential alternatives for gene transfection.

Malhotra, M. et al., 2009. Ultrafine chitosan nanoparticles as an efficient nucleic acid delivery system targeting neuronal cells. Drug development and industrial pharmacy, 35(6), pp.719-26. Available at: http://www.ncbi.nlm.nih.gov/pubmed/19514987 [Accessed April 7, 2012].

Abstract: Cell transfection with nanoscaled cationic polymeric particles using Chitosan has been extensively explored. Because of its properties such as cationic charges, biocompatibility, biodegradability, and low toxicity, it has been used as a potential gene, siRNA, protein (including antibodies), and drug carrier system.

Mao, H.-Q. et al., 2001. Chitosan-DNA nanoparticles as gene carriers: synthesis, characterization and transfection efficiency. Journal of Controlled Release, 70(3), pp.399-421. Available at: http://dx.doi.org/10.1016/S0168-3659(00)00361-8 [Accessed April 7, 2012].

Abstract: Chitosan-DNA nanoparticles were prepared using a complex coacervation process. The important parameters for the nanoparticle synthesis were investigated, including the concentrations of DNA, chitosan and sodium sulfate, temperature of the solutions, pH of the buffer, and molecular weights of chitosan and DNA. At an amino group to phosphate group ratio (N/P ratio) between 3 and 8 and a chitosan concentration of 100 μg/ml, the size of particles was optimized to ∼100–250 nm with a narrow distribution, with a composition of 35.6 and 64.4% by weight for DNA and chitosan, respectively. The surface charge of these particles was slightly positive with a zeta potential of +12 to +18 mV at pH lower than 6.0, and became nearly neutral at pH 7.2. The chitosan-DNA nanoparticles could partially protect the encapsulated plasmid DNA from nuclease degradation as shown by electrophoretic mobility analysis. The transfection efficiency of chitosan-DNA nanoparticles was cell-type dependent. Typically, it was three to four orders of magnitude, in relative light units, higher than background level in HEK293 cells, and two to ten times lower than that achieved by Lipofectamine^TM-DNA complexes. The presence of 10% fetal bovine serum did not interfere with their transfection ability. Chloroquine could be co-encapsulated in the nanoparticles at 5.2%, but with negligible enhancement effect despite the fact that chitosan only showed limited buffering capacity compared with PEI. The present study also developed three different schemes to conjugate transferrin or KNOB protein to the nanoparticle surface. The transferrin conjugation only yielded a maximum of four-fold increase in their transfection efficiency in HEK293 cells and HeLa cells, whereas KNOB conjugated nanoparticles could improve gene expression level in HeLa cells by 130-fold. Conjugation of PEG on the nanoparticles allowed lyophilization without aggregation, and without loss of bioactivity for at least 1 month in storage. The clearance of the PEGylated nanoparticles in mice following intravenous administration was slower than unmodified nanoparticles at 15 min, and with higher depositions in kidney and liver. However, no difference was observed at the 1-h time point.

McMahon, K.M. et al., 2011. Biomimetic high density lipoprotein nanoparticles for nucleic acid delivery. Nano letters, 11(3), pp.1208-14. Available at: http://www.ncbi.nlm.nih.gov/pubmed/21319839 [Accessed March 30, 2012].

Abstract: We report a gold nanoparticle-templated high density lipoprotein (HDL AuNP) platform for gene therapy that combines lipid-based nucleic acid transfection strategies with HDL biomimicry. For proof-of-concept, HDL AuNPs are shown to adsorb antisense cholesterylated DNA. The conjugates are internalized by human cells, can be tracked within cells using transmission electron microscopy, and regulate target gene expression. Overall, the ability to directly image the AuNP core within cells, the chemical tailorability of the HDL AuNP platform, and the potential for cell-specific targeting afforded by HDL biomimicry make this platform appealing for nucleic acid delivery.

Mozafari, M.R. & Omri, A., 2007. Importance of divalent cations in nanolipoplex gene delivery. Journal of pharmaceutical sciences, 96(8), pp.1955-66. Available at: http://www.ncbi.nlm.nih.gov/pubmed/17542023 [Accessed April 7, 2012].

Abstract: Gene therapy is a promising therapeutic strategy to combat genetic or acquired diseases at their root cause rather than just treating symptoms. It is well recognised that there is an urgent need for non-toxic and efficient gene delivery vectors to fully exploit the current potential of gene therapy in molecular medicine. Cell-specific targeting of bioactive nucleotides is a prerequisite to attain the concentration of nucleic acids required for therapeutic efficacy in the target tissue. Many metal ions such as Mg2+, Mn2+, Ba2+ and, most importantly, Ca2+ have been demonstrated to have significant roles in gene delivery. These inorganic cations show low toxicity, good biocompatibility and promise for controlled delivery properties, thus presenting a new alternative to toxic and immunogenic carriers. Recently, inorganic nanoparticles alone, or in combination with a colloidal particulate system such as nanoliposome, an advanced approach to gene delivery, were found to exert a positive effect on gene transfer. In this report, the role of the divalent cations in nucleic acid delivery, particularly with respect to the potential improvement of transfection efficiency of nanolipoplexes, is reviewed.

Prabha, S. et al., 2002. Size-dependency of nanoparticle-mediated gene transfection: studies with fractionated nanoparticles. International Journal of Pharmaceutics, 244(1-2), pp.105-115. Available at: http://dx.doi.org/10.1016/S0378-5173(02)00315-0 [Accessed April 7, 2012].

Abstract: Nanoparticles formulated from biodegradable polymers such as poly (lactic acid) and poly (d,l-lactide-co-glycolide) (PLGA) are being extensively investigated as non-viral gene delivery systems due to their sustained release characteristics and biocompatibility. PLGA nanoparticles for DNA delivery are mainly formulated using an emulsion-solvent evaporation technique. However, this formulation procedure results in the formation of particles with heterogeneous size distribution. The objective of the present study was to determine the relative transfectivity of the smaller- and the larger-sized fractions of nanoparticles in cell culture. PLGA nanoparticles containing a plasmid DNA encoding luciferase protein as a marker were formulated by a multiple emulsion-solvent evaporation method (mean particle diameter=97±3 nm) and were fractionated using a membrane (pore size: 100 nm) filtration technique. The particles that passed through the membrane were designated as the smaller-sized nanoparticles (mean diameter=70±2 nm) and the fraction that was retained on the membrane as the larger-sized nanoparticles (mean diameter=202±9 nm). The smaller-sized nanoparticles showed a 27-fold higher transfection than the larger-sized nanoparticles in COS-7 cell line and a 4-fold higher transfection in HEK-293 cell line. The surface charge (zeta potential), cellular uptake, and the DNA release were almost similar for the two fractions of nanoparticles, suggesting that some other yet unknown factor(s) is responsible for the observed differences in the transfection levels. The results suggest that the particle size is an important factor, and that the smaller-sized fraction of the nanoparticle formulation predominantly contributes towards their transfection.

Ravi Kumar, M. et al., 2004. Nanoparticle-mediated gene delivery: state of the art. Expert opinion on biological therapy, 4(8), pp.1213-24. Available at: http://informahealthcare.com/doi/abs/10.1517/14712598.4.8.1213 [Accessed April 7, 2012].

Abstract: With the development of genomic and proteomic technologies, the prospect for gene therapy has progressed rapidly. This has been partly possible due to the emergence of a diverse array of polymeric and non-polymeric nanoparticles that are being investigated for their ability to deliver genes and drugs. In this review, particles have been pragmatically divided as chitosan-related and chitosan-unrelated nanomaterials. The state of the art in terms of the development, characterisation and evaluation of their in vitro and/or in vivo potential is discussed for each of these various particles. Although substantial progress has been made, the potential of these particles in the clinical arena and human responses remain to be evaluated. It is hoped that this review will provide an impetus for further studies of these particles, with the ultimate intent that one or more of these diverse nanoparticle-based non-viral approaches for gene transfer will translate from “bench to bedside” in the future.

Sandhu, K.K. et al., 2002. Gold Nanoparticle-Mediated Transfection of Mammalian Cells. Bioconjugate Chemistry, 13(1), pp.3-6. Available at: http://dx.doi.org/10.1021/bc015545c [Accessed April 7, 2012].

Abstract: Mixed monolayer protected gold clusters (MMPCs) functionalized with quaternary ammonium chains efficiently transfect mammalian cell cultures, as determined through ?-galactosidase transfer and activity. The success of these transfection assemblies depended on several variables, including the ratio of DNA to nanoparticle during the incubation period, the number of charged substituents in the monolayer core, and the hydrophobic packing surrounding these amines. Complexes of MMPCs and plasmid DNA formed at w/w ratios of 30 were most effective in promoting transfection of 293T cells in the presence of 10% serum and 100 ?M chloroquine. The most efficient nanoparticle studied (MMPC 7) was ?8-fold more effective than 60 kDa polyethylenimine, a widely used transfection agent. Mixed monolayer protected gold clusters (MMPCs) functionalized with quaternary ammonium chains efficiently transfect mammalian cell cultures, as determined through ?-galactosidase transfer and activity. The success of these transfection assemblies depended on several variables, including the ratio of DNA to nanoparticle during the incubation period, the number of charged substituents in the monolayer core, and the hydrophobic packing surrounding these amines. Complexes of MMPCs and plasmid DNA formed at w/w ratios of 30 were most effective in promoting transfection of 293T cells in the presence of 10% serum and 100 ?M chloroquine. The most efficient nanoparticle studied (MMPC 7) was ?8-fold more effective than 60 kDa polyethylenimine, a widely used transfection agent.

Schiffelers, R.M. et al., 2004. Cancer siRNA therapy by tumor selective delivery with ligand-targeted sterically stabilized nanoparticle. Nucleic acids research, 32(19), p.e149. Available at: http://nar.oxfordjournals.org/cgi/content/abstract/32/19/e149 [Accessed March 13, 2012].

Abstract: Potent sequence selective gene inhibition by siRNA “targeted” therapeutics promises the ultimate level of specificity, but siRNA therapeutics is hindered by poor intracellular uptake, limited blood stability and non-specific immune stimulation. To address these problems, ligand-targeted, sterically stabilized nanoparticles have been adapted for siRNA. Self-assembling nanoparticles with siRNA were constructed with polyethyleneimine (PEI) that is PEGylated with an Arg-Gly-Asp (RGD) peptide ligand attached at the distal end of the polyethylene glycol (PEG), as a means to target tumor neovasculature expressing integrins and used to deliver siRNA inhibiting vascular endothelial growth factor receptor-2 (VEGF R2) expression and thereby tumor angiogenesis. Cell delivery and activity of PEGylated PEI was found to be siRNA sequence specific and depend on the presence of peptide ligand and could be competed by free peptide. Intravenous administration into tumor-bearing mice gave selective tumor uptake, siRNA sequence-specific inhibition of protein expression within the tumor and inhibition of both tumor angiogenesis and growth rate. The results suggest achievement of two levels of targeting: tumor tissue selective delivery via the nanoparticle ligand and gene pathway selectivity via the siRNA oligonucleotide. This opens the door for better targeted therapeutics with both tissue and gene selectivity, also to improve targeted therapies with less than ideal therapeutic targets.

Sokolova, V.V. et al., 2006. Effective transfection of cells with multi-shell calcium phosphate-DNA nanoparticles. Biomaterials, 27(16), pp.3147-53. Available at: http://dx.doi.org/10.1016/j.biomaterials.2005.12.030 [Accessed April 7, 2012].

Abstract: Coated calcium phosphate nanoparticles were prepared for cell transfection. A calcium phosphate nanoparticle served as core which was then coated with DNA for colloidal stabilisation. The efficiency of transfection could be considerably increased by adding another layer of calcium phosphate on the surface, thereby incorporating DNA into the particle and preventing its degradation within the cell by lysosomes. A subsequent outermost layer of DNA on the calcium phosphate gave a colloidal stabilisation. The efficiency of such multi-shell particles was significantly higher than that of simple DNA-coated calcium phosphate nanoparticles. The transfection efficiency of EGFP-encoding DNA was tested with different cell lines (T-HUVEC, HeLa, and LTK). The dispersions were stable and could be used for transfection after 2 weeks of storage at 4 degrees C without loss of efficiency.

Xu, R., Wang, X.-L. & Lu, Z.-R., 2010. New amphiphilic carriers forming pH-sensitive nanoparticles for nucleic acid delivery. Langmuir : the ACS journal of surfaces and colloids, 26(17), pp.13874-82. Available at: http://www.ncbi.nlm.nih.gov/pubmed/20672851 [Accessed April 7, 2012].

Abstract: Amphiphilic lipids are promising for efficient intracellular delivery of nucleic acids. In this study, two new amphiphilic carriers, EKHCO and EHHKCO, were designed and synthesized as multifunctional carriers for efficient intracellular delivery of nucleic acids. The critical micelle concentrations of EKHCO and EHHKCO were 9.50 and 6.87 microM, respectively. Dynamic light scattering showed that the surfactants complexed with plasmid DNA and siRNA to form stable nanoparticles at the concentrations below their critical micelle concentrations. The nanoparticles of the surfactants with pDNA and siRNA exhibited pH-sensitive hemolysis against rat red blood cells when the pH decreased from 7.4 to 5.5, the endosomal-lysosomal pH. The nanoparticles of EHHKCO showed more concentration-dependent pH sensitivity than those of EKHCO. The EHHKCO and EKHCO nanoparticles of both pNDA and siRNA exhibited low cytotoxicity of at physiological pH. Both EKHCO and EHHKCO resulted in high intracellular uptake of pDNA and siRNA. EKHCO and EHHKCO resulted in relatively lower luciferase expression efficiency in U87 cells than DOTAP but produced a much higher percentage of GFP expression in the transfected cells than DOTAP. Both EKHCO and EHHKCO mediated much higher gene silencing efficiency of luciferase and green fluorescence protein (GFP) than DOTAP. The surfactants were more effective for intracellular siRNA delivery than intracellular delivery of plasmid DNA. The pH-sensitive amphiphilic carriers are promising multifunctional carriers for intracellular delivery of nucleic acids.

Zhang, L. et al., 2010. Development of nanoparticles for antimicrobial drug delivery. Current medicinal chemistry, 17(6), pp.585-94. Available at: http://www.ncbi.nlm.nih.gov/pubmed/20015030.

Abstract: This review focuses on the development of nanoparticle systems for antimicrobial drug delivery. Numerous antimicrobial drugs have been prescribed to kill or inhibit the growth of microbes such as bacteria, fungi and viruses. Even though the therapeutic efficacy of these drugs has been well established, inefficient delivery could result in inadequate therapeutic index and local and systemic side effects including cutaneous irritation, peeling, scaling and gut flora reduction. Nanostructured biomaterials, nanoparticles in particular, have unique physicochemical properties such as ultra small and controllable size, large surface area to mass ratio, high reactivity, and functionalizable structure. These properties can be applied to facilitate the administration of antimicrobial drugs, thereby overcoming some of the limitations in traditional antimicrobial therapeutics. In recent years, encapsulation of antimicrobial drugs in nanoparticle systems has emerged as an innovative and promising alternative that enhances therapeutic effectiveness and minimizes undesirable side effects of the drugs. Here the current progress and challenges in synthesizing nanoparticle platforms for delivering various antimicrobial drugs are reviewed. We also call attention to the need to unite the shared interest between nanoengineers and microbiologists in developing nanotechnology for the treatment of microbial diseases.

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