共查询到20条相似文献,搜索用时 15 毫秒
1.
《Nanomedicine : nanotechnology, biology, and medicine》2014,10(1):35-44
The cationizable nature of ‘proton-sponge’ transfection agents facilitates pDNA delivery in several steps. Protonated amines account for electrostatic DNA binding and cellular uptake, buffering amines mediate polyplex escape from acidifying intracellular vesicles. As demonstrated with a sequence-defined library of oligo(ethanamino)amides containing selected oligoethanamino acids and histidines, the total protonation capacity as well as the cationization pH profile within the endolysosomal range have critical impact on gene transfer. Building blocks with even numbered amine groups (Gtt, Sph) exhibited higher total endolysosomal buffer capacity than odd number (Stp) analogs. Within the endolysosomal range, Gtt has the highest buffer capacity around pH 5, whereas Stp has its maximum around pH 7. Histidines increased the total buffer capacity, resulted in a more continuous cationization pH profile and greatly improved transgene expression in vitro and in vivo. Using receptor targeted and polyethylene glycol shielded polyplexes, better endosomal escape and > 100-fold enhanced transfection was detected.From the Clinical EditorProton-sponge transfection agents for pDNA delivery are characterized in this study, demonstrating over 100-fold enhanced transection and better endosomal escape by using receptor targeted and polyethylene glycol shielded polyplexes. 相似文献
2.
Julia Kloeckner Lina Prasmickaite Anders HØgset Kristian Berg 《Journal of drug targeting》2013,21(4):205-213
Epidermal growth factor receptor (EGFR) targeted DNA polyplexes, containing polyethylenimine (PEI) conjugated with EGF protein as cell-binding ligand for endocytosis and polyethylene glycol (PEG) for masking the polyplex surface charge, mediated a 3- to 30-fold higher luciferase gene expression in HUH7, HepG2 and A431 cell transfections than analogous untargeted PEG–PEI polyplexes. Transfection levels can be further enhanced by treatment of cells with amphiphilic photosensitizers followed by illumination. In this process photosensitizers localized in membranes of endocytic vesicles are activated by light, resulting in the destruction of endocytic membrane structures and releasing co-endocytosed polyplexes into the cell cytosol. Photochemical enhanced gene expression was observed in all cell lines, with the magnitude of enhancement depending on the particular PEI polyplex formulation and cell line, ranging between 2- and 600-fold. Importantly, improved gene transfer retained EGF receptor specificity, as demonstrated by comparison with ligand-free polyplexes and by receptor antibody or ligand competition experiments. These results suggest that this combined procedure enables a dual mode of targeting polyplexes: biological targeting via EGFR interaction, combined with physical targeting with light to direct a photochemical delivery of therapeutic genes to a desired location. 相似文献
3.
Epidermal growth factor receptor (EGFR) targeted DNA polyplexes, containing polyethylenimine (PEI) conjugated with EGF protein as cell-binding ligand for endocytosis and polyethylene glycol (PEG) for masking the polyplex surface charge, mediated a 3- to 30-fold higher luciferase gene expression in HUH7, HepG2 and A431 cell transfections than analogous untargeted PEG-PEI polyplexes. Transfection levels can be further enhanced by treatment of cells with amphiphilic photosensitizers followed by illumination. In this process photosensitizers localized in membranes of endocytic vesicles are activated by light, resulting in the destruction of endocytic membrane structures and releasing co-endocytosed polyplexes into the cell cytosol. Photochemical enhanced gene expression was observed in all cell lines, with the magnitude of enhancement depending on the particular PEI polyplex formulation and cell line, ranging between 2- and 600-fold. Importantly, improved gene transfer retained EGF receptor specificity, as demonstrated by comparison with ligand-free polyplexes and by receptor antibody or ligand competition experiments. These results suggest that this combined procedure enables a dual mode of targeting polyplexes: biological targeting via EGFR interaction, combined with physical targeting with light to direct a photochemical delivery of therapeutic genes to a desired location. 相似文献
4.
Polymer Side-Chain Degradation as a Tool to Control the Destabilization of Polyplexes 总被引:1,自引:1,他引:0
Funhoff AM van Nostrum CF Janssen AP Fens MH Crommelin DJ Hennink WE 《Pharmaceutical research》2004,21(1):170-176
PURPOSE: We purposed to design a cationic polymer that binds to pDNA to form polyplexes and that subsequently degrades within a few days at physiological pH and temperature, releasing the DNA in the cytosol of a cell. METHODS: We synthesized a new monomer carbonic acid 2-dimethylamino-ethyl ester 1-methyl-2-(2-methacryloylamino)-ethyl ester (abbreviated HPMA-DMAE) and the corresponding polymer. Hydrolysis of the carbonate ester of both the monomer and the polymer was investigated at 37 degrees C. The DNA condensing properties of the pHPMA-DMAE was studied using dynamic light scattering (DLS) and zeta potential measurements. Degradation of the polyplexes at 37 degrees C and pH 7.4 was monitored with DLS and gel electrophoresis. In vitro transfections were performed in COS-7 cell line. RESULTS: pHPMA-DMAE is able to condense DNA into small particles (110 nm) with a positive zeta potential. The half-life of the polymer and monomer at 37 degrees C and pH 7.4 was around 10 h whereas at pH 5, the half-life was 380 h. In line with this, due to hydrolysis of the side groups, pHPMA-DMAE-based polyplexes dramatically increased in size at 37 degrees C and pH 7.4 whereas at pH 5.0, only a very small increase was observed. Interestingly, intact DNA was released from the polyplexes after 48 h at pH 7.4 whereas all DNA remained bound to the polymer at pH 5.0. Polyplexes were able to transfect cells with minimal cytotoxicity if the endosomal membrane-disrupting peptide INF-7 was added to the polyplex formulation. CONCLUSIONS: Degradation of the cationic side-chains of a polymer is a new tool for time-controlled release of DNA from polyplexes, preferably within the cytosol and/or nucleus. 相似文献
5.
A series of cationic, methacrylamide polymers was tested for use as a biodegradable gene carrier in ovarian cancer. Tumor transfection activity of polyplexes consisting of a reporter gene and different methacrylamide polymers was assessed, after intraperitoneal injection in mice bearing an ovarian cancer xenograft. In this model, polyplexes based on poly(HPMA-DMAE) showed transfection activity similar to polyplexes based on the nondegradable and rather toxic polyethylenimine (PEI22). The tumor transfection activity of the pHPMA-DMAE polyplexes was remarkable considering their poor transfection activity in in vitro assays. Polyplexes based on pHPMA-DMAE were devoid of any cytotoxicity and mediated highest transfection activity at the highest N/P ratio investigated. Tumor cell gene expression after a single administration of these polyplexes rapidly declined within time, at a similar rate to that observed after injection with polyplexes based on PEI22. Incubation of the polyplexes with hyaluronic acid (HA), a polyanion accumulating in the ascitic fluid of ovarian cancer bearing mice, changed the physical characteristics of the pHPMA-DMAE and PEI22 polyplexes. The transfection activity of PEI22-based polyplexes, but not that of pHPMA-DMAE based polyplexes, was strongly impaired by HA. Differences in HA sensitivity might have contributed to the in vivo gene expression activities of pHPMA-DMAE- and PEI22-based polyplexes. pHPMA-DMAE-based polyplexes have potential for use in ovarian cancer therapy due to their considerable transfection activity, their low cytotoxicity, and their HA resistance. 相似文献
6.
de Wolf HK de Raad M Snel C van Steenbergen MJ Fens MH Storm G Hennink WE 《Pharmaceutical research》2007,24(8):1572-1580
Purpose Previously, we have shown that complexes of plasmid DNA with the biodegradable polymer poly(2-dimethylamino ethylamino)phosphazene
(p(DMAEA)-ppz) mediated tumor selective gene expression after intravenous administration in mice. In this study, we investigated
the effect of p(DMAEA)-ppz molecular weight on both in vitro and in vivo tumor transfection, as well as on complex induced toxicity.
Materials and Methods p(DMAEA)-ppz with a broad molar mass distribution was fractionated by preparative size exclusion chromatography. Polyplexes
consisting of plasmid DNA and the collected polymer fractions were tested for biophysical properties, (cyto)toxicity and transfection
activity.
Results Four p(DMAEA)-ppz fractions were collected with weight average molecular weights ranging from 130 to 950 kDa, and with narrow
molecular mass distributions (Mw/Mn from 1.1 to 1.3). At polymer-to-DNA (N/P) ratios above 6, polyplexes based on these polymers were all positively charged
(zeta potential 25–29 mV), and had a size of 80–90 nm. The in vitro cytotoxicity of the polyplexes positively correlated with polymer molecular weight. The in vitro transfection activity of the different polyplexes depended on their N/P ratio, and was affected by the degree of cytotoxicity,
as well as the colloidal stability of the different polyplexes. Intravenous administration of polyplexes based on the high
molecular weight polymers led to apparent toxicity, as a result of polyplex-induced erythrocyte aggregation. On the other
hand, administration of polyplexes based on low molecular weight p(DMAEA)-ppz’s (Mw 130 kDa) did not show signs of toxicity and resulted in tumor selective gene expression.
Conclusion Polymer molecular weight fractionation enabled us to optimize the transfection efficiency/toxicity ratio of p(DMAEA)-ppz polyplexes
for in vitro and in vivo tumor transfection. 相似文献
7.
Purpose Dynamic PEG-polycation copolymers that release PEG and degrade into small fragments after cell entry might present efficient
and biocompatible gene carriers.
Methods PEG-OEI-MK was synthesized by copolymerization of 5 kDa polyethyleneglycol (PEG) and 800 Da oligoethylenimines through acid-degradable
acetone-bis-(N-maleimidoethyl)ketal linkers (MK). To evaluate any benefit of the reversible over stable linkage, also the corresponding
pH-stable analog, PEG-OEI-BM, was synthesized via ether linkages. Luciferase and GFP expression plasmids were used for transfections, in vivo biocompatibility was evaluated by intravenous application of polymers in Balb/c mice.
Results PEG-OEI-MK showed efficient DNA binding as analyzed by ethidium bromide exclusion, resulting in formation of polyplexes with
sizes around 100 nm and surface charges of below 5 mV zeta potential. This surface shielding of PEG-OEI-MK polyplexes remained
stable at neutral pH 7.4, while polyplexes deshielded and aggregated at pH 5 within 15–30 min. Cell culture experiments demonstrated
reduced polymer toxicity compared to the non-PEGylated OEI-MK. Transfection experiments demonstrated reduced gene expression
of PEG-OEI-BM compared with the non-PEGylated analog OEI-BM, whereas the pH-reversible polymer PEG-OEI-MK mediated a significant
increased transfection efficiency over the non-PEGylated OEI-MK.
Conclusions PEG-OEI-MK mediated the highest gene transfer at lowest cytotoxicity levels and also best in vivo biocompatibility. 相似文献
8.
The clinical success of gene therapy intimately relies on the development of safe and efficient gene carrier systems. We found here that 1.8 kDa polyethylenimine (PEI) following hydrophobic modification with lipoic acid (LA) mediated nontoxic and highly potent in vitro gene transfection in both HeLa and 293T cells. 1.8 kDa PEI-LA conjugates were prepared with controlled degree of substitution (DS) by coupling LA to PEI using carbodiimide chemistry. Gel electrophoresis measurements showed that the DNA binding ability of 1.8 kDa PEI was impaired by lipoylation, in which an N/P ratio of 2/1 and 4-6/1 was required for 1.8 kDa PEI and 1.8 kDa PEI-LA conjugates, respectively, to completely inhibit DNA migration. Interestingly, dynamic light scattering measurements (DLS) revealed that PEI-LA conjugates condensed DNA into much smaller sizes (183-84 nm) than unmodified 1.8 kDa PEI (444-139 nm) at N/P ratios ranging from 20/1 to 60/1. These polyplexes revealed similar surface charges of ca. +22 to +30 mV. 1.8 kDa PEI-LA(2) polyplexes formed at an N/P ratio of 10/1 were stable against exchange with 12-fold excess of negatively charged dextran sodium sulfate (DSS) relative to DNA phosphate groups while 1.8 kDa PEI controls dissociated at 6-fold excess of DSS, indicating that lipoylation of 1.8 kDa PEI resulted in stronger binding with DNA. Importantly, DNA was released from 1.8 kDa PEI-LA(2) polyplexes upon addition of 10 mM dithiothreitol (DTT). Reduction-triggered unpacking of 1.8 kDa PEI-LA(2) polyplexes was also confirmed by DLS. MTT assays demonstrated that all PEI-LA conjugates and polyplexes were essentially nontoxic to HeLa and 293T cells up to a tested concentration of 50 μg/mL and an N/P ratio of 80/1, respectively. The in vitro gene transfection studies in HeLa and 293T cells showed that lipoylation of 1.8 kDa PEI markedly boosted its transfection activity. For example, 1.8 kDa PEI-LA(2) polyplexes displayed 400-fold and 500-fold higher levels of gene expression than unmodified 1.8 kDa PEI controls, which were ca. 2-fold and 3-fold higher than 25 kDa PEI controls, in serum-free and 10% serum media, respectively. The transfection efficiency decreased with increasing DS, following an order of 1.8 kDa PEI-LA(2) > 1.8 kDa PEI-LA(4) > 1.8 kDa PEI-LA(6) ? 1.8 kDa PEI. Confocal laser scanning microscopy (CLSM) studies corroborated that 1.8 kDa PEI-LA(2) delivered and released DNA into the nuclei of HeLa cells more efficiently than 25 kDa PEI. These nontoxic 1.8 kDa PEI-LA conjugates form a superb basis for the development of targeting, biocompatible and highly efficient carriers of gene delivery. 相似文献
9.
Zuidam NJ Posthuma G de Vries ET Crommelin DJ Hennink WE Storm G 《Journal of drug targeting》2000,8(1):51-66
The cationic polymer poly(2-(dimethylamino)ethyl methacrylate) (p(DMAEMA)) is able to efficiently bind and condense DNA and to mediate transfection of a variety of cell types. In this study, fluorescence activated cell sorting (FACS), confocal laser fluorescence microscopy (CSLM) and electron microscopy (EM) techniques were used to investigate in vitro the cellular interaction of p(DMAEMA)-based polyplexes with human ovarian carcinoma cells (OVCAR-3). Cellular association and subsequent internalization only occurred when the polyplexes exhibited a positive zeta potential. Small-sized polyplexes have an advantage over large-sized complexes regarding cellular entry. The effect of the presence of tertiary amine groups versus the presence of quatenary amine groups was evaluated by comparing p(DMAEMA) with its quaternary ammonium analogue poly(2-(trimethylamino)ethyl methacrylate) (p(TMAEMA)). The combined cellular interaction and transfection results suggest that the latter polymer does not have an intrinsic endosomal escape property, in contrast to the 'proton sponge' effect proposed for p(DMAEMA). PEGylation of p(DMAEMA) effectively shielded the surface charge and yielded a notably lower degree of cellular interaction. Data on the effects of the presence of endocytosis inhibitors and an endosome-disruptive peptide in the culture medium on the cellular interaction and transfection activity of p(DMAEMA)-based polyplexes support endocytosis as being the principal pathway for intracellular delivery of plasmid. Both the CLSM and EM studies did not reveal the presence of polyplexes or plasmid outside the endocytic vesicles or within the nucleus, suggesting that intracellular trafficking from the endosomes to the nucleus is a very inefficient process. 相似文献
10.
Aldawsari H Raj BS Edrada-Ebel R Blatchford DR Tate RJ Tetley L Dufès C 《Nanomedicine : nanotechnology, biology, and medicine》2011,7(5):615-623
The potential of gene therapy to treat cancer is currently limited by the low expression of therapeutic genes in the tumors. Because amino acids are known to have excellent properties in cell penetration and gene expression regulation, we investigated if the conjugation of arginine (Arg), lysine (Lys) and leucine (Leu) onto the surface of the gene delivery system polyethylenimine (PEI) could lead to an improved gene expression in tumors. The intravenous administration of Arg-, Lys- and Leu-bearing PEI polyplexes led to a significant increase of gene expression in the tumor, with a β-galactosidase expression amount at least threefold higher than that obtained after treatment with unmodified PEI polyplex. The three amino acid-bearing PEI polyplexes led to similar levels of gene expression in the tumor. The treatments were well tolerated by the mice. Arg-, Lys- and Leu-bearing PEI polyplexes are therefore highly promising gene delivery systems for cancer therapy.
From the Clinical Editor
In this paper, amino-acid based modulations of gene delivery enhancement are reported. Intravenous administration of Arg-, Lys- and Leu-bearing polyethylenimine polyplexes led to a significant increase of gene expression in the studied tumor model, which may enable the development of more efficient gene delivery strategies for future clinical applications. 相似文献11.
Ammar A. Y. Almulathanon Elisabetta Ranucci Paolo Ferruti Martin C. Garnett Cynthia Bosquillon 《Pharmaceutical research》2018,35(4):86
Purpose
This study aimed to further explore the mechanisms behind the ability of certain linear polyamidoamines (PAAs) to transfect cells with minimal cytotoxicity.Methods
The transfection efficiency of DNA complexed with a PAA of a molecular weight over 10 kDa or 25 kDa branched polyethyleneimine (BPEI) was compared in A549 cells using a luciferase reporter gene assay. The impact of endo/lysosomal escape on transgene expression was investigated by transfecting cells in presence of bafilomycin A1 or chloroquine. Cytotoxicity caused by the vectors was evaluated by measuring cell metabolic activity, lactate dehydrogenase release, formation of reactive oxygen species and changes in mitochondrial membrane potential.Results
The luciferase activity was ~3-fold lower after transfection with PAA polyplexes than with BPEI complexes at the optimal polymer to nucleotide ratio (RU:Nt). However, in contrast to BPEI vectors, PAA polyplexes caused negligible cytotoxic effects. The transfection efficiency of PAA polyplexes was significantly reduced in presence of bafilomycin A1 while chloroquine enhanced or decreased transgene expression depending on the RU:Nt.Conclusions
PAA polyplexes displayed a pH-dependent endo/lysosomal escape which was not associated with cytotoxic events, unlike observed with BPEI polyplexes. This is likely due to their greater interactions with biological membranes at acidic than neutral pH.12.
de Wolf HK Johansson N Thong AT Snel CJ Mastrobattista E Hennink WE Storm G 《Pharmaceutical research》2008,25(7):1654-1662
PURPOSE: Tumor gene expression after the intravenous (i.v.) administration of current polymer-based gene delivery systems is generally low and short-lived. Immune stimulatory CpG dinucleotides, present within the plasmid DNA of the polyplexes are likely to contribute to this. The effect of CpG replacement on the levels of transgene expression was studied, after the i.v. administration of polyethylenimine (PEI) polyplexes. METHODS: Tumor transfection and immune stimulation of PEI polyplexes containing plasmid DNA encoding for luciferase and rich in CpG motifs was monitored and compared to polyplexes containing the same gene but devoid of CpG motifs. Lipoplexes based on 1,2-dioleyl-3-trimethylammonium-propane/dioleoylphosphatidylethanolamine liposomes were included as a control. RESULTS: The replacement of CpGrich DNA by CpGfree DNA did neither affect the physical properties of the DNA complexes nor did it affect their in vitro transfection activity or cytotoxicity. The immune stimulation (interleukin-12) after i.v. administration of the PEI DNA complexes was low and unaffected by the presence of CpG motifs. The absence of CpG motifs within the different DNA complexes improved the degree and the duration of organ and tumor gene expression. CONCLUSION: The depletion of CpG dinucleotides within the plasmid DNA of polyplexes enhances the degree and duration of in vivo transgene expression. 相似文献
13.
非病毒基因载体聚乙二醇-聚乙烯亚胺共聚物缓冲容量的研究 总被引:1,自引:0,他引:1
目的考察非病毒基因载体PEI不同的分子量及接枝PEG的量对缓冲容量的影响。方法采用电位滴定的方法,根据公式β=dc(HCl)/dpH计算获得缓冲容量。结果分子量不同的PEI缓冲容量的最大值均〉8,且pH值随着分子量的增加而降低;同一分子量的PEI接枝不同量的PEG后,缓冲容量有明显的下降,接枝PEG量越多,缓冲容量下降越大。在生理pH4~7,缓冲容量差异减小。结论PEI分子量及接枝PEG的量对缓冲容量会产生一定影响.而缓冲容量又会影响PEI介导基因传递的能力。 相似文献
14.
The purpose of the present study was to investigate the influence of molecular weights on the chemical, biophysical, and biological properties of bioreducible oligoethylenimine conjugates. The cationic conjugates were synthesized by polyaddition between branched oligoethylenimine 800 Da (OEI) and the disulfide bond containing N,N′-cystamine bisacrylamide (CBA) linker. A correlation between the copolymer molecular weights and the polyplex transfection properties was found. The OEI–CBA copolymers differing in molecular weights (from 8.6 to 37 kDa) showed good plasmid DNA binding ability resulting in compact 90- to 150-nm-sized polyplexes. Colloidal stability of the polyplexes was lost in reductive environment. A low concentration of dithiothreitol of 5 µM was sufficient to render polyplexes unstable in size. Reducing conditions at physiological salt concentration triggered polyplex dissociation. The bioreducible polymers displayed much lower cytotoxicity (IC50 ∼ 100 μg/mL in cell culture) than branched polyethylenimine 25 kDa (BPEI) and linear polyethylenimine 22 kDa (LPEI). Reporter gene transfection experiments were done with CHO-K1 and B16-F10 cells. The largest (37 kDa) copolymer HC-6-8 demonstrated highest transfection levels among all the bioreducible copolymers, which was comparable with LPEI and much more effective than BPEI.Key words: bioreducible, gene delivery, PEI, polyplexes, synthetic vectors 相似文献
15.
Amir K. VarkouhiTwan Lammers Raymond M. SchiffelersMies J. van Steenbergen Wim E. HenninkGert Storm 《European journal of pharmaceutics and biopharmaceutics》2011,77(3):450-457
Cationic polymers are used as non-viral vectors for nucleic acid delivery. In this study, two biodegradable cationic polymers were evaluated for the purpose of siRNA delivery: pHPMA-MPPM (poly((2-hydroxypropyl) methacrylamide 1-methyl-2-piperidine methanol)) and TMC (O-methyl-free N,N,N-trimethylated chitosan). The silencing activity and the cellular cytotoxicity of polyplexes based on these biodegradable polymers were compared with those based on non-biodegradable pDMAEMA (poly(2-dimethylamino)ethyl methacrylate) and PEI (polyethylenimine) and with the regularly used lipidic transfection agent Lipofectamine. To promote endosomal escape, either the endosomolytic peptide diINF-7 was added to the formulations or photochemical internalization (PCI) was applied. Incubation of H1299 human lung cancer cells expressing firefly luciferase with polyplexes based on pHPMA-MPPM and TMC showed 30-40% silencing efficiency. This silencing activity was equal to or better than that obtained with the standard transfectants. Under all experimental conditions tested, the cytotoxicity of the biodegradable polymers was low. The application of PCI, as well as the addition of the diINF-7 peptide to the formulations increased their silencing activity up to 70-80%. This demonstrates that pHPMA-MPPM- and TMC-based polyplexes benefit substantially from endosomal escape enhancement. Importantly, the polyplexes retained their silencing activity in the presence of serum, and they showed low cytotoxicity. These biodegradable vectors are therefore attractive systems for further in vivo evaluations. 相似文献
16.
With the aim to improve the specificity and to reduce the cytotoxicity of polyethylenimine (PEI), we have synthesized the conjugates of the branched PEI (25 kDa) with transferrin. The transferrin-PEI (TP) conjugates with five compositions were synthesized using periodate oxidation method and confirmed by FT-IR spectroscopy and gel permeation chromatography. The free amine contents of TP conjugates, which were able to condense and deliver DNA, increased as the amount of PEI increased. TP/DNA polyplexes were characterized by measuring gel electrophoresis, ethidium bromide fluorescence quenching, particle size and zeta potential of complexes. Complete complexation of the polyplexes was observed above the N/P ratio of 5 in TP/ DNA, and above 3 in PEI/DNA, respectively. The zeta potential of the complexes decreased as the amount of transferrin in TP conjugates increased. Transfection efficiency of TP conjugates was evaluated in HeLa cell and Jurkat cell systems. Among the five compositions of TP conjugates, TP-2 system mediated a higher beta-galactosidase gene expression than PEI system in Jurkat cell which was known to express elevated numbers of transferrin receptors. From the results of the cell viability based on MTT assay, TP conjugates showed lower cytotoxicity compared with the PEI system. We expect that the TP conjugate can be used efficiently as a nonviral gene delivery vector. 相似文献
17.
Elke Kleemann Norman Jekel Lea Ann Dailey Susanne Roesler Ludger Fink Norbert Weissmann 《Journal of drug targeting》2013,21(8):638-651
The aim of this study was to elicit improved gene expression and decreased cytotoxicity for pulmonary gene therapy by replacing the commonly used carrier 25?kDa branched poly(ethylene imine) (BPEI) by two PEI derivatives, low-molecular-weight PEI (LMWPEI) and polyethylene glycol?grafted PEI (PEGPEI). All polymers were shown to condense DNA to spherical particles of approximately 100?nm. Biocompatibility was investigated in vitro and in vivo. Although transfection was less efficient with LMWPEI-DNA in vitro, this polyplex caused the highest luciferase expression in the mouse lung after intratracheal instillation. While PEGPEI luciferase expression in vitro was approximately three times higher when compared to BPEI, a transfection rate at the level of naked DNA was observed in vivo. LMWPEI polyplexes were located in both the bronchial and alveolar cells, whereas BPEI polyplexes were mainly detected in bronchial cells. LMWPEI combines low cytotoxicity with high transfection efficiency in the mouse lung in vivo, rendering it a promising strategy for pulmonary gene delivery. 相似文献
18.
Ferry Verbaan Inger van Dam Yoshinubu Takakura Mitsuru Hashida Wim Hennink Gert Storm Christien Oussoren 《European journal of pharmaceutical sciences》2003,20(4-5):419-427
The objective of this study was to assess the in vivo fate of poly(2-(dimethylamino)ethyl methacrylate) (pDMAEMA)-based polyplexes after intravenous administration into mice. Circulation kinetics and tissue distribution in terms of plasmid localization and transfection efficiency were assessed. To gain more insight into the observed biodistribution and gene expression profile, the interaction of pDMAEMA-based polyplexes with blood components (erythrocytes and albumin) was investigated in vitro. In the case of i.v. injection of positively charged polyplexes at a dose of 30 microg DNA most of the radioactivity was found in the lungs and the liver 60 min after injection. In the case of pDMAEMA/DNA polyplexes with a negative charge, uptake occurred mainly by the liver. Administration of positively charged complexes at a 30 microg DNA dose resulted in reporter gene expression primarily in the lungs. Injection of negatively charged complexes and naked plasmid did not result in luciferase expression in any of the organs examined. In vitro turbidity experiments showed the induction of a charge dependent aggregation process upon addition of albumin to the polyplexes pointing out to the involvement of aggregate formation in the dominant lung uptake of the positively charged polyplexes. Also, incubations of polyplexes after pre-incubation with a physiological concentration of albumin with washed erythrocytes confirmed that polyplexes induce the formation of extremely large structures. This paper underlines the need for the design of systems with reduced interaction with blood components to promote the delivery of DNA to target tissues outside the lungs. 相似文献
19.
We previously reported on a stearylated INF7 peptide (str-INF7), which enhances the endosomal escape of an octaarginine (R8)-modified liposomal particle encapsulating plasmid DNA (pDNA) in a fusion-independent manner. This study examined whether this peptide derivative enhanced the endosomal escape and gene expression of PEGylated liposomes encapsulating pDNA. We used a PEGylated, R8-modified multifunctional envelope-type nanodevice (R8-MEND) as a model for PEGylated liposomes. Polyethylene glycol 2000 (PEG2000) attached to two different anchors, distearoylphosphatidylethanolamine (DSPE-PEG) or dimyristoylphosphatidylethanolamine (DMPE-PEG), was used to modify the R8-MEND in the presence or absence of two different concentrations of str-INF7. Modification of the PEGylated R8-MEND with str-INF7 resulted in luciferase gene expression levels in HeLa cells that were 73-fold and 24-fold higher than the corresponding value for an unmodified MEND in the case of DSPE-PEG and DMPE-PEG, respectively. The endosomal escape of the PEGylated R8-MEND was improved by str-INF7, as confirmed by confocal laser scanning microscopy. Furthermore, modification with str-INF7 enhanced the hepatic gene expression of the R8-MEND modified with DSPE-PEG and DMPE-PEG by 95-fold and 1885-fold, respectively, after intravenous injection in mice. Collectively, these data demonstrate that str-INF7 can be a useful device for enhancing the endosomal escape even for PEGylated liposomes encapsulating pDNA. 相似文献
20.
Yanjie Bao Yi Jin Padmanabh Chivukula Jun Zhang Yun Liu Jian Liu Jean-Pierre Clamme Ram I. Mahato Dominic Ng Wenbin Ying Yiting Wang Lei Yu 《Pharmaceutical research》2013,30(2):342-351