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Cherng, Jong Yuh; Wetering, Petra van de; Talsma, H.; Crommelin, Daan J.A.; Hennink, Wim E.

doi:10.1023/a:1016054623543

Cited by 319 publications

(170 citation statements)

References 14 publications

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“…[31] The large aggregates of the complexes of DNA and PL were obtained at the C/A ratio of 3, while PtmL allowed the formation of nonaggregating complexes at the same C/A ratio (Figure 3). The quaternary ammonium groups seem to enhance both the electrostatic and sterical repulsion of complexes, and prevent the aggregation of the complexes at the same C/A ratio.…”

Section: Discussionmentioning

confidence: 97%

Effect of Physicochemical Properties of Polyplexes Composed of Chemically Modified PL Derivatives on Transfection Efficiency In Vitro

Kimura

Yamaoka

Iwase

et al. 2002

Macromolecular Bioscience

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Chemically modified poly‐L‐lysine (PL) derivatives with two essential features, which we have previously reported, have been used to examine the effect of physicochemical properties of their complexes with plasmid DNA on expression efficacy in in vitro gene transfection by osmotic shock procedure. PL derivatives having either Nε‐trimethyl lysine or 13.9% serine residue showed no transfection efficiency. Only when PL was modified in both ways, giving Nε‐trimethyl poly(L‐lysine‐co‐L‐serine), markedly enhanced gene expression was observed. The cellular uptake and localization of DNA in the cells were similar for each cationic polypeptide. The trimethylated polypeptide formed the loosest complexes of DNA among all of the polypeptides used, judging from the loss of EtBr fluorescence. It resulted in more effective DNA release from the complexes by adding heparin, and in the significant gene expression in in vitro transcription/translation systems, as compared with the other polypeptides. The gene transfection efficiency was found to depend on the physicochemical properties of the complexes, such as absence of aggregation, moderate compaction, and easy dissociation, which allow the efficient transcription.Transfection efficiency of pEGFP‐N1 complexes with (○) PL, (•) PLS, (□) PtmL, and (▪) PtmLS at various C/A ratios.magnified imageTransfection efficiency of pEGFP‐N1 complexes with (○) PL, (•) PLS, (□) PtmL, and (▪) PtmLS at various C/A ratios.

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Section: Discussionmentioning

confidence: 97%

Effect of Physicochemical Properties of Polyplexes Composed of Chemically Modified PL Derivatives on Transfection Efficiency In Vitro

Kimura

Yamaoka

Iwase

et al. 2002

Macromolecular Bioscience

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“…Since positively charged complexes interact with the cell surface through an electrostatic interaction with anionic substances on the cell surface, internalization events are affected by the zeta-potential of the polyplexes. It has been reported that polyplexes composed of polycations with MWs of several thousands exhibit high zeta-potentials [13,[26][27][28][29][30], irrespective of the polycations used. PK and PR are known to form polyplexes with pDNA in a different manner [31].…”

Section: Discussionmentioning

confidence: 99%

Quantitative comparison between poly(L-arginine) and poly(L-lysine) at each step of polyplex-based gene transfection using a microinjection technique

Hashimoto

Kawazu

Nagasaki

et al. 2012

Science and Technology of Advanced Materials

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Among the well-studied polypeptide-type gene carriers, transfection efficiency is empirically known to be higher for poly(L-arginine) (PR) than poly(L-lysine) (PK). The big difference between PR and PK should be determined at one of the intracellular trafficking steps based on the different charge densities, structures or PKa values. However, the endosomal escape and the intranuclear transcription efficiency in living cells have not been clarified yet. In this study, a novel method for quantifying the intranuclear transcription efficiency and the nuclear transport of the polyplex is established based on the nuclear and the cytosolic microinjection technique, and the results for PK and PR with different molecular weights (MWs) are compared in living cells. The intranuclear transcription efficiency is the same in PR and PK and it decreases rapidly with increasing MW, in spite of the commonly measured transfection efficiency. The transcription efficiency is strongly suppressed at high MW and strongly correlates with the polyplex forming ability expressed as a critical ratio of the number of polypeptide cationic groups to the number of pDNA anionic groups. When considered with the results of the cellular uptake and in vitro transfection with or without chloroquine, the rate-limiting step for their gene transfer is the buffering effect-independent endosomal escape.

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“…Nevertheless, it can be concluded that cellular uptake occurs on 293T cells since luciferase was detected after transfection with p(DMAPEMA) complexes in the presence of endosome disruptive agent (chloroquine). [3] The performances of three polymers (Exgen 500, p(DMAEMA) and p(DMAPEMA)) were confirmed by transfection with a pEGFP plasmid on BHK21 cells and analysis by flow cytometry. This method is more straightforward for determining transfection efficiency.…”

Section: In Vitro Transfection Studiesmentioning

confidence: 95%

“…The hydrolysis ratio of the polymers was determined by integration of 1 H NMR spectra ( Figure 2). The integral between 3.6 and 4.5 ppm, corresponding to the protons H4 (3) , H5 (3) , H4 0 (4) and H5 0 (4) , was normalized to a value of 4, then this area was split into two regions: 3.6-3.9 ppm (a-alcoholic methylene H5 0 (4) ) and 3.9-4.5 ppm (methylenes H4 (3) , H5 (3) and H4 0 (4) ). The hydrolysis ratio was calculated according to Equation (1).…”

Section: Kinetics Of Hydrolysismentioning

confidence: 99%

“…Better transfection results were obtained with polyplexes which were able to be dissociated by electrostatic interactions rather than ones which DNAse digestion has been well established with several polycations, and in particular with p(DMAEMA). [7] It has also been shown that the toxicity of p(DMAEMA)-based polyplexes is negligible at the concentration corresponding to the maximum efficiency, [3] whereas p(DMAEMA) alone is seriously cytotoxic. [8] In several studies, p(DMAEMA) was found to be an efficient transfecting agent for different cell lines (OVCAR-3, [7] COS-7, [3] BHK21 [9] ) though the mechanism of transfection is not still well understood.…”

Section: Introductionmentioning

confidence: 99%

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Hydrolyzable p(DMAPEMA) Polymers for Gene Delivery

Véron

Ganée

Ladavière

et al. 2006

Macromolecular Bioscience

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The efficiency of cationic polymers as transfectants is thought to be closely related to their DNA association/dissociation properties. An incomplete polymer-DNA dissociation could explain the relatively low gene expression obtained with p(DMAEMA) polymers. Our approach was to synthesize a p(DMAEMA) analogue, p(DMAPEMA), bearing an hydrolyzable cationic group incorporated into the pendant chain with a view to improving transfection. The complexation of DNA with both polymers was studied by agarose gel electrophoresis, size and zeta potential measurements, as well as the dissociation of the polyplexes, after treatment by an anionic polymer, sodium hydroxide or heat. The transfection efficiencies of the polyplexes were evaluated with 293T and BHK21 cells in comparison with Exgen 500. P(DMAPEMA) polymers were able to complex DNA and to release it in a free intact form after an alkaline treatment or storage at 37 degrees C. Poly(aspartic acid) was unable to dissociate p(DMAPEMA) based polyplexes, in contrast to p(DMAEMA) ones. No transfection was obtained with p(DMAPEMA) with both cell lines. A slow hydrolysis under physiological conditions resulting in the absence of DNA unpacking or endosomal entrapment could explain these results. Better transfection results were obtained with polyplexes which were able to be dissociated by electrostatic interactions rather than ones which required the hydrolysis mechanism to release free DNA into cells. Scheme of hydrolyzable p(DMAPEMA) polymer.

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Abstract: PDMAEMA is an interesting vector for the design of in vivo and ex vivo gene transfection systems.

Cited by 319 publications

References 14 publications

Effect of Physicochemical Properties of Polyplexes Composed of Chemically Modified PL Derivatives on Transfection Efficiency In Vitro

Effect of Physicochemical Properties of Polyplexes Composed of Chemically Modified PL Derivatives on Transfection Efficiency In Vitro

Quantitative comparison between poly(L-arginine) and poly(L-lysine) at each step of polyplex-based gene transfection using a microinjection technique

Hydrolyzable p(DMAPEMA) Polymers for Gene Delivery

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