Ferrocene-Containing Cationic Lipids: Influence of Redox State on Cell Transfection

Abbott, Nicholas L.; Jewell, Christopher M.; Hays, Melissa E.; Kondo, Yukishige; Lynn, David M.

doi:10.1021/ja054038t

Cited by 63 publications

(127 citation statements)

References 17 publications

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“…This lipid can be reversibly cycled between its reduced state (net charge of +1) and its oxidized state (net charge of +3) by either oxidation or reduction of the ferrocene groups present at the end of each hydrophobic chain [35–41]. Our past studies have demonstrated that the oxidation state of BFDMA significantly affects the interaction of this lipid with DNA [39, 40] and the efficiency with which lipoplexes of BFDMA and DNA transfect cells [37, 38, 41]. In particular, our past studies have identified a range of lipid concentrations over which lipoplexes formed from reduced BFDMA mediate high levels of transgene expression in vitro , whereas lipoplexes formed from oxidized BFDMA (over the same range of concentrations) yield negligible levels of transgene expression [37, 38, 41].…”

Section: Introductionmentioning

confidence: 99%

“…Our past studies have demonstrated that the oxidation state of BFDMA significantly affects the interaction of this lipid with DNA [39, 40] and the efficiency with which lipoplexes of BFDMA and DNA transfect cells [37, 38, 41]. In particular, our past studies have identified a range of lipid concentrations over which lipoplexes formed from reduced BFDMA mediate high levels of transgene expression in vitro , whereas lipoplexes formed from oxidized BFDMA (over the same range of concentrations) yield negligible levels of transgene expression [37, 38, 41]. Our previous physical characterization experiments also reveal that the oxidation state of BFDMA influences the zeta potentials and nanostructures of lipoplexes formed from BFDMA [39, 40].…”

Section: Introductionmentioning

confidence: 99%

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Addition of ascorbic acid to the extracellular environment activates lipoplexes of a ferrocenyl lipid and promotes cell transfection

Aytar¹,

Müller²,

Golan³

et al. 2012

Journal of Controlled Release

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The level of cell transfection mediated by lipoplexes formed using the ferrocenyl lipid bis(11-ferrocenylundecyl)dimethylammonium bromide (BFDMA) depends strongly on the oxidation state of the two ferrocenyl groups of the lipid (reduced BFDMA generally mediates high levels of transfection, but oxidized BFDMA mediates very low levels of transfection). Here, we report that it is possible to chemically transform inactive lipoplexes (formed using oxidized BFMDA) to “active” lipoplexes that mediate high levels of transfection by treatment with the small-molecule reducing agent ascorbic acid (vitamin C). Our results demonstrate that this transformation can be conducted in cell culture media and in the presence of cells by addition of ascorbic acid to lipoplex-containing media in which cells are growing. Treatment of lipoplexes of oxidized BFDMA with ascorbic acid resulted in lipoplexes composed of reduced BFDMA, as characterized by UV/vis spectrophotometry, and lead to activated lipoplexes that mediated high levels of transgene expression in the COS-7, HEK 293T/17, HeLa, and NIH 3T3 cell lines. Characterization of internalization of DNA by confocal microscopy and measurements of the zeta potentials of lipoplexes suggested that these large differences in cell transfection result from (i) differences in the extents to which these lipoplexes are internalized by cells and (ii) changes in the oxidation state of BFDMA that occur in the extracellular environment (i.e., prior to internalization of lipoplexes by cells). Characterization of lipoplexes by small-angle neutron scattering (SANS) and by cryogenic transmission electron microscopy (cryo-TEM) revealed changes in the nanostructures of lipoplexes upon the addition of ascorbic acid, from aggregates that were generally amorphous, to aggregates with a more extensive multilamellar nanostructure. The results of this study provide guidance for the design of redox-active lipids that could lead to methods that enable spatial and/or temporal control of cell transfection.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Addition of ascorbic acid to the extracellular environment activates lipoplexes of a ferrocenyl lipid and promotes cell transfection

Aytar¹,

Müller²,

Golan³

et al. 2012

Journal of Controlled Release

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“…In a related example, Abbott and coworkers developed ferrocene-containing cationic lipids with reduction/oxidation (redox) sensitive nucleic acid binding capacity. 24 The authors showed that chemical or electrochemical oxidation/reduction of the ferrocene groups could reversibly alter the charge of the lipids and thereby produce shifts between lamellar (reduced) and amorphous (oxidized) lipoplex morphologies, which was directly correlated with deactivation/activation of gene transfer. 25 Subsequent studies demonstrated the utility of these approaches in cell patterning, as the authors spatially controlled gene transfection in COS-7 cultures by incubating the cells with inactivated lipoplexes and selectively delivering ascorbic acid to a subpopulation of cells isolated by a hollow cylinder (among other methods); the ascorbic acid activated lipoplexes in the subpopulation and facilitated efficient gene transfer.…”

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confidence: 99%

Catch and release: photocleavable cationic diblock copolymers as a potential platform for nucleic acid delivery

et al. 2014

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“…A previous study showed that ferrocene-lipid conjugates in the reduced form were taken up better than the oxidised ferrocenium conjugates. [65] With regard to the mechanism of cytotoxicity of the metallocenes that were investigated in this study, two conclusions can be drawn. First, the charge of the metallocene does not play a role; that is, the unspecific interaction of a positively charged cobaltocenium cation with the polyanionic DNA can probably be discounted.…”

Section: Discussionmentioning

confidence: 92%

Enhanced Cellular Uptake and Cytotoxicity Studies of Organometallic Bioconjugates of the NLS Peptide in Hep G2 Cells

et al. 2009

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SPACE INVADERS: Organometallic fragments such as the ferrocenyl group (shown in red in the picture) help to enhance cellular entry of NLS peptides. Eventually, these nontoxic conjugates find their way to the cellular nucleus as shown by fluorescence microscopy studies in this work. Intracellular delivery to biomolecular targets is still a major challenge in molecular and cell biology. We recently found that attaching an organometallic group, namely the cobaltocenium cation, to the SV 40 large T antigen nuclear localisation signal (NLS) greatly enhances cellular uptake of the conjugate (Noor et al., Angew. Chem. Int. Ed. 2005, 45, 2429). In addition, nuclear localisation of the conjugate was observed. In this work, we present a thorough investigation of this novel cellular delivery system with respect to the nature of the metal complex and the peptide sequence. A number of ferrocene ((Fe(II)), neutral metal complex) and cobaltocenium ((Co(III)), cationic metal complex) bioconjugates with both the NLS wild-type sequence PKKKRKV and a scrambled sequence (NLS(scr), KKVKPKR) were prepared by solid-phase peptide synthesis (SPPS). Cellular and nuclear uptake of these bioconjugates was studied by fluorescence microscopy on living Hep G2 cells. In addition, cytotoxicity screening on the conjugates was carried out, as the toxic effects of several simple metallocenes have been noted previously. Rapid cellular uptake as well as nuclear localisation was observed for the metal-NLS conjugates, but not for any dipeptide controls, the metal-NLS(scr) conjugates or any metal-free conjugates. It thus appears that the presence of a metallocene, but not its charge, and the correct NLS sequence is essential for cellular uptake. Fluorescence microscopy co-localisation studies did not reveal a significant endosomal entrapment of the conjugates. The metallocene not only provides a hydrophobic handle for membrane translocation but also facilitates the localisation and distribution of the conjugate in the cytoplasm. The NLS peptide on the other hand is responsible for the nuclear localisation of the bioconjugate. Finally, none of the conjugates were found to be toxic up to the highest concentrations that was tested (1 mM) against the Hep G2 cells that were used in this study. In conclusion, this work supports metallocene-NLS bioconjugates, in particular with the very robust cobaltocenium group, as a simple but potent, nontoxic system for cellular uptake and nuclear delivery. Concurrently, our finding is relevant to the still-unresolved question of cytotoxicity of metallocenes because it excludes binding and/or damage to the DNA as a mechanism of metallocene cytotoxicity. This finding is confirmed by a combined yeast cytotoxicity/genotoxicity assay, which also shows very little toxic effects for all organometal-NLS conjugates that were tested.

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Ferrocene-Containing Cationic Lipids: Influence of Redox State on Cell Transfection

Cited by 63 publications

References 17 publications

Addition of ascorbic acid to the extracellular environment activates lipoplexes of a ferrocenyl lipid and promotes cell transfection

Addition of ascorbic acid to the extracellular environment activates lipoplexes of a ferrocenyl lipid and promotes cell transfection

Catch and release: photocleavable cationic diblock copolymers as a potential platform for nucleic acid delivery

Enhanced Cellular Uptake and Cytotoxicity Studies of Organometallic Bioconjugates of the NLS Peptide in Hep G2 Cells

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