Design concepts of polyplex micelles for <scp><i>in vivo</i></scp> therapeutic delivery of plasmid DNA and messenger RNA

Uchida, Satoshi; Kataoka, Kazunori

doi:10.1002/jbm.a.36614

Cited by 84 publications

(84 citation statements)

References 170 publications

(360 reference statements)

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“…Overall, cellular uptake of all vectors, pDNA and mRNA, was over 90%; therefore, the bottleneck for a successful pDNA transfection seems to be the nuclear entry, despite our systems contains P, which favors the transcription process and the entry of the pDNA into the nucleus, thanks to their nuclear localization signals [ 26 ]. Unfortunately, an effective nuclear transport system has yet to be stablished [ 27 ]. Among pDNA vectors, the highest transfection efficacy was observed with HA-containing vectors in both cell lines, and in ARPE-19 especially with the vector pDNA-HA-SLN1.…”

Section: Discussionmentioning

confidence: 99%

Nucleic Acid Delivery by Solid Lipid Nanoparticles Containing Switchable Lipids: Plasmid DNA vs. Messenger RNA

et al. 2020

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The development of safe and effective nucleic acid delivery systems remains a challenge, with solid lipid nanoparticle (SLN)-based vectors as one of the most studied systems. In this work, different SLNs were developed, by combination of cationic and ionizable lipids, for delivery of mRNA and pDNA. The influence of formulation factors on transfection efficacy, protein expression and intracellular disposition of the nucleic acid was evaluated in human retinal pigment epithelial cells (ARPE-19) and human embryonic kidney cells (HEK-293). A long-term stability study of the vectors was also performed. The mRNA formulations induced a higher percentage of transfected cells than those containing pDNA, mainly in ARPE-19 cells; however, the pDNA formulations induced a greater protein production per cell in this cell line. Protein production was conditioned by energy-dependent or independent entry mechanisms, depending on the cell line, SLN composition and kind of nucleic acid delivered. Vectors containing 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) as unique cationic lipid showed better stability after seven months, which improved with the addition of a polysaccharide to the vectors. Transfection efficacy and long-term stability of mRNA vectors were more influenced by formulation-related factors than those containing pDNA; in particular, the SLNs containing only DOTAP were the most promising formulations for nucleic acid delivery.

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Section: Discussionmentioning

confidence: 99%

Nucleic Acid Delivery by Solid Lipid Nanoparticles Containing Switchable Lipids: Plasmid DNA vs. Messenger RNA

et al. 2020

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“…An odd-even effect of the amynoethylene repeated units has been described for different nucleic acid payload. PAsp containing even-numbered of amynoethylene units showed higher transfection with pDNA, whereas, an odd-numbered unit sustainably increased mRNA expression and enhanced resistance against [129,130]. PEG-PAsp polyplexes have displayed enhanced stability and lower cytotoxicity; for instance, IVT mRNA complexed with PEG-PAsp(DET) was intranasally administrated to mice for delivering brain-derived neurotrophic factor (BDNF), showing protein expression in nasal tissues for nearly two days [131].…”

Section: Polymeric Systemsmentioning

confidence: 99%

Nanomedicines to Deliver mRNA: State of the Art and Future Perspectives

et al. 2020

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The use of messenger RNA (mRNA) in gene therapy is increasing in recent years, due to its unique features compared to plasmid DNA: Transient expression, no need to enter into the nucleus and no risk of insertional mutagenesis. Nevertheless, the clinical application of mRNA as a therapeutic tool is limited by its instability and ability to activate immune responses; hence, mRNA chemical modifications together with the design of suitable vehicles result essential. This manuscript includes a revision of the strategies employed to enhance in vitro transcribed (IVT) mRNA functionality and efficacy, including the optimization of its stability and translational efficiency, as well as the regulation of its immunostimulatory properties. An overview of the nanosystems designed to protect the mRNA and to overcome the intra and extracellular barriers for successful delivery is also included. Finally, the present and future applications of mRNA nanomedicines for immunization against infectious diseases and cancer, protein replacement, gene editing, and regenerative medicine are highlighted.

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“…Delivery of genes and other functional nucleic acids as a powerful tool for basic research [ 1 – 3 ], in biomedical/therapeutic applications and biotechnology [ 4 – 7 ], has been practiced intensely for decades. Corresponding methods have been constantly advanced [ 8 – 11 ], facilitating efficient cellular delivery of different types of nucleic acids such as plasmid DNA (pDNA), small interfering RNA (siRNA), small hairpin RNA (shRNA), and, more recently, single-guide RNA (sgRNA) as well as in vitro transcribed messenger RNA (IVT-mRNA) [ 9 , 12 – 15 ]. For numerous applications, the simultaneous delivery of more than a single nucleic acid is advantageous or even mandatory.…”

Section: Introductionmentioning

confidence: 99%

Strategies for simultaneous and successive delivery of RNA

2020

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Advanced non-viral gene delivery experiments often require co-delivery of multiple nucleic acids. Therefore, the availability of reliable and robust co-transfection methods and defined selection criteria for their use in, e.g., expression of multimeric proteins or mixed RNA/DNA delivery is of utmost importance. Here, we investigated different co- and successive transfection approaches, with particular focus on in vitro transcribed messenger RNA (IVT-mRNA). Expression levels and patterns of two fluorescent protein reporters were determined, using different IVT-mRNA doses, carriers, and cell types. Quantitative parameters determining the efficiency of co-delivery were analyzed for IVT-mRNAs premixed before nanocarrier formation (integrated co-transfection) and when simultaneously transfecting cells with separately formed nanocarriers (parallel co-transfection), which resulted in a much higher level of expression heterogeneity for the two reporters. Successive delivery of mRNA revealed a lower transfection efficiency in the second transfection round. All these differences proved to be more pronounced for low mRNA doses. Concurrent delivery of siRNA with mRNA also indicated the highest co-transfection efficiency for integrated method. However, the maximum efficacy was shown for successive delivery, due to the kinetically different peak output for the two discretely operating entities. Our findings provide guidance for selection of the co-delivery method best suited to accommodate experimental requirements, highlighting in particular the nucleic acid dose-response dependence on co-delivery on the single-cell level.

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Design concepts of polyplex micelles for in vivo therapeutic delivery of plasmid DNA and messenger RNA

Cited by 84 publications

References 170 publications

Nucleic Acid Delivery by Solid Lipid Nanoparticles Containing Switchable Lipids: Plasmid DNA vs. Messenger RNA

Nucleic Acid Delivery by Solid Lipid Nanoparticles Containing Switchable Lipids: Plasmid DNA vs. Messenger RNA

Nanomedicines to Deliver mRNA: State of the Art and Future Perspectives

Strategies for simultaneous and successive delivery of RNA

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