Enhanced nuclear import and transfection efficiency of plasmid DNA using streptavidin-fused importin-β

Nagasaki, Takeshi; Kawazu, Takeshi; Tachibana, Taro; Tamagaki, Seizo; Shinkai, Seiji

doi:10.1016/j.jconrel.2004.11.024

Cited by 29 publications

(21 citation statements)

References 26 publications

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“…Importin-plays an important role in nuclear transport. 35,36 Nagasaki et al 23 were the first to prepare streptavidin-fused importin-and conjugate it with biotin-attached plasmid-DNA. They have reported that the resultant importin-increases the nuclear transport of the plasmid-DNA.…”

Section: Resultsmentioning

confidence: 99%

“…CD, 12 microinjection, 23 and confocal laser-scanning microscopy 24,25 were performed, and those experimental details were described elsewhere. The hydrodynamic radius (R h ) in each SPG and the complex was determined using a Zetasizer Nano-ZS (Malvern Instruments Ltd.) with 100 mM Tris or 100 mM NaCl buffer at 37 C. Microinjection assay was carried out in order to clarify whether the small complex can be transported through nuclear pore.…”

Section: Spgmentioning

confidence: 99%

“…This is consistent with the size selectivity and the cationic nature being favorable for nuclear transport. Nagasaki et al 23 have reported a microinjection study for plasmid DNA/ importin-conjugate, showing that the conjugate is more localized in nucleus and thus has higher expression efficiency than intact plasmid DNA. We can expect the same effect for the importin-conjugated SPG.…”

Section: Spg-am-rgdmentioning

confidence: 99%

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Delivery of Antisense Oligonucleotides to Nuclear Telomere RNA by Use of a Complex between Polysaccharide and Polynucleotide

Minari

Kubo

Ohba

et al. 2007

Bulletin of the Chemical Society of Japan

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Telomerase, which is highly activated in neoplastic cells, can be a target for antisense therapy, and for that purpose, antisense oligonucleotides (AS ODNs) have to be effectively delivered into cellular nucleus where the target telomerase is present. The present work shows a new strategy to deliver AS ODNs to nucleus by use of a novel complex made from a natural polysaccharide schizophyllan (SPG) and AS ODNs. Nuclear transport is strictly regulated by the nuclear pore size and the related proteins. If the molecular weight of SPG is decreased, the SPG/AS ODN complex should be easily transported, although the stability of the complex decreases with a decrease in the molecular weight. We optimized the molecular weight of SPG to be 25 K. Furthermore, we attached importin-(a nuclear transport protein) to the side chain of SPG by use of a streptavidin-biotin interaction. When this complex was added to Jurkat cells, the telomerase activity was more suppressed than the naked dose, indicating that the importin-in the complex induced the nuclear transport of the complexed AS ODN and the AS ODN inhibited the telomerase. The present work shows a new methodology for nuclear anti-sense therapy that should be important in future anti-cancer therapies.Telomere shortening determines cellular lifetime and eventually induces apoptosis; therefore, the activation of telomerase and the resultant extension of telomere are essential for immortalization of neoplastic cells. This fact implies that telomerase can be a target for anti-cancer therapy. Telomerase is a ribonucleoprotein complex containing an RNA subunit that serves as a template for reverse transcription of telomere DNA. Corey et al. 1,2 showed that the oligonucleotides (ODNs) complementary to the template RNA can block the transcription process and therefore suppress neoplastic growth. This event happens only when ODNs are effectively delivered into cellular nucleus, because the target telomerase is present in nucleus. This strategy can be regarded as an antisense therapy, although the target RNA is present in nucleus, while the target RNAs for conventional antisense therapy are present in cytosol. To make this strategy practicable, three issues should be considered: (1) improving instability of AS ODNs in biological fluids; (2) increasing the cellular uptake of AS ODNs; (3) delivering AS ODNs to target organelle.

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Section: Resultsmentioning

confidence: 99%

Section: Spgmentioning

confidence: 99%

Section: Spg-am-rgdmentioning

confidence: 99%

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Delivery of Antisense Oligonucleotides to Nuclear Telomere RNA by Use of a Complex between Polysaccharide and Polynucleotide

Minari

Kubo

Ohba

et al. 2007

Bulletin of the Chemical Society of Japan

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“…Thus, modification of the pDNA with proteins (i.e. importins) via avidin-biotin interactions represents an alternative strategy [55,56]. The other strategy is the compaction of pDNA with NLS-conjugated cationic polymers [57,58] or the cationically charged NLS itself [59][60][61][62][63].…”

Section: Control Of the Intracellular Trafficking For Pdna Deliverymentioning

confidence: 99%

“Programmed packaging” for gene delivery

Hyodo

Sakurai

Akita

et al. 2014

Journal of Controlled Release

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We report on the development of a Multifunctional Envelope-type Nano Device (MEND) based on our packaging concept "Programmed Packaging" to control not only intracellular trafficking but also the biodistribution of encapsulated compounds such as nucleic acids/proteins/peptides. Our strategy for achieving this is based on molecular mechanisms of cell biology such as endocytosis, vesicular trafficking, etc. In this review, we summarize the concept of programmed packaging and discuss some of our recent successful examples of using MENDs. Systematic evolution of ligands by exponential enrichment (SELEX) was applied as a new methodology for identifying a new ligand toward cell or mitochondria. The delivery of siRNA to tumors and the tumor vasculature was achieved using pH sensitive lipid (YSK05), which was newly designed and optimized under in vivo conditions. The efficient delivery of pDNA to immune cells such as dendritic cells has also been developed using the KALA ligand, which can be a breakthrough technology for DNA vaccine. Finally, ss-cleavable and pH-activated lipid-like surfactant (ssPalm) which is a lipid like material with pH-activatable and SS-cleavable properties is also introduced as a proof of our concept.

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“…Hence delivery of therapeutic plasmid DNA to a nonproliferating cell nucleus is an inefficient process. Nuclear import of signal-mediated pDNA using nuclear localization signal (NLS) for non-viral gene delivery has been attempted [57][58][59]. …”

Section: (C) Plasmid Dna Uptake In Nucleusmentioning

confidence: 99%