Discovery of Metabolically Stabilized Electronegative Polyacridine-PEG Peptide DNA Open Polyplexes

Fernandez, Christian A.; Baumhover, Nicholas J.; Anderson, Kevin B.; Rice, Kevin G.

doi:10.1021/bc900514s

Cited by 13 publications

(11 citation statements)

References 39 publications

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“…This may be the result of saturating DNAse in the blood 2 or by blocking uptake of the scavenger receptor as has been reported previously 28 . Interestingly, the DNAse protection afforded to pGL3 by PEGylated polyacridine peptides in blood is not as critical as when dosing polyplexes via local administration, such as intramuscle electroporation 29 .…”

Section: Discussionmentioning

confidence: 99%

High-affinity PEGylated polyacridine peptide polyplexes mediate potent in vivo gene expression

2012

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PEGylated polyacridine peptides bind to plasmid DNA with high affinity to form unique polyplexes that possess a long circulatory half-life and are hydrodynamically (HD)-stimulated to produce efficient gene expression in the liver of mice. We previously demonstrated that (Acr-Lys)6-Cys-PEG5kDa stabilizes a 1 μg pGL3 dose for up to 1 hr in the circulation, resulting in HD-stimulated (saline only) gene expression in the liver, equivalent in magnitude to direct-HD dosing of 1 μg of pGL3 (Fernandez C.A. et al. Gene Therapy 2011). In the present study we report that increasing the spacing of Acr with either 4 or 5 Lys residues, dramatically increases the stability of PEGylated polyacridine peptide polyplexes in the circulation allowing maximal HD-stimulated expression for up to 5 hrs post-DNA administration. Co-administration of a decoy dose of 9 μg of non-expressing DNA polyplex with 1 μg of pGL3 polyplex further extended the HD-stimulated expression to 9 hrs. This structure-activity relationship study defines the PEGylated polyacridine peptide requirements for maintaining fully transfection competent plasmid DNA in the circulation for 5 hrs and provides an understanding as to why polyplexes or lipoplexes prepared with PEI, chitosan or Lipofectamine are inactive within 5 min following i.v. dosing.

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Section: Discussionmentioning

confidence: 99%

High-affinity PEGylated polyacridine peptide polyplexes mediate potent in vivo gene expression

2012

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“…135 As mentioned in the previous section, the coupling of NLSs can be used to facilitate the passage of large molecules across the nuclear envelope to enhance their TE. 121,122,136,137 Another emerging approach is to employ protein transduction domains (PTDs), 138 which contain non-classical NLSs and effectively condense the DNA or RNA using their cationic domains. Commonly used PTDs include TAT (GRKKRRQRRRPQ) peptides derived from transactivator of transcription, antennapedia, and VP22 protein that have been shown to efficiently deliver DNA into the nuclei.…”

Section: Intracellular Targetingmentioning

confidence: 99%

Understanding nano-bio interactions to improve nanocarriers for drug delivery

et al. 2014

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“…Acridines were added either by conjugating acridine to a finished polylysine peptide 195 , or building the peptide using an acridine-lysine amino acid 196 . The latter method was determined to be more effective.…”

Section: Polyacridine Peptidesmentioning

confidence: 99%

Nonviral gene delivery to the liver

Crowley¹

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Diseases of the liver have a large impact on human health. Genetic disorders, metabolic disorders, alcoholism, cancer, or infections can all impair liver function. If serious enough, a liver transplant may be necessary, a major surgical procedure which requires lifelong immune suppression and relies on the availability of donor livers. Gene therapy is being intensively studied as a potential method to treat many disorders, including disorders of the liver. While viral gene therapy has seen some success, possible side effects make it risky, so nonviral gene delivery vectors are being developed. Unfortunately, these nonviral vectors do not yet have the efficiency of the viral vectors. Nonviral gene delivery vectors face many challenges in vivo. The vectors must protect DNA from nucleases while it moves through the bloodstream, they must avoid nonspecific uptake, they must be enter the correct cells, and must enter the nucleus before the DNA can be expressed. If any step of this process fails, there will be very little, if any, expression, and it may be impossible to determine what went wrong. One impediment to nonviral gene delivery research is the transition from in vitro studies to in vivo studies. The cancer derived cell lines most often used for in vitro transfections are rapidly dividing, which makes nuclear entry much easier than in the whole animal. While primary cells would be a more accurate model of the in vivo environment, the number of cells that can be obtained from tissues is small, and primary cells usually cannot be cultured for long. This limits the number of experiments that can be done with each preparation of cells. To overcome this, we have miniaturized transfection assays, including the transfection of mouse primary hepatocytes with luciferase in 384 well plates. Because fewer cells are needed, more experiments can be performed with each liver preparation. Another issue introduced by the differences between in vitro and in vivo research is circulatory stability. In vitro, large particles with strong positive charges are desired, because they sink down onto the cells and are attracted to the negatively charged cellular membranes. However, in vivo these particles will aggregate serum proteins and become lodged in narrow capillary beds in the lungs or other organs, often causing toxicity. While this behavior can usually be overcome through PEGylation, improving a particle's circulatory half-life will still improve its chances of finding the correct target. Scavenger receptors found on liver nonparenchymal cells are very efficient at removing negatively charged particles from the bloodstream. We have shown that dosing large amounts of PEGylated polyacridine DNA polyplex can saturate the scavenger receptors and improve circulatory half-life. We have also shown that large doses of PEGylated peptide, with or without acridine groups, can inhibit scavenger receptor uptake through the formation of peptide-protein nanoparticles. By inhibiting scavenger receptor uptake, DNA can be successfully hydrodynami...

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Discovery of Metabolically Stabilized Electronegative Polyacridine-PEG Peptide DNA Open Polyplexes

Cited by 13 publications

References 39 publications

High-affinity PEGylated polyacridine peptide polyplexes mediate potent in vivo gene expression

High-affinity PEGylated polyacridine peptide polyplexes mediate potent in vivo gene expression

Understanding nano-bio interactions to improve nanocarriers for drug delivery

Nonviral gene delivery to the liver

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