Cationic Polymer Intercalation into the Lipid Membrane Enables Intact Polyplex DNA Escape from Endosomes for Gene Delivery

Sriram Vaidyanathan, Junjie Chen, Bradford G. Orr, Mark M. Banaszak Holl

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Developing improved cationic polymer-DNA polyplexes for gene delivery requires improved understanding of DNA transport from endosomes into the nucleus. Using a FRET-capable oligonucleotide molecular beacon (OMB), we monitored the transport of intact DNA to cell organelles. We observed that for effective (jetPEI) and ineffective (G5 PAMAM) vectors, the fraction of cells displaying intact OMB in the cytosol (jetPEI ≫ G5 PAMAM) quantitatively predicted the fraction expressing transgene (jetPEI ≫ G5 PAMAM). Intact OMB delivered with PAMAM and confined to endosomes could be released to the cytosol by the subsequent addition of L-PEI, with a corresponding 10-fold increase in transgene expression. These results suggest that future vector development should optimize vectors for intercalation into, and destabilization of, the endosomal membrane. Finally, the study highlights a two-step strategy in which the pDNA is loaded in cells using one vector and endosomal release is mediated by a second agent.

Original languageEnglish
Pages (from-to)1967-1978
Number of pages12
JournalMolecular Pharmaceutics
Volume13
Issue number6
DOIs
Publication statusPublished - 6 Jun 2016
Externally publishedYes

Keywords

  • FRET molecular beacon
  • gene delivery
  • gene therapy
  • linear poly(ethylenimine)
  • membrane intercalation
  • molecular beacon
  • polyplex-cell membrane interactions
  • polyplexes

Cite this

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title = "Cationic Polymer Intercalation into the Lipid Membrane Enables Intact Polyplex DNA Escape from Endosomes for Gene Delivery",
abstract = "Developing improved cationic polymer-DNA polyplexes for gene delivery requires improved understanding of DNA transport from endosomes into the nucleus. Using a FRET-capable oligonucleotide molecular beacon (OMB), we monitored the transport of intact DNA to cell organelles. We observed that for effective (jetPEI) and ineffective (G5 PAMAM) vectors, the fraction of cells displaying intact OMB in the cytosol (jetPEI ≫ G5 PAMAM) quantitatively predicted the fraction expressing transgene (jetPEI ≫ G5 PAMAM). Intact OMB delivered with PAMAM and confined to endosomes could be released to the cytosol by the subsequent addition of L-PEI, with a corresponding 10-fold increase in transgene expression. These results suggest that future vector development should optimize vectors for intercalation into, and destabilization of, the endosomal membrane. Finally, the study highlights a two-step strategy in which the pDNA is loaded in cells using one vector and endosomal release is mediated by a second agent.",
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Cationic Polymer Intercalation into the Lipid Membrane Enables Intact Polyplex DNA Escape from Endosomes for Gene Delivery. / Vaidyanathan, Sriram; Chen, Junjie; Orr, Bradford G.; Banaszak Holl, Mark M.

In: Molecular Pharmaceutics, Vol. 13, No. 6, 06.06.2016, p. 1967-1978.

Research output: Contribution to journalArticleResearchpeer-review

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AB - Developing improved cationic polymer-DNA polyplexes for gene delivery requires improved understanding of DNA transport from endosomes into the nucleus. Using a FRET-capable oligonucleotide molecular beacon (OMB), we monitored the transport of intact DNA to cell organelles. We observed that for effective (jetPEI) and ineffective (G5 PAMAM) vectors, the fraction of cells displaying intact OMB in the cytosol (jetPEI ≫ G5 PAMAM) quantitatively predicted the fraction expressing transgene (jetPEI ≫ G5 PAMAM). Intact OMB delivered with PAMAM and confined to endosomes could be released to the cytosol by the subsequent addition of L-PEI, with a corresponding 10-fold increase in transgene expression. These results suggest that future vector development should optimize vectors for intercalation into, and destabilization of, the endosomal membrane. Finally, the study highlights a two-step strategy in which the pDNA is loaded in cells using one vector and endosomal release is mediated by a second agent.

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