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Investigation of enzyme-sensitive lipid nanoparticles for delivery of siRNA to blood-brain barrier and glioma cells.

Bruun J, Larsen TB, Jølck RI, Eliasen R, Holm R, Gjetting T, Andresen TL - Int J Nanomedicine (2015)

Bottom Line: Second, the positively charged LNPs are masked with a negatively charged PEGylated (poly(ethylene glycol)) cleavable lipopeptide, which contains a recognition sequence for matrix metalloproteinases (MMPs), a class of enzymes often expressed in the tumor microenvironment and inflammatory BBB conditions.This work describes the development of this two-stage nanocarrier-system and evaluates the performance in brain endothelial and glioblastoma cells with respect to uptake and gene silencing efficiency.The ability of activation by MMP-triggered dePEGylation and charge shift is demonstrated to substantially increase the uptake and the silencing efficiency of the LNPs.

View Article: PubMed Central - PubMed

Affiliation: Department of Micro- and Nanotechnology, Center for Nanomedicine and Theranostics, Technical University of Denmark, DTU Nanotech, Lyngby, Denmark.

ABSTRACT
Clinical applications of siRNA for treating disorders in the central nervous system require development of systemic stable, safe, and effective delivery vehicles that are able to cross the impermeable blood-brain barrier (BBB). Engineering nanocarriers with low cellular interaction during systemic circulation, but with high uptake in targeted cells, is a great challenge and is further complicated by the BBB. As a first step in obtaining such a delivery system, this study aims at designing a lipid nanoparticle (LNP) able to efficiently encapsulate siRNA by a combination of titratable cationic lipids. The targeted delivery is obtained through the design of a two-stage system where the first step is conjugation of angiopep to the surface of the LNP for targeting the low-density lipoprotein receptor-related protein-1 expressed on the BBB. Second, the positively charged LNPs are masked with a negatively charged PEGylated (poly(ethylene glycol)) cleavable lipopeptide, which contains a recognition sequence for matrix metalloproteinases (MMPs), a class of enzymes often expressed in the tumor microenvironment and inflammatory BBB conditions. Proteolytic cleavage induces PEG release, including the release of four glutamic acid residues, providing a charge switch that triggers a shift of the LNP charge from weakly negative to positive, thus favoring cellular endocytosis and release of siRNA for high silencing efficiency. This work describes the development of this two-stage nanocarrier-system and evaluates the performance in brain endothelial and glioblastoma cells with respect to uptake and gene silencing efficiency. The ability of activation by MMP-triggered dePEGylation and charge shift is demonstrated to substantially increase the uptake and the silencing efficiency of the LNPs.

No MeSH data available.


Related in: MedlinePlus

Schematic presentation showing the gene delivery by dual modified LNPs.Notes: The nanoparticle is modified with angiopep for receptor-mediated uptake in LRP-1 expressing cells (left pathway) and a MMP-cleavable lipopeptide for activation in tumor tissue microenvironment (right pathway). Intra- or extracellular cleavage of the lipopeptide dePEGylates the LNP and reverses the surface charge from negative to positive leading to increased uptake and endosomal escape.Abbreviations: LNPs, lipid nanoparticles; LRP-1, low-density lipoprotein receptor-related protein-1; PEG, poly(ethylene glycol).
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f1-ijn-10-5995: Schematic presentation showing the gene delivery by dual modified LNPs.Notes: The nanoparticle is modified with angiopep for receptor-mediated uptake in LRP-1 expressing cells (left pathway) and a MMP-cleavable lipopeptide for activation in tumor tissue microenvironment (right pathway). Intra- or extracellular cleavage of the lipopeptide dePEGylates the LNP and reverses the surface charge from negative to positive leading to increased uptake and endosomal escape.Abbreviations: LNPs, lipid nanoparticles; LRP-1, low-density lipoprotein receptor-related protein-1; PEG, poly(ethylene glycol).

Mentions: Here we present PCL-decorated LNPs as a two-stage vehicle for delivery of siRNA to brain tumors. For receptor-mediated transport across the BBB, the MMP-sensitive LNP vehicle is functionalized with angiopep that provides dual targeting to both brain endothelial and glioma cells (Figure 1, left panel). Once at the tumor site, the receptor-mediated uptake is supported by MMP-triggered proteolytical activation of the LNP, which results in removal of the protective PEG coating and negative charge, thus favoring cellular endocytosis and release of siRNA (Figure 1, right panel). We investigate this double-functionalized siRNA delivery vehicle and evaluate the influence of the lipid anchor for its potential to effectively target and mediate protein knockdown in brain endothelial and glioblastoma cell cultures.


Investigation of enzyme-sensitive lipid nanoparticles for delivery of siRNA to blood-brain barrier and glioma cells.

Bruun J, Larsen TB, Jølck RI, Eliasen R, Holm R, Gjetting T, Andresen TL - Int J Nanomedicine (2015)

Schematic presentation showing the gene delivery by dual modified LNPs.Notes: The nanoparticle is modified with angiopep for receptor-mediated uptake in LRP-1 expressing cells (left pathway) and a MMP-cleavable lipopeptide for activation in tumor tissue microenvironment (right pathway). Intra- or extracellular cleavage of the lipopeptide dePEGylates the LNP and reverses the surface charge from negative to positive leading to increased uptake and endosomal escape.Abbreviations: LNPs, lipid nanoparticles; LRP-1, low-density lipoprotein receptor-related protein-1; PEG, poly(ethylene glycol).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4590347&req=5

f1-ijn-10-5995: Schematic presentation showing the gene delivery by dual modified LNPs.Notes: The nanoparticle is modified with angiopep for receptor-mediated uptake in LRP-1 expressing cells (left pathway) and a MMP-cleavable lipopeptide for activation in tumor tissue microenvironment (right pathway). Intra- or extracellular cleavage of the lipopeptide dePEGylates the LNP and reverses the surface charge from negative to positive leading to increased uptake and endosomal escape.Abbreviations: LNPs, lipid nanoparticles; LRP-1, low-density lipoprotein receptor-related protein-1; PEG, poly(ethylene glycol).
Mentions: Here we present PCL-decorated LNPs as a two-stage vehicle for delivery of siRNA to brain tumors. For receptor-mediated transport across the BBB, the MMP-sensitive LNP vehicle is functionalized with angiopep that provides dual targeting to both brain endothelial and glioma cells (Figure 1, left panel). Once at the tumor site, the receptor-mediated uptake is supported by MMP-triggered proteolytical activation of the LNP, which results in removal of the protective PEG coating and negative charge, thus favoring cellular endocytosis and release of siRNA (Figure 1, right panel). We investigate this double-functionalized siRNA delivery vehicle and evaluate the influence of the lipid anchor for its potential to effectively target and mediate protein knockdown in brain endothelial and glioblastoma cell cultures.

Bottom Line: Second, the positively charged LNPs are masked with a negatively charged PEGylated (poly(ethylene glycol)) cleavable lipopeptide, which contains a recognition sequence for matrix metalloproteinases (MMPs), a class of enzymes often expressed in the tumor microenvironment and inflammatory BBB conditions.This work describes the development of this two-stage nanocarrier-system and evaluates the performance in brain endothelial and glioblastoma cells with respect to uptake and gene silencing efficiency.The ability of activation by MMP-triggered dePEGylation and charge shift is demonstrated to substantially increase the uptake and the silencing efficiency of the LNPs.

View Article: PubMed Central - PubMed

Affiliation: Department of Micro- and Nanotechnology, Center for Nanomedicine and Theranostics, Technical University of Denmark, DTU Nanotech, Lyngby, Denmark.

ABSTRACT
Clinical applications of siRNA for treating disorders in the central nervous system require development of systemic stable, safe, and effective delivery vehicles that are able to cross the impermeable blood-brain barrier (BBB). Engineering nanocarriers with low cellular interaction during systemic circulation, but with high uptake in targeted cells, is a great challenge and is further complicated by the BBB. As a first step in obtaining such a delivery system, this study aims at designing a lipid nanoparticle (LNP) able to efficiently encapsulate siRNA by a combination of titratable cationic lipids. The targeted delivery is obtained through the design of a two-stage system where the first step is conjugation of angiopep to the surface of the LNP for targeting the low-density lipoprotein receptor-related protein-1 expressed on the BBB. Second, the positively charged LNPs are masked with a negatively charged PEGylated (poly(ethylene glycol)) cleavable lipopeptide, which contains a recognition sequence for matrix metalloproteinases (MMPs), a class of enzymes often expressed in the tumor microenvironment and inflammatory BBB conditions. Proteolytic cleavage induces PEG release, including the release of four glutamic acid residues, providing a charge switch that triggers a shift of the LNP charge from weakly negative to positive, thus favoring cellular endocytosis and release of siRNA for high silencing efficiency. This work describes the development of this two-stage nanocarrier-system and evaluates the performance in brain endothelial and glioblastoma cells with respect to uptake and gene silencing efficiency. The ability of activation by MMP-triggered dePEGylation and charge shift is demonstrated to substantially increase the uptake and the silencing efficiency of the LNPs.

No MeSH data available.


Related in: MedlinePlus