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Solid formulation of cell-penetrating peptide nanoparticles with siRNA and their stability in simulated gastric conditions
Stockholm University, Faculty of Science, Department of Neurochemistry.
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2012 (English)In: Journal of Controlled Release, ISSN 0168-3659, E-ISSN 1873-4995, Vol. 162, no 1, 1-8 p.Article in journal (Refereed) Published
Abstract [en]

Cell-penetrating peptides (CPPs) are short cationic peptides that have been extensively studied as drug delivery vehicles for proteins, nucleic acids and nanoparticles. However, the formulation of CPP-based therapeutics into different pharmaceutical formulations and their stability in relevant biological environments have not been given the same attention. Here, we show that a newly developed CPP, PepFect 14 (PF14), forms non-covalent nanocomplexes with short interfering RNA (siRNA), which are able to elicit efficient RNA-interference (RNAi) response in different cell-lines. RNAi effect was obtained at low siRNA doses with a unique kinetic profile. Furthermore, we utilized the solid dispersion technique to formulate PF14/siRNA nanocomplexes into solid formulations that were as active as the freshly prepared nanocomplexes in solution. Importantly, the freshly prepared nanocomplexes and solid formulations were stable after incubation with simulated gastric fluid having a pH of 1.2 and containing proteolytic enzymes. These results demonstrate the activity of PF14 in delivering and protecting siRNA in different pharmaceutical forms and biological environments.

Place, publisher, year, edition, pages
2012. Vol. 162, no 1, 1-8 p.
Keyword [en]
Cell penetrating peptide, siRNA, Solid formulation, Acid stability, Gastric fluid
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
URN: urn:nbn:se:su:diva-75482DOI: 10.1016/j.jconrel.2012.06.006ISI: 000307769200001OAI: oai:DiVA.org:su-75482DiVA: diva2:516797
Available from: 2012-04-19 Created: 2012-04-19 Last updated: 2017-12-07Bibliographically approved
In thesis
1. Cell-penetrating peptides; chemical modification, mechanism of uptake and formulation development
Open this publication in new window or tab >>Cell-penetrating peptides; chemical modification, mechanism of uptake and formulation development
2012 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Gene therapy holds the promise of revolutionizing the way we treat diseases. By using recombinant DNA and oligonucleotides (ONs), gene functions can be restored, altered or silenced according to the therapeutic need. However, gene therapy approaches require the delivery of large and charged nucleic acid-based molecules to their intracellular targets across the plasma membrane, which is inherently impermeable to such molecules. In this thesis, two chemically modified cell-penetrating peptides (CPPs) that have superior delivery properties for several nucleic acid-based therapeutics are developed. These CPPs can spontaneously form nanoparticles upon non-covalent complexation with the nucleic acid cargo, and the formed nanoparticles mediate efficient cellular transfection. In paper I, we show that an N-terminally stearic acid-modified version of transportan-10 (PF3) can efficiently transfect different cell types with plasmid DNA and mediates efficient gene delivery in-vivo when administrated intra muscularly (i.m.) or intradermaly (i.d.). In paper II, a new peptide with ornithine modification, PF14, is shown to efficiently deliver splice-switching oligonucleotides (SSOs) in different cell models including mdx mouse myotubes; a cell culture model of Duchenne’s muscular dystrophy (DMD). Additionally, we describe a method for incorporating the PF14-SSO nanoparticles into a solid formulation that is active and stable even when stored at elevated temperatures for several weeks. In paper III, we demonstrate the involvement of class-A scavenger receptor subtypes (SCARA3 & SCARA5) in the uptake of PF14-SSO nanoparticles, which possess negative surface charge, and suggest for the first time that some CPP-based systems function through scavenger receptors. In paper IV, the ability of PF14 to deliver siRNA to different cell lines is shown and their stability in simulated gastric acidic conditions is highlighted.

Taken together, these results demonstrate that certain chemical modifications can drastically enhance the activity and stability of CPPs for delivering nucleic acids after spontaneous nanoparticle formation upon non-covalent complexation. Moreover, we show that CPP-based nanoparticles can be formulated into convenient and stable solid formulations that can be suitable for several therapeutic applications. Importantly, the involvement of scavenger receptors in the uptake of such nanoparticles is presented, which could yield novel possibilities to understand and improve the transfection by CPPs and other gene therapy nanoparticles.

Place, publisher, year, edition, pages
Stockholm: Department of Neurochemistry, Stockholm University Stockholm University, 2012. 86 p.
Keyword
cell penetrating peptides, gene therapy, scavenger receptors, pharmaceutical formulation, solid dispersion
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Research subject
Neurochemistry with Molecular Neurobiology
Identifiers
urn:nbn:se:su:diva-75537 (URN)978-91-7447-464-0 (ISBN)
Public defence
2012-06-11, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
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Note
At the time of doctoral defence the following paper was unpublished and had a status as follows: Paper nr 4: SubmittedAvailable from: 2012-05-10 Created: 2012-04-20 Last updated: 2012-04-24Bibliographically approved

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Ezzat, KariemLangel, Ülo
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